method of conducting quality control to determine abnormalities in the operation of a microfluidic s

专利摘要:
feedback control in microfluidic systems. the present invention relates to systems and methods for controlling fluids in microfluidic systems. in some modalities, fluid control involves the use of feedback from one or more processes or events that occur in the microfluidic system. for example, a detector can detect one or more fluids in a measurement zone of a microfluidic system and one or more signals, or a pattern of signals, that correspond to the fluid can be generated. in some cases, the signal or signal pattern may correspond to an intensity, a duration, a position in time in relation to a second position in time or in relation to another process, and / or a period of time between events. using this data, a control system can determine whether the subsequent fluid flow in the microfluidic system should be modulated. in some modalities, these and other methods can be used to perform quality control to determine abnormalities in the operation of the microfluidic system.
公开号:BR112012026406B1
申请号:R112012026406
申请日:2011-04-15
公开日:2020-06-09
发明作者:Steinmiller David；Linder Vincent
申请人:Opko Diagnostics Llc；
IPC主号:

专利说明:

Descriptive Report of the Invention Patent for METHOD OF CONDUCTING QUALITY CONTROL TO DETERMINE ABNORMALITIES IN THE OPERATION OF A MICROFLUID SYSTEM.
FIELD
[001] Systems and methods for controlling fluids in microfluidic systems are described in general. In some modalities, fluid control involves using feedback from one or more processes or events that occur in the microfluidic system. BACKGROUND
[002] Fluid handling plays an important role in fields such as chemistry, microbiology and biochemistry. Such fluids can include liquids or gases and can provide reagents, solvents, reactive agents, or rinses to chemical or biological processes. Although various microfluidic methods and devices, such as microfluidic assays, can provide economical, sensitive and accurate analytical platforms, fluid manipulations - such as mixing multiple fluids, introducing samples, introducing reagents, storing reagents, , fluid separation, waste collection, fluid extraction for off-chip analysis, and / or transferring fluids from one chip to the next - can add a level of cost and sophistication. Therefore, advances in the field that could reduce costs, simplify use, provide quality control for the analysis being performed, and / or improve fluid handling in microfluidic systems should be beneficial.
SUMMARY
[003] Systems and methods for controlling fluids in microfluidic systems are described in general. In some modalities, fluid control involves the use of feedback from one or more
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2/101 processes or events that occur in the microfluidic system. The objective of the present invention involves, in some cases, related products, alternative solutions to a particular problem, and / or a plurality of different uses of one or more systems and / or articles.
[004] In a set of modalities, a series of methods is provided. In one embodiment, a method comprises the initiation of fluid detection in a first measurement zone of a microfluidic system. The method involves detecting a first fluid and a second fluid in the first measurement zone and the formation of a first signal that corresponds to the first fluid and a second signal that corresponds to the second fluid. A first signal pattern is transmitted to a control system, wherein the first signal pattern comprises at least two of: a) an intensity of the first signal;
b) a duration of the first signal; c) a position of the first signal in time in relation to a second position in time; and d) an average period of time between the first and second signals. The method also involves determining whether the flow of fluid in the microfluidic system should be modulated based at least in part on the first signal pattern.
[005] In another embodiment, a method comprises the detection of a first fluid and a second fluid in a first measurement zone of a microfluidic system, wherein the detection step comprises the detection of at least two a) opacity the first fluid; b) a volume of the first fluid; c) a flow of the first fluid; d) a position of detecting the first fluid in time in relation to a second position in time; and e) an average period of time between the detection of the first and second fluids. The method involves determining whether the flow of fluid in the microfluidic system should be modulated based at least in part on the detection step.
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[006] In another embodiment, a method of carrying out quality control to determine abnormalities in the operation of a microfluidic system comprises the detection of a first fluid in a first measurement zone of the microfluidic system and the formation of a first signal that corresponds to the first fluid. The method also involves transmitting the first signal to a control system, comparing the first signal to a reference signal, thereby determining the presence of abnormalities in the operation of the microfluidic system, and the determination must be stopped. made in the microfluidic system based at least in part on the results of the comparison step.
[007] Other advantages and new features of the present invention will become apparent from the detailed description below of various non-limiting modalities of the invention when considered in conjunction with the attached figures.
BRIEF DESCRIPTION OF THE DRAWINGS
[008] The non-limiting modalities of the present invention will be described by way of example with reference to the attached figures, which are schematic and should not be drawn to scale. In the figures, each identical or nearly identical component illustrated is typically represented by a single numeral. For the sake of clarity, not every component is labeled in each figure, nor is each component of each embodiment of the invention shown where the illustration is not necessary to allow elements versed in the state of the art to understand the invention. In the figures:
[009] Figure 1 is a block diagram showing a microfluidic system and a variety of components that can be part of a sample analyzer according to a modality;
[0010] Figure 2 is a graph showing the measurement of optical density as a function of time according to a modality;
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[0011] Figure 3 is a perspective view of a cassette that includes a fluidic connector according to an embodiment;
[0012] Figure 4 is an exploded view of a cassette according to an embodiment;
[0013] Figure 5 is a schematic view of a cassette according to an embodiment;
[0014] Figure 6 is a diagram showing a microfluidic system of a cassette that includes a fluidic connector according to a modality;
[0015] Figure 7 is a schematic view of a part of a sample analyzer according to an embodiment;
[0016] Figure 8 is a block diagram showing a control system of a sample analyzer associated with a variety of different components according to a modality;
[0017] Figure 9 is a schematic diagram showing a microfluidic system of a cassette according to a modality; and [0018] Figure 10 is a graph showing the measurement of optical density as a function of time according to a modality. DETAILED DESCRIPTION
[0019] The systems and methods for controlling fluids in microfluidic systems are described in general. In some modalities, fluid control involves using feedback from one or more processes or events that occur in the microfluidic system. For example, a detector can detect one or more fluids that pass through a measurement zone of a microfluidic system and one or more signals, or a pattern of signals, that correspond to the fluid can be generated. In some cases, the signal or signal pattern may correspond to an intensity (for example, an indication of the type of fluid passing through the detector), a duration (for example, an indication of fluid volume and / or flow ), a po
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5/101 timing in relation to another position in time or in relation to another process that occurred in the microfluidic system (for example, when a certain fluid passed through the detector after a valve was activated), and / or a average time period between events (for example, between two consecutive signals). Using this data, a control system can determine whether the subsequent fluid flow in the microfluidic system should be modulated. In some modalities, these and other methods can be used to perform quality control to determine abnormalities in the operation of the microfluidic system.
[0020] As described in more detail below, in some modalities an analysis done on a device can be recorded to essentially produce a fingerprint of the analysis, and all or parts of the fingerprint can be used to provide feedback to the microfluidic system. For example, an analysis fingerprint can include signals from each fluid (for example, passing across, through, above, below, etc.) on one detector or multiple detectors, which can be statically positioned in a measurement zone or in multiple measurement zones of a device. The signals can be a measurement, for example, of the transmission of light passing through fluids. Since the different fluids used in the analysis can have different volumes, flow rates, compositions, and other characteristics, the fluids can produce signals that have different intensities and durations, which are reflected in the fingerprint. In this way, the fingerprint can be used to identify, for example, the fluids used in the analysis, the timing of the fluids (for example, when particular fluids were introduced in certain regions of the device), and the interaction between the fluids (for example , mixing). This data can be used to provide feedback to modulate the subsequent fluid flow
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6/101 hot in the microfluidic system, and in some cases, to perform quality control to determine whether all or parts of the analysis were performed correctly.
[0021] The systems and methods described here can find application in a variety of fields. In some cases, systems and methods can be used to perform quality control to determine, for example, a correct sequence of events that occur in the microfluidic system. If an incorrect sequence of events is determined, the feedback control can, for example, cancel the test being performed in the microfluidic system and / or alert the user to the abnormality. In addition and / or alternatively, the systems and methods described herein can be used to modulate fluid flow, such as mixing, introducing or removing fluids in certain channels or reservoirs in the microfluidic system, activating one or more components such as a valve, pump, vacuum, or heater, and other processes. These and other processes can be applied to a variety of microfluidic systems such as, for example, microfluidic point of care diagnostics platforms, microfluidic laboratory chemical analysis systems, high throughput detection systems, fluidic control systems in crop cultures. cells or bioreactors, among others. The articles, systems, and methods described herein can be particularly useful, in some cases, where a cheap, robust, disposable microfluidic device is desired.
[0022] In addition, the feedback control described here can be used to perform any appropriate process in a microfluidic system, such as a chemical and / or biological reaction. As a specific example, feedback control can be used to control reagent transport in antibody assays that employ unstable reaction precursors, such as the solution assay
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7/101 silver described in the examples section. Other advantages are described in more detail below.
[0023] A number of exemplifying systems and methods are now described.
[0024] Figure 1 shows a block diagram 10 of a microfluidic system and several components that can provide feedback control according to a set of modalities. The microfluidic system can include, for example, a device 20 operatively associated with one or more components such as a fluid flow source 40 such as a pump (for example, to introduce one or more fluids into the device and / or to control fluid flows), optionally a fluid flow source 40 such as a pump or vacuum that can be configured to apply either positive pressure or vacuum (for example, to move / remove one or more fluids from within / from the cassette and / or to control fluid flows), a valve system 28 (for example, to drive one or more valves), a detection system 34 (for example to detect one or more fluids and / or processes) , and / or a temperature regulating system 41 (for example, to heat and / or cool one or more regions of the device). The components can be external or internal to the microfluidic device, and can optionally include one or more processors to control the component or the component system. In certain embodiments, one or more of such components and / or processors are associated with a sample analyzer 47 configured to process and / or analyze a sample contained in the microfluidic system.
[0025] In general, as used here, a component that is operatively associated with one or more other components indicates that such components are directly connected to each other, in direct physical contact with each other without being connected
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8/101 or fixed to each other, or are not directly connected to each other or in contact with each other, but are mechanically interconnected, electrically (including through electromagnetic signals transmitted through space), or fluidly interconnected (for example , through channels such as pipes) in order to cause or allow the associated components in this way to perform their intended functionality.
[0026] The components shown illustratively in the figure
1, as well as other optional components, can be operatively associated with a control system 50. In some embodiments, the control system can be used to control liquids and / or regulate quality control by using feedback from one or more more events that occur in the microfluidic system. For example, the control system can be configured to receive input signals from one or more components, calculate and / or control various parameters, compare one or more signals or a signal pattern with the pre-programmed signals or values in the control system. control, and / or send signals to one or more components to modulate the fluid flow / control operation of the microfluidic system. Specific examples of feedback control are provided below.
[0027] The control system can also be optionally associated with other components such as an interface 54, an identification system 56, an external communication unit 58 (for example, a USB), and / or other user components, such as as described in more detail below.
[0028] The microfluidic device, for example, 20, can have any appropriate configuration of channels and / or components to perform a desired analysis. In a set of modalities, the microfluidic device 20 contains stored reagents that can be used to perform a chemical and / or biological reaction (for example,
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9/101 example, an immunoassay). The microfluidic device can include, for example, an optional reagent inlet 62 in fluid communication with an optional reagent storage area 64. The storage area can include, for example, one or more channels and / or reservoirs that can, in some embodiments, be partially or completely filled with fluids (for example, liquids and gases, including immiscible reagents such as reagent solutions and washing solutions, optionally separated by immiscible liquids, as described in more detail below). The microfluidic device may also include an optional sample or reagent loading area 66, such as a fluid connector that can be used to connect reagent storage area 64 to an optional measurement zone 68 (for example, a reaction area ). The measurement zone, which can include one or more zones (for example, detection regions) to detect a component in a sample, can be in fluid communication with an optional disposal area 70 and coupled to outlet 72. In a set of modalities, the fluid can flow in the direction of the arrows shown in the figure. A description and additional examples of these and other components are provided in more detail below.
[0029] In some embodiments, sections 71 and 77 of the microfluidic device are not in fluid communication with each other before introducing a sample into the microfluidic device. In some cases, sections 71 and 77 are not in fluid communication with each other prior to the first use of the microfluidic device, where, in the first use, the sections are placed in fluid communication with each other. In other embodiments, however, sections 71 and 77 are in fluid communication with each other before first use and / or before introducing a sample into the microfluidic device. Other configurations of microfluidic devices are also posPetition 870200007448, of 1/16/2020, p. 12/164
10/101 levels.
[0030] As shown in the exemplary embodiment illustrated in figure 1, one or more sources of fluid flow 40, such as a pump and / or a vacuum or another pressure control system, valve system 28, detection system 34, temperature control system 41, and / or other components can be operatively associated with one or more of the reagent inlet 62, the reagent2 storage area 64, the sample or reagent loading area 66, the zone measurement 68, the discharge area 70, the outlet 72, and / or other regions of the microfluidic device 20. The detection of processes or events in one or more regions of the microfluidic device can produce a signal or a pattern of signals that can be transmitted to the control system 50. Based at least in part on the signal (s) received by the control system, this feedback can be used to manipulate fluids within and / or between each of these regions of the microfluidic device, t al how to control one or more of a pump, a vacuum, a valve system, a detection system, a temperature regulating system, and / or other components. In some cases, feedback can determine abnormalities that have occurred in the microfluidic system, and the control system can send a signal to one or more components to cause all or parts of the system to be stopped. Consequently, the quality of the processes being performed in the microfluidic system can be controlled by using the systems and methods described here.
[0031] In some modalities, feedback control involves the detection of one or more events or processes that occur in a microfluidic system. A variety of detection methods can be used, as described in more detail below. Detection may involve, for example, the determination of at least one ca
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11/101 characteristic of a fluid, a component within a fluid, the interaction between components within the regions of the microfluidic device, or a condition within a region of the microfluidic device (for example, temperature, pressure, humidity). For example, detection may involve the detection of an opacity of one or more fluids, a concentration of one or more components in a fluid, a volume of one or more fluids, a flow rate of one or more fluids, a position of detection of a first fluid in time in relation to a second position in time, and an average period of time between the detection of a first fluid and a second fluid. The detection of one or more characteristics, conditions, or events can, in some modalities, result in the generation of one or more signals, which can optionally be further processed and transmitted to the control system. As described in more detail here, one or more signals can be compared to one or more signals, values or pre-programmed limits in the control system, and can be used to provide feedback to the microfluidic system.
[0032] A variety of signals or signal patterns can be generated and / or determined (eg, measured) by using the systems and methods described here. In a set of modalities, a signal includes an intensity component. The intensity can indicate or be used to indicate, for example, one or more of: the concentration of a component in a fluid, an indication of the type of fluid being detected (for example, a type of sample such as blood versus urine, or a physical characteristic of the fluid such as a liquid versus a gas), the amount of a component in a fluid, and the volume of a fluid. In some cases, the intensity is determined by the opacity of a fluid or component. In other embodiments, the intensity is determined by the use of a marker or label such as a fluorescent marker or label.
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[0033] In some modalities, a frequency of the signals can be generated and / or determined. For example, a series of signals in which each has an intensity (for example, above or below a threshold intensity) can be measured by a detector. This number can be compared with a number of signals or values (with the intensity above or below the limit intensity) pre-programmed in a control system or other unit. Based at least in part on this comparison, the control system can start, stop, or change a condition such as the modulation of fluid flow in the microfluidic system.
[0034] In some modalities, a signal duration is generated and / or determined. The duration of a signal can indicate or be used to indicate, for example, one or more of: the volume of a fluid, the flow rate of a fluid, a characteristic of a component within a fluid (for example, how long a component has some activity, such as chemiluminescence, fluorescence, and others), and for how long a particular fluid has been positioned in a specific region of the microfluidic device.
[0035] In some modalities, a position of a signal in time in relation to a second position in time or in relation to another process or event (for example, that occurs in the microfluidic system) is generated and / or determined. For example, a detector can detect when a certain fluid passes through the detector (for example, a first position in time), and the timing of that signal can be related to a second position in time (for example, when detection started; a certain amount of time after a process has occurred, etc.). In another example, a detector can detect when a certain fluid has passed through the detector after (or before) a component of the microfluidic system (for example, a valve) is activated. In one modality, the
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13/101 opening a valve may indicate that the mixing of the reagents is about to occur, and thus the position of the signal in time may give some indication of when a certain fluid passes through the detector after (or before) the mixing of the reagents . If the position of the fluid signal occurs within some time after (or before) the mixing of the reagents, for example, this may indicate that the analysis is working correctly. In another example, a detector can detect when a second fluid passes through the detector after a first fluid has passed through the detector. In other embodiments, a position of a signal in time is determined in relation to a particular event or process that is occurring or has occurred in the microfluidic system (for example, the beginning of the analysis, the initiation of the fluid flow, the initiation of detection in the microfluidic system, when a user inserts the microfluidic device into an analyzer, etc.).
[0036] In another set of modes, an average time between signals or events is generated and / or determined. For example, the average period of time between two signals can be measured, where each of the signals can independently correspond to one or more characteristics or conditions described here. In other embodiments, the average time between the first and the last of a series of similar signals is determined (for example, the average time between a series of wash fluids that pass through a detector).
[0037] In certain modalities, a signal pattern is generated and / or determined. The signal pattern may include, for example, at least two (or, in other embodiments, at least three, or at least four) of a signal strength, a frequency of the signals, a duration of a signal, a position of a signal in time in relation to a second position in time or in relation to another process or event that occurs (or has occurred) in the microfluidic system
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14/101 co, and an average period of time between two or more signals or events. In other embodiments, the signal pattern comprises at least two (or, in other embodiments, at least three, or at least four) of an intensity of a first signal, a duration of the first signal, a position of the first signal at the time in relation to a second position in time; an intensity of a second signal, a duration of the second signal, a position of the second signal in time with respect to a second position in time, and an average period of time between the first and second signals. The pattern of signals may indicate, in some modalities, whether a particular event or process is occurring correctly within the microfluidic system. In other embodiments, the signal pattern indicates whether a particular process or event has occurred in the microfluidic system. In still other embodiments, a pattern of signals may indicate a particular sequence of events.
[0038] A variety of signals or signal patterns, such as those described above, can be generated and / or determined and can be used alone or in combination to provide feedback to control one or more processes, such as flow modulation of fluid in a microfluidic system. That is, the control system or any other appropriate unit can determine, in some embodiments, whether the flow of fluid in the microfluidic system should be modulated based at least in part on the signal pattern. For example, determining whether fluid flow should be modulated based at least in part on a signal pattern that includes an intensity of a first signal and a time position of the first signal in relation to a second position in time may involve the use of both of these pieces of information to make a decision as to whether or not the fluid flow should be modulated. For example, these signals can be compared to one or more reference signals (for example, a
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15/101 limit intensity or an intensity range, and a time limit position or range of positions in time, in relation to a second position in time) that can be pre-programmed or preset in the control system. If each of the measured signals falls within the values or the respective limit ranges, a decision should be made whether the fluid flow can be made. Only one of the parameters to be considered (for example, only an intensity of the first signal or only a position in the time of the first signal) that satisfies a limit value or range may not be enough information to make a decision on whether or not to modulate the fluid flow, because it may not provide enough information about the fluid or the components that caused the signal (s) for the purposes described here. For example, in some cases the fluid or component detected cannot be sufficiently identified for the purposes described here unless a signal pattern is taken into account.
[0039] In certain modalities, one or more measured signals are processed or manipulated (for example, before or after transmission, and / or before being compared to a signal or reference value). It should be appreciated, therefore, that when a signal is transmitted (for example, to a control system), compared (for example, with a signal or a reference value), or else used in a feedback process, that signal raw can be used or a processed / manipulated signal can be used based (at least in part) on the raw signal. For example, in some cases, one or more signals derived from a measured signal can be calculated (for example, using a differentiator, or some other appropriate method) and used to provide feedback. In other cases, the signals are normalized (for example, by subtracting a measured signal from a background signal). In a set of modes, a signal comprises an inclination
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16/101 nation or mean slope, for example, a mean slope of intensity as a function of time.
[0040] In some cases, the measured signal can be converted into a digital signal with the use of an analog to digital converter so that any other additional signal processing can be performed by a digital computer or digital signal processor. Although in one embodiment all signal processing is performed digitally, the present invention is thus not limited, since analogous processing techniques can alternatively be used. For example, a digital to analog converter can be used to produce an output signal. The signals can be processed in a time domain (one-dimensional signals), in the spatial domain (multidimensional signals), in the frequency domain, in the autocorrelation domain, or in any other appropriate domain. In some cases, signals are filtered, for example, using a linear filter (a linear transformation of a measured signal), a non-linear filter, a causal filter, a non-causal filter, a time-invariant filter, a variant filter time, or other appropriate filters. It should be understood that the signals, patterns, and their use in the feedback described here are exemplary and that the invention is not limited in this regard.
[0041] Once a signal or a signal pattern is determined, the signal (s) can (optionally) be transmitted to a control system. In some cases, the control system compares the signal or the signal pattern with a second set of signals. The second signal or signal pattern can be, for example, the signal (s) determined previously in the microfluidic system, or signal (s) or reference value (s) that can be pre-programmed in the system control unit or another unit of the microfluidic system. In some cases, a reference signal or signal pattern includes one or more
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17/101 plus limit values or a range of limit values. The control system can compare a first signal or signal pattern with a second signal or signal pattern (for example, reference signals), and determines whether to start, stop, or modulate one or more events or series of events in the system microfluidic. That is, the signal or the measured signal pattern can be used by the control system to generate an activation signal and provide feedback control to the microfluidic system. For example, the control system can determine whether to modulate the fluid flow (for example, flow, mixing, interrupting a flow of one or more fluids) in one or more regions of the microfluidic system. Other conditions such as the modulation of temperature, pressure, humidity, or other conditions can also be controlled. This modulation can be performed, in certain modalities, by the control system that sends one or more activation signals to an appropriate component of the microfluidic system (for example, a valve, pump, vacuum, heater, or other component) to activate that or another component. Any appropriate electronic valve activation circuit can be used to receive an activation signal and convert the activation signal to a voltage, current, or other signal capable of activating the component. In certain embodiments, the control system can determine whether or not an operation of one or more components of the microfluidic system should cease. In some cases, the control system can determine whether or not to stop an analysis or part of an analysis that is being done in the microfluidic system.
[0042] In some modalities, a method of carrying out feedback control may involve initiating the detection of fluids in a first measurement zone of a microfluidic system. A first fluid and a second fluid can be detected in the first measurement zone and a first signal that corresponds to the first flow
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18/101 do and a second signal corresponding to the second fluid can be formed. A first pattern of signals can be transmitted to a control system, wherein the first pattern of signals comprises at least two out of an intensity of the first signal, a duration of the first signal, a position of the first signal in time with respect to a second position in time, and an average period of time between the first and second signals. A decision whether to modulate the fluid flow in the microfluidic system can be made based at least in part on the first signal pattern.
[0043] It should be understood that although much of this description describes the use of signals or signal patterns, the invention is not so limited and that aspects of feedback control or other processes that involve determining the characteristics, conditions or events that involve fluids or components may not require the generation, determination (eg measurement) or analysis of signals or signal patterns in some modalities.
[0044] In some embodiments, a method of performing feedback involves the detection of a first fluid and a second fluid in a first measurement zone of a microfluidic system, in which the detection step comprises the detection of at least two (or at least three) out of an opacity of the first fluid, a volume of the first fluid, a flow of the first fluid, a detection position of the first fluid in time relative to a second position in time, and an average period of time between detection of the first and second fluids. A decision whether to modulate the fluid flow in the microfluidic system can be made based at least in part on the detection step.
[0045] In some modalities, the feedback control can be used to modulate the same condition, event, or type of condition or event that were first detected. For example,
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19/101 The concentration of a component in a fluid can be determined, and a signal can be generated and transmitted to a control system, which determines whether or not the concentration of the same component should be increased or decreased in the region of the microfluidic device. In another example, the flow rate of a fluid in a channel is measured, and based at least in part on the generated measurement signal, the source of fluid flow (eg a vacuum or a pump) or a valve is used to modulate the flow in that same channel. In such and other embodiments, the generated signal can be compared to a predetermined signal or to values that indicate a desired value or range of conditions (for example, concentration, flow). Control of feedback may involve a feedback loop (a feedback loop, for example, positive or negative) in some cases. In other cases, feedback control does not involve a feedback loop.
[0046] In other modalities, however, (including many of the examples described here) the control of feedback is based at least in part on the determination of one or more first conditions or events that occur in the microfluidic system, and the signals of one or more more conditions or events are used to control a second, different set of conditions or events that occur (or the events that will occur) in the microfluidic system. In certain embodiments, the second different set of conditions or events does not substantially affect the first set of conditions or events (for example, as opposed to the above examples involving the modulation of a component's concentration or flow in a channel). In some cases, detection occurs in a measurement zone, and feedback from the measurement zone is used to modulate fluid flow in a different region of the microfluidic system. For example, the detection of any fluid passing through a detection system can trigger control over whether a particular valve is activated or not.
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20/101 to allow a flow of one or more different fluids in a different region of the microfluidic system. In a particular embodiment, the detection of a first fluid (for example, which passes transversely) in a reaction area can cause the mixing of the second and third fluids in a mixing region of the microfluidic system. The second and third fluids can initially be positioned in a different region (for example, a storage region) of the microfluidic system where the detection and production of the signal used to provide feedback occurs. In another example, measuring the optical density of a sample flowing through a measurement zone (for example, a first condition) gives an indication of whether the sample was introduced at the right time and / or in the presence of the correct type or volume Sample. One or more signals from this measurement can be compared to one or more preset values, and based (at least in part) on this feedback and comparison, a control system can stop the fluid flow in the microfluidic system (for example, a second different condition) if the measured signals fall out of range with the preset values. In some of these and other modalities, the first condition or event has passed after the detection step, in such a way that the feedback control does not substantially modulate that same condition, event, or type of condition or event that produced the signal used for the feedback.
[0047] In one embodiment, one or more methods of controlling feedback such as proportional control, integral control, proportional-integral control, derived control, proportional-derived control, integral-derived control, and proportional-integral control derivative, can be used by a control system to modulate fluid flow. Feedback control may involve a feedback loop in some modalities. In some cases that
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21/101 involve one or more of the aforementioned feedback control methods, an activation signal (which can be used to modulate fluid flow, for example, by activating a component of the microfluidic system) can be generated based on at least partly in a signal that is the difference between a pre-programmed limit value or signal (which may be indicative of a future action to be taken) and a feedback signal that is measured by a detector.
[0048] The detection of a condition or event that occurs in a microfluidic system can take a variety of forms. In some cases, detection occurs continuously. In other modalities, detection occurs periodically; however, in other modalities, detection occurs sporadically. In some cases, detection occurs about a specific event or condition.
[0049] As described herein, detection can occur in any appropriate position with respect to a microfluidic device. In some cases, one or more detectors are stationary with respect to a microfluidic device during use and / or during detection. For example, a stationary detector can be positioned adjacent to a particular region of the microfluidic device, such as a detection region or the measurement zone, where one or more events (for example, a chemical or biological reaction) occur. The detector can detect, for example, the passage of fluids through the measurement zone. Additionally or alternatively, the detector can detect the connection or association of other components in that region (for example, the connection of a component to the surface of the measurement zone). In some embodiments, a stationary detector can monitor multiple measurement zones simultaneously. For example, a detector such as a camera can be used to image an entire microfluidic device, or a large portion of the device, and only certain areas of the device scrutinize
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22/101 do. Components such as optical fibers can be used to transmit light from multiple measurement zones to a single detector.
[0050] In other modalities, a detector is removably positioned with respect to the microfluidic device during use and / or during detection. For example, a detector can be physically moved through different regions of the microfluidic device to detect the movement of fluids through the device. For example, a detector can track the movement of certain fluids and / or components in the channels of the microfluidic device. Alternatively, the fluidic device can move relative to a stationary detector. Other configurations and uses of the detectors are also possible.
[0051] Examples of signals or signal patterns that can be used to control feedback are shown in the example mode illustrated in figure 2. Figure 2 is a graph showing the detection of various fluids as they flow in a region of a device (for example, a channel) and pass through a detector. Graph 100 shows the measurement of optical density in arbitrary units (y-axis) as a function of time (x-axis). In certain modalities, the transmission and / or absorbance of a fluid, for example, can be detected as it passes through a region of a microfluidic system. An optical density equal to zero can indicate that the maximum light transmission (for example, low absorbance) and a higher optical density can indicate low transmission (for example, higher absorbance). Since different fluids flowing through the detector may have different susceptibilities to light transmission or absorbance, the detection of specific fluids, including their volumes, flows, and types of fluids, can be determined.
[0052] For example, as shown illustratively in the
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23/101 figure 2, a first fluid that produces signal 110 can pass through the detector around time = 0.1 second up to about 700 seconds. (Time = 0 seconds can indicate, for example, initiation of detection). The first fluid 110 has a particular intensity 112 (for example, an optical density of about 0.23). If a particular type of fluid that has a specific intensity or scale of intensities has to flow through the detector at a particular point in time (for example, at a time about 400 seconds after the initiation of detection) or between a certain period of time (for example, some time between 0 and 800 seconds), confirmation that this process has occurred can be detected. For example, the first fluid 110 may, in some embodiments, be a particular type of sample that must be introduced into the microfluidic device to perform a particular analysis. If the type of sample is associated with a particular intensity (for example, whole blood will give an optical density of about 0.23), the type of sample can be checked by determining whether or not that sample has an intensity within a range. allowed range.
[0053] In addition, the proper introduction of the sample into the device at a correct time (for example, at the beginning of the analysis) can be verified by determining where the sample signal occurs as a function of time (along the x-axis). For example, the time when the sample reaches the measurement zone (observed at an OD that has some range or intensity) can be monitored. If the sample takes too long to enter the measurement zone, this could indicate, for example, a leak or an obstruction in the system. If the sample takes a long time to reach the first measurement zone or there is too much time between the sample or parts of the sample that reach multiple measurement zones (which can be positioned in parallel or in series), the test can be canceled.
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[0054] Additionally, the volume of the first fluid producing signal 110 can be determined and verified by measuring the time period 114 of the signal. If the particular process to be performed on the microfluidic device requires a sample that has a particular volume, this can be verified. For example, a sample that has a particular volume (for example, 10 pl) can be expected, corresponding to a predicted range of the flow time (for example, a signal that has some duration) at a certain intensity (for example, OD Sample). The test can ensure that the user correctly loads the sample into the fluid connector or other appropriate sample introduction device. If the duration of the sample signal is too short (which may indicate that not enough sample has been introduced) or too long (which may indicate that too much sample has been introduced) the test may be canceled and / or the results may be neglected.
[0055] If, for example, the intensity, the time period, or the positioning of the signal 110 that results from the first fluid are incorrect, the control system can activate a secondary process that can, for example, modulate the fluid flow in the microfluidic system. For example, in a set of modalities, the control system may determine that once an incorrect type or volume of sample has been introduced into the device, or introduced into the device at an incorrect time, the analysis to be made by the microfluidic device must be canceled. In other embodiments, the cancellation can occur due to a problem with the device (for example, an obstruction in the channels that does not allow fluid to flow at a particular flow), or a problem with an analyzer used to analyze the device (for example , the malfunction of one or more components such as a valve, a pump, or a vacuum).
[0056] The analysis can be canceled, for example, by modulating the
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25/101 fluid flow in the microfluidic system (for example, by sending a signal to a pump or a vacuum to stop the flow of fluids), cutting power to certain components of the system, ejecting the microfluidic device / cassette from the analysis system (for example, automatically or by informing a user to do so), or by other processes.
[0057] In other modalities, an abnormality that occurs in the system causes the occurrence of a secondary event, but does not cancel the analysis. In some cases, a user may be alerted that an abnormality has occurred in the system. The user can be informed that the test results must not be reliable, that the analysis needs to be performed again, that the analysis may take a long time to be performed, or that the user must take some action. In some cases, the user may be notified and then prompted to verify whether one or more processes in the microfluidic system, or analysis being performed, should be continued or not. Other methods of quality control are also possible.
[0058] In a set of modalities, a method of carrying out quality control to determine abnormalities in the operation of a microfluidic system includes the detection of a first fluid (for example, which passes transversely) in a first measurement zone of the microfluidic system and forms a first signal that corresponds to the first fluid, and transmits the first signal to a control system. The first signal can be compared to a reference signal, thereby determining the presence of abnormalities in the operation of the microfluidic system. The method may include determining whether the operation of the microfluidic system should be stopped based at least in part on the results of the comparison step. In some cases, the control system can determine whether or not to interrupt an analysis or a part of an analysis that is
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26/101 being made in the microfluidic system.
[0059] As shown illustratively in figure 2, the type of fluid that through a detector can be determined at least in part by the intensity of the signal generated by the fluid. For example, while the signal 110 of a first fluid has a high intensity (for example, a low light transmission), a second series of signal-producing fluids 120, 122 and 124 has a relatively low intensity (for example, a high light transmission). The graph also indicates the relative separation between the first signal production fluid 110 and the second signal production fluids 120, 122 and 124. For example, the difference between time period 125 and time period 114 can give a indication of how quickly the second set of fluids is flowing through the detector after the first fluid has finished passing through the detector. In some embodiments, this difference in time can be compared to one or more signals or reference values (for example, a predetermined amount of separation time or time range that is supposed to occur between the first fluid and the second fluids). A difference in time that does not match the signal or reference value, or falls within a permissible range, may indicate that an abnormality has occurred in the microfluidic system. For example, if the time difference between time periods 125 and 114 is too long, this may indicate that the flow of fluid has been obstructed (for example, due to an obstruction in a channel by an air bubble or other means) , but cleared later in the microfluidic device. In some embodiments, this can influence the test being performed, and in this way the control system can determine whether or not one or more processes should be stopped or modified in the microfluidic system.
[0060] As shown illustratively in figure 2, the
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27/101 fluid worlds of signal production 120, 122 and 124 are separated by peaks 126, 128 and 130. These peaks represent the fluids that are fluid between the second fluids. As described in more detail here, in some cases these separation fluids can be fluids that are immiscible with the fluids they are separating. For example, in a set of modalities, the second signal production fluids 120, 122 and 124 are washing solutions that pass through the measurement zone. These washing fluids can be separated by immiscible (separating) fluids (for example, air plugs) that produce signals 126, 128 and 130. Washing solutions can have a relatively high transmission and, therefore, a relatively optical density low, while air plugs may have a relatively lower light transmission (e.g., a relatively higher optical density) due to light scattering as these fluids pass through the detector. Because of the different susceptibility of these fluids to light transmission, the different fluids (including the type, phase, volume, flow rate of the fluid) can be different. In addition, the sequence of second fluids passing through the detector can have a time period 134, which can optionally be compared to a time period or range of the ideal time period and can optionally be used to control feedback.
[0061] In certain modalities, the number of washes (peaks and valleys) is counted and a control system cancels the analysis if the predicted number is not observed. Few washes can mean that the reagents have evaporated during storage of the device (which indicates a leak) or a problem with the fluid connector connection. Very few washes can also indicate that the correct number was not loaded on the device when manufacturing the device. Too many washes should indicate that tampons
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28/101 of the wash broke during storage.
[0062] Figure 2 also shows a third signal-producing fluid 135 that passes through the measurement zone after the second fluids flow. Since the third fluid has an optical density similar to that of the second set of fluids, the third fluid can be identified or distinguished from other liquids at least in part by its time period 136, which can give an indication of the fluid volume. The position of the time period 136 along the time line (or in relation to one or more other signals present) can also give an indication of the fluid being flowed through the measurement zone. For example, the analysis can be designed in such a way that a fluid that provides a certain optical density (for example, ~ 0.01) and duration (for example, ~ 200 seconds at a particular flow rate to be used or the pressure to be applied ) will occur between 900 seconds and 1,200 seconds after starting the analysis. These parameters can be pre-programmed in the control system, and compared with the signal 135 measured by the detector.
[0063] The third signal-producing fluid 135 can be any appropriate fluid, and in some cases is the reagent to be used in a chemical and / or biological reaction to be performed on the microfluidic device. For example, as described in more detail below, the third fluid can be a detection antibody that can bind with one or more components in the sample. In other embodiments, however, a detection antibody is bound with a component of the sample before the sample flows through the detector. Other binding configurations of a detection antibody are also possible, and in some embodiments no detection antibody is used at all.
[0064] After the third fluid is flowed through the measurement zone, a series of fourth signal-producing fluids 140, 142,
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144, 146, 148 and 150 can flow through the measurement zone. Each of the fourth fluids can be separated by an immiscible fluid (for example, air caps) producing signals 154. In certain embodiments, the frequency of signals that have a certain limit (for example, air caps for producing signals 154 that have a limit above an optical density of 0.05 and / or a series of fluid quarters that have an optical density below 0.01) can be used to activate one or more events in the microfluidic system.
[0065] In some cases, the intensity and frequency of a series of fluids can be combined with a total period of time between the first and the last of such fluids (for example, the time period 158 covering the series of quarters fluids). For example, the feedback or activation of an event can be based at least in part on the frequency of the signals (for example, peaks) observed in combination with one or more periods of time between adjacent signals, and / or in combination with the intensity of the signals, and / or in combination with the time period between the first and the last signal of that type or intensity. Optionally, one or more of the signals can be used in combination with the average position of the signals in relation to the time range of the events along the timeline (for example, the average time 158 between signals 140 and 150 in relation to a or more other signals or landmarks (for example, time = zero)).
[0066] In some embodiments, the event that is triggered by a signal pattern is the modulation of the fluid flow within the microfluidic system. For example, one or more of a pump, vacuum, valve system, or other component can be driven based at least in part on the presence or absence of a particular pattern of signals. As an example, a signal pattern can trigger a valve that allows one or more fluids to flow
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30/101 to a particular channel of the microfluidic device. For example, activation of the valve can allow two fluids that are kept separate during the storage of liquids in the device to mix in a common channel. In a particular embodiment, a mixed fluid includes an amplification reagent that allows amplification of a signal in a measurement zone of the device. Specific examples are provided in more detail below.
[0067] As described herein, a detector can not only detect the passage of fluids through a region of a microfluidic device, but can also detect the presence or absence of an event or condition that occurs in a region of the device microfluidic. For example, in some cases a link event is detected. In other embodiments, the accumulation and / or deposition of a component in a particular region of the microfluidic device is detected. And in still other modalities the amplification of a signal is detected. Such processes can take place at any appropriate position within a region of a device. For example, the event or condition can occur within a fluid positioned in the device region, on a channel surface or a device chamber, or in a component positioned within the device region (for example, on a a granule, in a gel, in a membrane).
[0068] In some cases, the progression of the event or condition can be determined, and optionally compared to one or more signals or reference values (which can be pre-programmed in the control system). For example, as shown illustratively in figure 2, a peak 160 may form due to the configuration of a signal (for example, an opaque layer) in a measurement zone. This peak slope can be measured and compared with one or more control values to determine whether a correct process is
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31/101 or not occurring or occurred in the measurement zone. For example, if the slope of peak 160 is within a particular range of acceptable values, this may indicate that there was no abnormality in the storage of the reagents that were used in part to produce the signal.
[0069] In a set of modalities, peak 160 indicates an amplification reagent entering the measurement zone. The analysis can be designed and configured in such a way that the amplification reagent enters the measurement zone within a certain period of time after a certain event has occurred (for example, with the activation of a valve). In some cases, the amplification reagent must have some optical density associated with it (for example, a low optical density if the reagent is a clear liquid). If the reagent takes too long to reach the measurement zone and / or if the initial optical density is too high, the test can be canceled. If the reagent has a high optical density (for example, dark or opaque), this could indicate that the reagent has spoiled (for example, during the storage of the reagent in the device).
[0070] In some embodiments, a device may include multiple measurement zones (for example, in parallel or in series). A measurement zone can be used as a negative control. For example, minimal binding or deposition of a substance (for example, an opaque layer) and therefore of a low optical density in some embodiments, can be expected in the negative control measurement zone. If a detector measures a high optical density in the negative control measurement zone, this may indicate, for example, a non-specific connection. In some cases, the signal from that measurement zone can be considered background and be subtracted from the signals in the other measurement zones to overcome the alloy
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32/101 non-specific information that can occur throughout the system. If the fund is too high, the test can be canceled. This may, for example, indicate a problem with the amplification reagents or other reagents used in the analysis.
[0071] In some embodiments, a device may include a measurement zone used as a positive control. Positive control may, in some embodiments, include a known amount of analyte linked to the measurement zone (for example, the channel walls), and the level of the optical density signals at some point in time, the slope of those signals, or the change in the slope of these signals in the zone may fall within a predicted range. These ranges can be determined when calibrating a specified batch of devices. In some cases, as described in more detail here, that information can be included in the batch-specific information transferred to an analyzer by using a batch-specific label, such as a barcode, a memory bar, or a label. radio frequency identification (RFID). If the reference levels for these measurement zones fall outside the range, the test can be canceled. Similar to the above, these signals can also be used to adjust the test signal (for example, increasing the test signal slightly if these signals are high, decreasing the test signal if these signals are low).
[0072] The presence of obstructions such as bubbles or other components during one or more events (for example, amplification, mixing) and / or at one or more unexpected positions in time can indicate problems in the analysis, such as a leak in a valve . These bubbles or other components can be detected as peaks that have some intensity in the optical density pattern (which can be similar to the air buffer peaks used during washing). If these are observed in unexpected places, the test
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33/101 can be canceled.
[0073] It should be understood that when the optical density (for example, transmission or absorbance) was determined in figure 2, in other modalities other types of signals can be measured using an appropriate detector. The signals can be produced in the absence of a label (such as when measuring optical density), or produced when using a label. A variety of different labels can be used, such as fluorescent markers, dyes, quantum dots, magnetic particles, and other labels known in the art.
[0074] As shown illustratively in figure 2, in some modalities an analysis made on a device can be recorded to essentially produce a fingerprint of the analysis. All parts of the fingerprint can be used to provide feedback to the microfluidic system. In some cases, the fingerprint includes signs of the passage of substantially all fluids used in an analysis through a region of the device. Since the different fluids used in the analysis can have different volumes, flow rates, compositions, and other characteristics, these properties can be reflected in the fingerprint. In this way, the fingerprint can be used to identify, for example, the fluids used in the analysis, the timing of the fluids (for example, when particular fluids have been introduced in certain regions of the device), the interaction of the fluids (for example, the mixing). In some embodiments, the fingerprint can be used to identify the type of analysis performed on the device and / or the test format (for example, a sandwich test versus a competitive test) of the analysis.
[0075] In a set of modalities, the fingerprint as a whole (for example, the general form, the duration, and the timing of
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34/101 all signals) is used to perform quality control at the end of the analysis. For example, the fingerprint can be compared to a fingerprint of the control to determine if the analysis was done correctly after all fluids have been fluid. The control system can, in some cases, notify the user if the analysis was done correctly (for example, through a user interface).
[0076] In other embodiments, a detector can be positioned within certain regions of a microfluidic system and can only determine the presence or passage of certain, but not all, fluids through the detector. For example, a detector can be positioned in a mixing region to determine the proper mixing of fluids. If the fluids are mixed correctly (for example, a mixed fluid that has a certain property such as a certain concentration or volume is produced) or mixed at an appropriate point in time in relation to one or more other events that occur in the analysis, the Feedback control can allow the mixed fluid to flow into another region of the device. If the mixed fluid does not have one or more desired or predetermined characteristics, the feedback control can prevent the mixed fluid from flowing into the region and, in some embodiments, can initiate a second set of fluids to be mixed and transported to the region .
[0077] In certain modalities, the feedback control includes the use of two or more detectors. A first detector can determine a first set of signals, and a second detector can determine a second set of signals. The first and second sets of signals can be compared with each other, and / or each can be compared with a set of signals or reference values that can be pre-programmed in a control system. For example, a device can include a plurality of
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35/101 measurement zones, where each measurement zone is associated with a detector that measures signals in that region. In some cases, the system is designed and configured in such a way that a first detector determines an analysis fingerprint that substantially matches the analysis fingerprint of a second detector. If the fingerprints do not match, however, this may indicate that an abnormality has occurred within the system. In some cases, the first and / or second detectors may detect the passage of all fluids used in the analysis through a region of the device, or only certain (but not all) fluids that pass through a region of the device, such as described above. In other modalities, controlling feedback, or determining a value in general, may involve using the signals detected from multiple measurement zones. For example, flow can be determined by measuring how long a bubble or leading edge of a fluid takes to travel between two measurement zones.
[0078] The control of feedback and other processes and methods described herein can be performed using any appropriate microfluidic system, such as those described in more detail below. In some cases, the microfluidic system includes a device or cassette that can be configured to be inserted into a microfluidic sample analyzer. Figures 3-6 illustrate several exemplary modalities of cassette 20 for use with an analyzer. As shown illustratively in these figures, cassette 20 can be substantially in the form of a card (i.e., similar to a card key) with a substantially rigid plate-like structure.
[0079] Cassette 20 can be configured to include a fluidic connector 220, which, as shown in the example mode illustrated in Figure 3, can fit on one end of the cassette
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20. In certain embodiments, the fluidic connector can be used to introduce one or more fluids (for example, a sample or a reagent) into the cassette.
[0080] In a set of modalities, the fluidic connector is used to fluidly connect two (or more) channels of the cassette during the first use, in which the channels are not connected before the first use. For example, the cassette may include two channels that are not in fluid communication before the first use of the cassette. Unconnected channels can be advantageous in certain cases, such as storing different reagents in each channel. For example, a first channel can be used to store dry reagents and a second channel can be used to store wet reagents. When the channels are physically separated from each other, it is possible to enhance the long-term stability of the reagents stored in each of the channels, for example, keeping the reagent (s) stored in dry form protected against moisture that can be produced by the reagent (s) stored in the wet form. In the first use, the channels can be connected through the fluid connector to allow a fluid communication between the channels of the cassette. For example, the fluidic connector can puncture seals that cover cassette inlets and / or outlets to allow insertion of the fluidic connector into the cassette.
[0081] As used herein, before the first use of the cassette means a moment or moments before the cassette is first used by an intended user after commercial sale. The first use can include any (any) step (s) that require the user to manipulate the device. For example, first use may involve one or more steps such as drilling a sealed inlet to introduce a reagent into the cassette, connecting two or more channels to cause fluid communication
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37/101 between channels, preparing the device (for example, loading reagents into the device) before analyzing a sample, loading a sample into the device, preparing a sample in a region of the device, running a reaction with a sample, the detection of a sample, etc. The first use, in this context, does not include manufacturing or other preparatory or quality control steps performed by the cassette manufacturer. Those skilled in the art are well aware of the meaning of the first use in this context, and can easily determine whether a cassette of the invention has experienced the first use or not. In a set of embodiments, the cassette of the invention is disposable after first use (for example, after completion of an assay), and is particularly evident when such devices are used for the first time, because it is typically impractical to use the devices in absolute (for example, to run a second test) after the first use.
[0082] A cassette can be attached to a fluidic connector using a variety of mechanisms. For example, the fluidic connector can include at least one non-fluidic feature complementary to a cassette-shaped feature a non-fluidic connection between the fluidic connector and the cassette with the fixation. The complementary non-fluidic feature can be, for example, a protruding feature of the fluidic connector and corresponding complementary cavities in the cassette, which can assist the user in aligning the fluidic connector with the cassette. In some cases, the feature creates substantial resistance to the movement of the fluidic connector in relation to the cassette and / or alignment element when the alignment element receives the fluidic component (for example, with the insertion of the fluidic component in the alignment element) and / or during the intended use of the device. The fluidic connector and / or cassette can
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38/101 optionally include one or more characteristics such as fitting characteristics (for example, cutouts), grooves, openings for inserting clamps, cross-closing mechanisms, pressure inserts, friction inserts, threaded connectors such as screw inserts, inserts pressure switches, adherent fittings, magnetic connectors, or other appropriate coupling mechanisms. Connecting the fluidic connector to the cassette may involve the formation of a liquid-impermeable and / or air-impermeable seal between the components. The attachment of a fluidic connector to a cassette can be reversible or irreversible.
[0083] As shown, cassette 20 can be configured to include a fluidic connector 220. In particular, cassette 20 can include a fluidic connector alignment element 202 that is configured to receive and couple with connector 220. For example, For example, the alignment element can be constructed and arranged to couple the fluidic connector and thereby position the connector in a predetermined stipulated configuration in relation to the cassette. As shown in the illustrative embodiments of figure 3, the cassette may include an alignment element that extends more or less perpendicular to the cassette. In other embodiments, the alignment element may extend more or less parallel to the cassette.
[0084] In some embodiments, the configuration of the alignment element and the fluidic connector can be adapted to allow the insertion of the fluidic connector in the alignment element by a sliding movement. For example, the fluidic connector can slide against one or more surfaces of the alignment element when the fluidic connector is inserted into the alignment element.
[0085] Fluidic connector 220 may include a substantially U-shaped channel that may contain a fluid and / or reagent (e.g., a fluid sample) before being connected to the cassette.
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The channel can be housed between two protective components that form the connector. In some embodiments, the fluidic connector can be used to collect a sample from the patient before the fluidic connector is connected to the cassette. For example, with a blood sample, the fluid connector can be configured to pierce a patient's finger to collect the sample in the channel. In other embodiments, the fluidic connector does not contain a sample (or reagent) prior to connection to the cassette, but simply allows fluid communication between two or more channels on the cassette with the connection. In one embodiment, the U-shaped channel is formed with a capillary tube. The fluidic connector can also include other channel configurations, and in some embodiments, it can include more than one channel, which can be fluidly connected or not connected to each other.
[0086] As shown illustratively in the exploded view of figure 4, cassette 20 may include a cassette body 204 that includes at least one channel 206 configured to receive a sample or a reagent. The cassette body 204 can also include the latches 208 positioned at one end that couple with the fluid connector alignment element 202 for a pressure fit.
[0087] Cassette 20 can also include the top and bottom covers 210 and 212, which can, for example, be made of a transparent material. In some embodiments, a cover may be in the form of a biocompatible adhesive and may be made of a polymer (for example, PE, COC, PVC) or an inorganic material, for example. In some cases, one or more covers are in the form of a cling film (for example, a tape). For some applications, the material and dimensions of a cover are chosen in such a way that the cover is substantially impermeable to vapor
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40/101 water. In other modalities, the coating may be non-adherent, but it can be thermally bonded to the microfluidic substrate by the direct application of heat, laser energy, or ultrasonic energy. Any (Any) input (s) and / or output (s) of a cassette channel can be sealed (for example, by placing a sticker over the input (s) and / or the output (s) ) when using one or more coatings In some cases, the coat substantially seals one or more reagents stored in the cassette.
[0088] As illustrated, the cassette body 204 may include one or more ports 214 coupled to channel 206 on the cassette body 204. These ports 214 can be configured to align with the substantially U-shaped channel 222 on fluid connector 220 when fluid connector 220 is coupled to cassette 20 to fluidly connect channel 206 on body 204 of the cassette with channel 222 on the connector fluidic 220. As shown, a cover 216 can be provided over ports 214 and the cover 216 can be configured to be patched or else opened (for example, by connector 220 or other means) to fluidly connect the two channels 206 and 222. In addition, a cover 218 can be provided on the cover port 219 (for example, a vacuum port) on the body 204 of the cassette. As indicated in more detail below, port 219 can be configured to fluidly connect a fluid flow source 40 with channel 206 to move a sample through the cassette. The cover 218 over port 219 can be configured to be perforated or open to fluidly connect channel 206 with fluid flow source 40.
[0089] The cassette body 204 can optionally include a liquid-containing region, such as a disposal area, including an absorbent material 217 (e.g., a disposal pad). In some modalities, the liquid containment region inPetição 870200007448, of 1/16/2020, p. 47/124
41/101 includes regions that capture one or more liquids that flow into the cassette, while allowing gases or other fluids in the cassette to pass through the region. This can be achieved, in some embodiments, by positioning one or more absorbent materials in the liquid containment region to absorb the liquids. This configuration can be useful for removing air bubbles from a fluid stream and / or for separating hydrophobic liquids from hydrophilic liquids. In certain embodiments, the liquid containment region prevents liquids from passing through the region. In some of such cases, the liquid containment region can act as a disposal area by capturing substantially all of the liquid in the cassette, thereby preventing liquids from leaving the cassette (for example, by allowing gases to escape from an outlet the cassette). For example, the disposal area can be used to store the sample and / or reagents in the cassette after they have passed through channel 206 during sample analysis. These and other arrangements can be useful when the cassette is used as a diagnostic tool, because the liquid-containing region can prevent a user from being exposed to potentially harmful fluids in the cassette.
[0090] Figure 5 shows a cassette that has a certain configuration of channels and includes several components of a microfluidic system for handling fluids. Figure 6 shows another example of a configuration of channels that can be part of a cassette. As shown illustratively in Figures 5 and 6, in some embodiments a cassette may include a first channel 206 and a second channel 207 spaced from the first channel. In one embodiment, channels 206, 207 vary in dimension in greater cross section from about 50 micrometers to about 500 micrometers, although other channel sizes and configurations can be used, as described in more detail below.
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[0091] The first channel 206 may include one or more measurement zones used to analyze the sample. For example, in an illustrative embodiment, channel 206 includes four measurement zones 209 that are used during sample analysis (see figure 6).
[0092] In certain modalities, one or more measurement zones are in the form of winding regions (for example, regions involving winding channels). A winding region can, for example, be defined by an area of at least 0.25 mm ² , at least 0.5 mm ² , at least 0.75 mm ² , or at least 1.0 mm ² , where at least minus 25%, 50% or 75% of the area of the winding region comprises an optical detection passage. A detector that allows the measurement of a single signal across more than one of the adjacent segments of the winding region can be positioned adjacent to the winding region.
[0093] As described herein, the first channel 206 and / or the second channel 207 can be used to store one or more reagents used to process and analyze the sample prior to the first use of the cassette. In some embodiments, dry reagents are stored in one channel or section of a cassette and wet reagents are stored in a second channel or section of the cassette. Alternatively, two separate sections or channels of a cassette can both contain dry reagents and / or wet reagents. The reagents can be stored and / or arranged, for example, as a liquid, a gas, a gel, a plurality of particles, or a film. Reagents can be positioned in any appropriate part of a cassette, including, but not limited to, a channel, reservoir, surface, and / or membrane, which can optionally form part of a reagent storage area . A reagent can be associated with a cassette (or the components of a cassette) in any appropriate manner. For example, reagents can be cross-linked (for example, covalent or ionic
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43/101), absorbed, or adsorbed (fisissorvidos) on a surface inside the cassette. In a particular embodiment, all or part of the channel (such as a fluid passage from a fluid connector or a cassette channel) is coated with an anticoagulant (for example, heparin). In some cases, a liquid is contained within a channel or reservoir in a cassette before first use and / or before introducing a sample into the cassette.
[0094] In some embodiments, stored reagents may include fluid buffers positioned in a linear order so that, during use, as fluids flow to a reaction site, they are distributed in a predetermined sequence. A cassette designed to perform an assay, for example, may serially include a rinse fluid, an antibody labeled fluid, a rinse fluid, and an amplification fluid, all stored in the same. While the fluids are stored, they can be kept separated by substantially immiscible separation fluids (for example, a gas such as air) so that fluid reagents that must react normally with each other when in contact can be stored in a common channel .
[0095] The reagents can be stored in a cassette for several periods of time. For example, a reagent can be stored for more than 1 hour, for more than 6 hours, for more than 12 hours, for more than 1 day, for more than 1 week, for more than 1 month, for more than 3 months, for more than 6 months, for more than 1 year, or for more than 2 years. Optionally, the cassette can be treated in an appropriate manner in order to prolong storage. For example, the cassettes that store the reagents contained therein can be vacuum sealed, stored in a dark environment, and / or stored at low temperatures (for example, below 0 degrees Celsius). The duration of storage depends on one or more factors,
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44/101 such as the particular reagents used, the shape of the stored reagents (for example, wet or dry), the dimensions and materials used to form the substrate and cover layer (s), the adhesion method the substrate and the layer (s) of the cover, and how the cassette is treated or stored as a whole.
[0096] As illustrated in the example mode shown in figures 5 and 6, channels 206 and 207 cannot be in fluid communication with each other until fluid connector 220 is coupled to cassette 20. In other words, the two channels , in some modalities, they are not in fluid communication with each other before the first use and / or before the introduction of a sample in the cassette. In particular, as illustrated, the substantially U-shaped channel 222 of connector 220 can fluidly connect the first and second channels 206, 207 in such a way that reagents in the second channel 207 can pass through the U-shaped channel 22 and move selectively to measurement zones 209 on the first channel 206. In other embodiments, the two channels 206 and 207 are in fluid communication with each other before the first use, and / or before introducing a sample into the cassette , but the fluidic connector also connects the two channels (for example, to form a closed-loop system) with first use.
[0097] In some embodiments, a cassette described herein may include one or more microfluidic channels, although such cassettes are not limited to microfluidic systems and may relate to other types of fluidic systems. Microfluidic, as used herein, refers to a cassette, device, apparatus or system that includes at least one fluid channel that has a maximum cross-sectional dimension less than 1 mm, and a relationship between length and greatest cross-sectional dimension of at least 3: 1. A microfluidic channel, as used herein, is a channel that meets the requirement 870200007448, of 1/16/2020, p. 51/124
45/101 those criteria.
[0098] The cross-sectional dimension (for example, a diameter) of the channel is measured perpendicular to the direction of the fluid flow. Most of the fluid channels in the cassette components described here have dimensions in maximum cross-section less than 2 mm, and in some cases less than 1 mm. In a set of modalities, all fluid channels in a cassette are microfluidic or have a greater cross-sectional dimension of no more than 2 mm or 1 mm. In another set of modalities, the maximum cross-sectional dimension of the channel (s) is less than 500 microns, less than 200 microns, less than 100 microns, less than 50 microns, or less than 25 microns. In some cases the dimensions of the channel can be chosen in such a way that the fluid can flow freely through the article or the substrate. The dimensions of the channel can also be chosen, for example, to allow a certain volumetric or linear flow of fluid in the channel. Naturally, the number of channels and the format of the channels can be varied by any appropriate method known to those skilled in the art. In some cases, more than one channel or capillary can be used.
[0099] A channel can include a feature on or within an article (for example, a cassette) that at least partially directs the flow of a fluid. The channel can have any shape in appropriate cross section (circular, oval, triangular, irregular, square or rectangular, or another) and can be covered or uncovered. In the modalities where it is completely covered, at least part of the channel can have a cross section that is completely included, or the entire channel can be completely included along its entire length with the exception of its entrance (s) and output (s). A channel can also have an aspect ratio (length to dimension in average cross section) of at least 2: 1, plus tipiPetition 870200007448, from 1/16/2020, pg. 52/124
46/101 of at least 3: 1.5: 1, or 10: 1 or more.
[00100] The cassettes described here can include channels or channel segments positioned on one or two sides of the cassette. In some cases, channels are formed on one surface of the cassette. The channel segments can be connected by an intermediate channel that passes through the cassette. In some embodiments, channel segments are used to store reagents in the device prior to the first use by an end user. The specific geometry of the channel segments and the positions of the channel segments within the cassettes can allow fluid reagents to be stored for extended periods of time without mixing, even during routine handling of the cassettes, such as during transport of the cassettes, and when the cassettes are subjected to shock or physical vibration.
[00101] In certain embodiments, a cassette includes optical elements that are manufactured on one side of a cassette opposite a series of fluidic channels. An optical element is used to refer to a feature formed or positioned either in an article or on a cassette that is provided for and used to change the direction (for example, through refraction or reflection), the focus, the polarization, and / or another property of electromagnetic radiation incident in relation to the light incident on the article or the cassette in the absence of the element. For example, an optical element can comprise a lens (for example, concave or convex), a mirror, a grid, a groove, or another feature formed or positioned or on a cassette. A cassette itself without a unique characteristic, however, should not constitute an optical element, even if one or more properties of the incident light may change with the interaction with the cassette. Optical elements can guide the incident light that passes through the cassette in such a way that most of the light is scattered away
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47/101 of the specific areas of the cassette, such as intermediate parts between the fluid channels. By decreasing the amount of light incident on these intermediate parts, the amount of noise in a detection signal can be decreased when using certain optical detection systems. In some embodiments, the optical elements comprise triangular grooves formed on or within a surface of the cassette. The tracing angle of the triangular grooves can be chosen in such a way that the normal incident light on the surface of the cassette is redirected to an angle depending on the refractive indices of the external medium (eg air) and the material of the cassette. In one embodiment, one or more optical elements are positioned between adjacent segments of a winding region of a measurement zone.
[00102] A cassette can be made of any material suitable for forming a channel. Non-limiting examples of materials include polymers (e.g., polyethylene, polystyrene, polymethyl methacrylate, polycarbonate, poly (dimethylsiloxane), PTFE, PET, and a cycloolefin copolymer), glass, quartz, and silicon. The material that forms the cassette and any associated components (for example, a cover) can be hard or flexible. Those skilled in the art can immediately select the appropriate material (s) based, for example, on their rigidity, their inactivity (for example, freedom from degradation by) a fluid to be passed through ( s) same (s), its robustness at a temperature at which a particular device must be used, its transparency / opacity to light (for example, in the ultraviolet and visible regions), and / or the method used to provide characteristics in the material. For example, for injection molded or extruded articles, the material used can include a thermoplastic (eg polypropylene, polycarbonate, acrylonitrile butyrene-styrene, nylon 6), an elastomer (eg polyisoprene
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48/101 no, isobutene-isoprene, nitrile, neoprene, ethylene-propylene, hipalon, silicone), a thermoset (e.g., epoxy, unsaturated, phenolic polyesters), or combinations thereof.
[00103] In some embodiments, the material and dimensions (for example, the thickness) of a cassette and / or cover are chosen in such a way that they are substantially impermeable to water vapor. For example, a cassette designed to store one or more fluids in it prior to first use may include a cover that comprises a material that is known to provide a high vapor barrier, such as a sheet of metal, certain polymers, certain ceramics and combinations of these. In other cases, the material is chosen based at least in part on the format and / or configuration of the cassette. For example, certain materials can be used to form planar devices, while other materials are more suitable for forming devices that are curved or irregularly shaped.
[00104] In some examples, a cassette comprises a combination of two or more materials, such as those listed above. For example, the cassette channels can be formed of polystyrene or other polymers (for example, by means of injection molding) and a biocompatible tape can be used to seal the channels. The biocompatible tape or flexible material can include a material that is known to improve the vapor barrier properties (for example, a sheet of metal, polymers or other materials that are known to be endowed and high vapor barriers), and may allow optionally access to the inputs and outputs when punching or tearing the tape. A variety of methods can be used to seal a microfluidic channel or parts of a channel, or to join multiple layers of a device, including, but not limited to, the use of adhesives, the use of adhesive tapes, gluing, agglutination,
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49/101 lamination of materials, or by mechanical methods (for example, stapling, locking mechanisms, etc.).
[00105] In some examples, a cassette comprises a combination of two or more separate layers (or cassettes) mounted on each other. Independent channel networks (such as sections 71 and 77 of figure 1A), which can optionally include reagents stored in them prior to first use, can be included on separate layers (or cassettes). The separate layers can be assembled together by any suitable means, such as by the methods described herein, to form a single cassette. In some embodiments, two or more channel networks are fluidly connected on first use, for example, by using a fluidic connector. In other embodiments, the two or more channel networks are fluidly connected before first use.
[00106] A cassette described here can have any appropriate volume to perform an analysis such as a chemical and / or biological reaction or another process. The entire volume of a cassette includes, for example, all reagent storage areas, measurement zones, liquid containment regions, disposal areas, as well as any fluid connectors, and the fluid channels associated with them. In some embodiments, small amounts of reagents and samples are used and the entire volume of the fluidic device is, for example, less than 10 ml, 5 ml, 1 ml, 500 pl, 250 pl, 100 pl, 50 pl, 25 pl , 10 pl, 5 pl, or 1 pl. A cassette described here can be portable and, in some embodiments, carried by hand. The length and / or width of the cassette may, for example, be less than or equal to 20 cm, 15 cm, 10 cm, 8 cm, 6 cm, or 5 cm. The thickness of the cassette can be, for example, less than or equal to 5 cm, 3 cm, 2 cm, 1 cm, 8 mm, 5 mm, 3 mm, 2 mm, or 1 mm. Advantageously, portable devices may be suitable for use
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50/101 in care point configurations.
[00107] It should be understood that the cassettes and their respective components described here are exemplifiers and that other configurations and / or types of cassettes and components can be used with the systems and methods described here.
[00108] The methods and systems described herein can involve a variety of different types of analysis, and can be used to determine a variety of different samples. In some cases, an analysis involves a chemical and / or biological reaction. In some embodiments, a chemical and / or biological reaction involves bonding. Different types of connection can occur on the cassettes described here. Bonding may involve the interaction between a corresponding pair of molecules that exhibit mutual affinity or binding capacity, typically specific or non-specific binding or interaction, including biochemical, physiological, and / or pharmaceutical interactions. The biological bond defines a type of interaction that occurs between pairs of molecules that include proteins, nucleic acids, glycoproteins, carbohydrates, hormones, and others. Specific examples include antibody / antigen, antibody / hapten, enzyme / substrate, enzyme / inhibitor, enzyme / cofactor, binding protein / substrate, carrier protein / substrate, lecithin / carbohydrate, receptor / hormone, receptor / effector, complementary filaments of nucleic acid, nucleic acid protein / repressor / inducer, cell surface ligand / receptor, virus / ligand, etc. The bond can also occur between proteins or other components and cells. In addition, the devices described herein can be used for other fluid analysis (which may or may not involve binding and / or reactions) such as component suppression, concentration, etc.
[00109] In some cases, a heterogeneous reaction (or test) can occur on a cassette; for example, a liaison partner can
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51/101 be associated with a channel surface, and the complementary connection partner may be present in the fluid phase. Other solid-phase assays that involve the affinity reaction between proteins or other biomolecules (for example, DNA, RNA, carbohydrates), or molecules that occur unnaturally, can also be performed. Non-limiting examples of typical reactions that can be performed on a cassette include chemical reactions, enzymatic reactions, immunobased reactions (eg, antigen-antibody), and cell-based reactions.
[00110] Non-limiting examples of analytes that can be determined (for example, detected) when using the cassettes described herein include specific proteins, viruses, hormones, drugs, nucleic acids and polysaccharides; specifically antibodies, for example, IgD, IgG, IgM or IgA immunoglobulins for HTLV-I, HIV, hepatitis A, B and non-A / non-B, rubella, measles, Human Parvovirus B19, mumps, malaria, chickenpox or leukemia; human and animal hormones, for example, thyroid stimulating hormone (TSH), thyroxine (T4), luteinizing hormone (LH), follicle stimulating hormones (FSH), testosterone, progesterone, human chorionic gonadotropin, estradiol; other proteins or peptides, for example troponin I, c-reactive protein, myoglobin, brain natriuretic protein, prostate specific antigen (PSA), free PSA, complexed PSA, proPSA, EPCA-2, PCADM-1, ABCA5, hK2, beta-MSP (PSP94), AZGP1, Appendix A3, PSCA, PSMA, JM27, PAP; drugs, for example, paracetamol or theophylline; marker nucleic acids, for example, PCA3, TMPRS-ERG, polysaccharides such as cell surface antigens for the typing of HLA tissue and bacterial material from the cell wall. Chemicals that can be detected include explosives such as TNT, nerve agents, and environmentally harmful compounds such as polychlorinated biphenyls (PCBs), dioxi
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52/101 nas, hydrocarbons and MTBE. Typical sample fluids include physiological fluids such as human or animal whole blood, blood serum, blood plasma, semen, tears, urine, sweat, saliva, cerebrospinal fluid, vaginal secretions; in-vitro liquids used in research or environmental liquids such as aqueous liquids suspected of being contaminated by the test.
[00111] In some embodiments, one or more reagents that can be used to determine a sample test (for example, a test binding partner to be determined) are stored in a channel or in a cassette chamber before first use in order to carry out a test or a specific test. In cases where an antigen is being analyzed, an antibody or a corresponding aptamer can be the binding partner associated with a surface of a microfluidic channel. If an antibody is the analyte, then an appropriate antigen or aptamer can be the binding partner associated with the surface. When a disease condition is being determined, it may be preferable to place the antigen on the surface and test it for an antibody that is produced in the individual. Such antibodies can include, for example, antibodies to HIV.
[00112] In some modalities, a cassette is adapted and arranged to perform an analysis that involves the accumulation of an opaque material in a region of a microfluidic channel, the exposure of the region to light, and the determination of light transmission through the material opaque. An opaque material can include a substance that interferes with the transmittance of light at one or more wavelengths. An opaque material does not merely reflect light, but reduces the amount of transmission through the material, for example, by absorbing or reflecting light. Different opaque materials or different amounts of an opaque material can allow transmittance of less than, for example,
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53/101 example, 90, 80, 70, 60, 50, 40, 30, 20, 10 or 1 percent of the light that illuminates the opaque material. Examples of opaque materials include molecular layers of metal (for example, elemental metal), ceramic layers, polymeric layers, and layers of an opaque substance (for example, a dye). The opaque material may, in some cases, be a metal that can be deposited non-electrolytically. Such metals may include, for example, silver, copper, nickel, cobalt, palladium, and platinum.
[00113] An opaque material that forms in a channel can include a series of discontinuous independent particles that together form an opaque layer, but in one embodiment, it is a continuous material that takes on a generally planar shape. The opaque material can have a dimension (for example, an extension of the length), for example, greater than or equal to 1 micron, greater than or equal to 5 microns, greater than 10 microns, greater than or equal to 25 microns, or greater than or equal to 50 microns. In some cases, the opaque material extends across the width of the channel (for example, a measurement zone) containing the opaque material. The opaque layer may have a thickness, for example, less than or equal to 10 microns, less than or equal to 5 microns, less than or equal to 1 micron, less than or equal to 100 nanometers or less than than or equal to 10 nanometers. Even at these small thicknesses, a detectable change in transmittance can be achieved. The opaque layer can provide an increase in the sensitivity of the assay when compared to techniques that do not form an opaque layer.
[00114] In a set of modalities, a cassette described here is used to perform an immunoassay (for example, for human IgG or PSA) and optionally uses silver enhancement for signal amplification. In such an immunoassay, after applying a sample containing human IgG to a reaction site or an analysis site, the
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54/101 link between human IgG and anti-human IgG can occur. One or more reagents, which can optionally be stored in a channel of the device before use, can then flow over that binding pair complex. One of the stored reagents can include a metal colloid solution (for example, a gold-conjugated antibody) that specifically binds to the antigen to be detected (for example, human IgG). This metal colloid can provide a catalytic surface for the deposition of an opaque material, such as a layer of metal (for example, silver), on a surface in the region of analysis. The metal layer can be formed using a two-component system: a metal precursor (for example, a solution of silver salts) and a reducing agent (for example, hydroquinone, chloro-hydroquinone, pyrogallol, methanol, 4- aminophenol and phenidone), which can optionally be stored in different channels before use.
[00115] Once a positive or negative pressure differential is applied to the system, the silver salt and reduction solutions can merge at a channel intersection, where they mix (for example, due to diffusion) in a channel , and then flow over the analysis region. Therefore, if antibody-antigen binding occurs in the region of analysis, the flow of the metal precursor solution through the region may result in the formation of an opaque layer, such as a silver layer, due to the presence of the associated catalytic metal colloid. with the antibody-antigen complex. The opaque layer may include a substance that interferes with the transmittance of light at one or more wavelengths. An opaque layer that is formed in the channel can be detected optically, for example, by measuring a reduction in light transmittance across a part of the analysis region (for example, a sinuous channel region) compared to a part of an area which does not include the antibody or antigen. Al
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55/101 ternatively, a signal can be obtained by measuring the change in light transmittance as a function of time, since the film is being formed in an analysis region. The opaque layer can provide an increase in the sensitivity of the assay when compared to techniques that do not form an opaque layer. In addition, several amplification chemicals that produce optical signals (for example, absorbance, fluorescence, glow or flash chemiluminescence, electrochemiluminescence), electrical signals (for example, the electrical resistance or conductivity of metal structures created by a non-electrolytic process ) or magnetic signals (for example, magnetic granules) can be used to allow the detection of a signal by a detector.
[00116] Various types of fluids can be used with the cassettes described here. As described herein, fluids can be introduced into the cassette on first use, and / or be stored inside the cassette before first use. Fluids include liquids such as solvents, solutions and suspensions. Fluids also include gases and gas mixtures. When multiple fluids are contained in a cassette, the fluids can be separated by another fluid which is preferably substantially immiscible in each of the first two fluids. For example, if a channel contains two different aqueous solutions, a buffer for separating a third fluid can be substantially immiscible in both aqueous solutions. When aqueous solutions are to be kept separate, the substantially immiscible fluids that can be used as separators may include gases such as air or nitrogen, or hydrophobic liquids that are substantially immiscible with the aqueous fluids. Fluids can also be chosen based on fluid reactivity with adjacent fluids. For example, an inert gas such as nitrogen can be used in some modalities and can
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56/101 help to preserve and / or stabilize all adjacent fluids. An example of a substantially immiscible fluid for separating aqueous solutions is perfluorodecalin. The choice of a fluid separator can also be made based on other factors, including any effect that the fluid separator may have on the surface tension of the adjacent fluid plugs. It may be preferable to maximize the surface tension within any fluid plug to promote retention of the fluid plug as a single continuous unit under varying environmental conditions such as variations in vibration, shock and temperature. The separator fluids can also be inert to a reaction site (for example, the measurement zone) to which the liquids will be provided. For example, if a reaction site includes a biological binding partner, a fluid separator such as air or nitrogen can have almost no effect on the binding partner. The use of a gas (for example, air) as a fluid separator can also provide space for expansion within a channel of a fluidic device if the liquids contained in the device expand or contract due to changes such as temperature variations (including freezing) or pressure.
[00117] As described here, a cassette can be configured to operate with an analyzer in some modalities. For example, the cassette shown illustratively in Figure 5 can have a cam surface along a side of the cassette. In this particular embodiment, the cam surface includes a notch 230 formed at one end of the cassette. The other end of the cassette includes a curved surface 232. That cam surface of the cassette can be configured to interact with a sample analyzer in such a way that the analyzer can detect the presence of the cassette inside the analyzer casing and / or position the cassette inside the analyzer.
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57/101
[00118] Figure 7 shows an example of a 301 analyzer that can be configured to receive a cassette. The analyzer can include a fluid flow source 40 (for example, a pressure control system) that can be fluidly connected to channels 206, 207, 222 (for example, from figure 6) to pressurize the channels so moving the sample and / or other reagents through the channels. In particular, the fluid flow source 40 can be configured to initially move a sample and / or a reagent from the substantially U-shaped channel 222 to the first channel 206. The fluid flow source 40 can also be used to move the reagents in the second channel 207 through the substantially U-shaped channel 222 and to the first channel 206. After the sample and reagents pass through measurement zones 209 and are analyzed, the fluid flow source 40 can be configured to move fluids into the absorbent material 217 of cassette 200. In one embodiment, the source of fluid flow is a vacuum system. It should be understood, however, that other sources of fluid flow such as valves, pumps, and / or other components can be used. The 301 analyzer can be used in a variety of ways to process and analyze a sample placed inside the analyzer. In a particular embodiment, since a mechanical component configured to interface with the cassette indicates that cassette 20 is loaded correctly in analyzer 301, the identification reader reads and identifies the information associated with cassette 20. Analyzer 301 can be configured to compare the information with the data stored in a control system to ensure that you have the calibration information for this particular sample (such as a calibration curve or predicted values for any measurements made during a test). If the analyzer does not have the appropriate calibration information, the analyzer can send a request to the user
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58/101 to load the specific information needed. This information can be loaded by using, for example, the same identification reader that reads information from the cassette. It could also be loaded using a separate ID reader or by some other method. The analyzer can also be configured to review the expiration date information associated with the cassette and to cancel the analysis if the expiration date has expired.
[00119] In one embodiment, once the analyzer has determined that the cassette can be analyzed, a fluid flow source such as a vacuum distributor can be configured to contact the cassette to ensure a fluid impermeable seal around a vacuum port and cassette exhaust ports. In one embodiment, an optical system can take initial measurements to obtain reference readings. Such reference readings can be taken with both light sources (eg 82, 86 in figure 7) enabled and disabled.
[00120] To initiate the movement of the sample, the fluid flow source 40 (for example, a vacuum system) can be activated, which can quickly change the pressure within channel 206, 207 (for example, reduced to about -30kPa). This pressure reduction within the channel can direct the sample to channel 206 and through each of the 209A-209D measurement zones (see figure 6). After the sample reaches the final measurement zone 209D, the sample can continue to flow into the liquid containment region 217.
[00121] In a particular embodiment, the sample 301 microfluidic analyzer is used to measure the level of a prostate specific antigen (PSA) in a blood sample. In this mode, four measurement zones 209A-209D can be used to analyze the sample. For example, in a first measurement zone, the channel walls can be blocked with a block protein
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59/101 chemo (such as bovine serum albumin) in such a way that little or protein in the blood sample binds to the walls of measurement zone 209 (with the exception perhaps of some non-specific binding that can be eliminated). This first measurement zone can act as a negative control.
[00122] In a second measurement zone 209, the walls of channel 206 can be coated with a large predetermined amount of a prostate specific antigen (PSA) to act as an elevated or positive control. Due to the fact that the blood sample passes through the second measurement zone 209, little or no PSA protein in the blood can bind to the channel walls. The gold-conjugated signal antibodies in the sample can be dissolved within the fluid connector tube 222 or can be fluid from any other appropriate location. These antibodies cannot yet be limited to the PSA in the sample, and thus can bind to the PSA on the channel walls to act as an elevated or positive control.
[00123] In a third measurement zone 209, the walls of channel 206 can be coated with a small predetermined amount of PSA to act as a low control. As the blood sample flows through this measurement zone 209, no PSA protein in the sample binds to the channel wall. Antibodies to gold-conjugated signals in the sample can be dissolved inside the 222 fluid connector tube (which are not yet attached to the PSA in the sample) or can be fluid from any other appropriate location, and can bind to the PSA on the channel walls to act as a low control.
[00124] In a fourth measurement zone 209, the walls of channel 206 can be coated with the capture antibody, an anti-PSA antibody, which binds to a different epitope on the PSA protein of the
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60/101 that the gold-conjugated signal antibody. As the blood sample flows through the fourth measurement zone, the PSA proteins in the blood sample can bind to the anti-PSA antibody in a way that is proportional to the concentration of these proteins in the blood. Thus, in one embodiment, the first three measurement zones 209 can act as controls and the fourth measurement zone 209 can actually test the sample. In other embodiments, different numbers of measurement zones can be provided, and an analysis can optionally include including more than one measurement zone that actually tests the sample.
[00125] In some cases, measurements from a region that analyzes the sample (for example, the fourth measurement zone described above) can be used to determine not only the concentration of an analyte in a sample, but also as a control . For example, a limit measurement can be established at an early stage of amplification. Measurements above this value (or below this value) may indicate that the concentration of the analyte is outside the desired range for the assay. This technique can be used to identify, for example, if a high dose Hook effect is occurring during the analysis, that is, when a very high concentration of analyte provides a low artificial reading.
[00126] In other modalities, different numbers of measurement zones can be provided, and an analysis can optionally include more than one measurement zone that actually tests the sample. Additional measurement zones can be used to measure additional analytics so that the system can run multiple runs simultaneously on a single sample.
[00127] In a particular embodiment, a 10 microliter blood sample takes about eight minutes to flow through the four 209 measurement zones. The beginning of this analysis can be calculated
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61/101 when the pressure inside channel 206 is about -30kPa. During this time, the optical system 80 is measuring the light transmission for each measurement zone, and in one mode these data can be transmitted to a control system about every 0.1 second. When using reference values, these measurements can be converted using the following formulas:
[00128] Transmission = (l-ld) / (lr-ld) (1)
[00129] Where:
[00130] l = the intensity of the light transmitted through a measurement zone at a given point in time
[00131] ld = the intensity of the light transmitted through a measurement zone with the light source deactivated
[00132] lr = a reference intensity (that is, the intensity of the light transmitted in a measurement zone with the light source activated, or before the beginning of an analysis when only the air is in the channel and
[00133] Optical density = - log (Transmission) (2)
[00134] In this way, when using these formulas, the optical density in a 209 measurement zone can be calculated.
[00135] As described herein, a variety of methods can be used to control the flow of fluid in a cassette, including the use of pumps, voids, valves, and other components associated with an analyzer. In some cases, fluid control can also be performed at least in part by one or more components within the cassette, such as when using a valve positioned inside the cassette, or when using specific fluids and channel configurations with the cassette. In a set of modalities, fluid flow control can be achieved based at least in part on the influence of the channel geometry and the viscosity of one or more fluids (which can be stored) within the cassette.
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[00136] One method includes flowing a low fluid viscosity plug and a high fluid viscosity plug into a channel that includes a flow constriction region and a non-constriction region. In one embodiment, the low viscosity fluid flows at a first flow in the channel and the flow is not substantially affected by the fluid flowing in the flow constriction region. When the high viscosity fluid flows from the non-constricted region to the flow constricted region, fluid flow rates decrease substantially, since flow rates, in some systems, are influenced by the higher viscosity fluid flowing in the smaller section area system (for example, the flow constriction region). This causes the low viscosity fluid to flow at a second flow slower than its original flow, for example, at the same flow rate at which the high viscosity fluid flows in the flow constriction region.
[00137] For example, a method of fluid flow control may involve the flow of a first fluid from a first part of the channel to a second part of the channel in a microfluidic system, in which a fluid passage defined by the first part of the channel has a cross-sectional area larger than a cross-sectional area of a fluid passage defined by the second part of the channel, and the flow of a second fluid into a third part of the channel in the microfluidic system in fluid communication with the first and second channel parts, where the viscosity of the first fluid is different from the viscosity of the second fluid, and where the first and second fluids are substantially incompressible. Without stopping the first or second fluids, a volumetric flow rate of the first and second fluids can be decreased by a factor of at least 3, at least 10, at least 20, at least 30, at least 40, or at least 50 in the microfluidic system as a result of the flow of the first fluid from the first part of the channel to the second part of the channel, in comparison
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63/101 tion to the absence of the flow of the first fluid from the first part of the channel to the second part of the channel. A chemical and / or biological interaction involving a component of the first or second fluids can occur in a first measurement zone in fluid communication with parts of the channel when the first and second liquids are flowing at reduced flow.
[00138] Therefore, when designing microfluidic systems with flow constriction regions positioned in particular locations and choosing appropriate fluid viscosities, a fluid can be caused to accelerate or decelerate at different locations within the system without the use of valves and / or without external control. In addition, the length of the channel parts can be chosen to allow a fluid to remain in a particular area of the system for a certain period of time. Such systems are particularly useful for carrying out chemical and / or biological tests, as well as other applications where the timing of the reagents is important.
[00139] Any appropriate fluid flow source of flow can be used to promote or maintain fluid flow in a microfluidic system or cassette described herein. In some cases, the fluid flow source is part of a microfluidic sample analyzer. A fluid flow can be configured to pressurize a channel in a cassette to move a sample through the channel. In an illustrative embodiment, the fluid flow source is a vacuum system and includes a vacuum source or pump, two vacuum reservoirs that can be separated by a vacuum regulator, and a distributor to provide a fluid connection between the reservoirs vacuum and the cassette. The dispenser 48 may also include one or more fluid connections to one or more ports on the cassette. For example, the distributor can provide a fluid connection between a port and a valve.
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64/101 la (such as a solenoid valve). The opening and closing of this valve can control where air can enter the cassette, thereby serving as a breathing valve in certain modalities.
[00140] As mentioned above, in one embodiment, the vacuum source is a pump, such as a solenoid-operated diaphragm pump. In other embodiments, the fluid flow can be directed / controlled through the use of other types of pumps or fluid flow sources. For example, in one embodiment, a syringe pump can be used to create a vacuum by pulling the syringe plunger in an outward direction. In other embodiments, positive pressure is applied to one or more cassette inlets to provide a source of fluid flow.
[00141] In some embodiments, fluid flow occurs by applying a substantially constant non-zero pressure drop (ie, AP) through an inlet and outlet of a cassette. In a set of modalities, an entire analysis is performed by applying a substantially constant non-zero pressure drop (ie, AP) across a cassette inlet and outlet. A substantially constant non-zero pressure drop can be achieved, for example, by applying positive pressure at the inlet or reduced pressure (for example, a vacuum) at the outlet. In some cases, a substantially constant non-zero pressure drop is achieved when the fluid flow does not occur predominantly by capillary forces and / or without the use of activating valves (for example, without changing an area in cross section of an air channel). a fluid passage from the cassette). In some embodiments, during essentially the entire analysis done on the cassette, a substantially constant non-zero pressure drop may be present, for example, through a
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65/101 input to a measurement zone (which can be connected to a fluidic connector) and an output downstream of the measurement zone (for example, an output downstream of a liquid containment region), respectively.
[00142] In one embodiment, a vacuum source is configured to pressurize a channel to about -60kPa (about 2/3 atmosphere). In another embodiment, the vacuum source is configured to pressurize a channel to about -30kPa. In certain embodiments, the vacuum sources are configured to pressurize a channel, for example, between -100kPa and -70kPa, between -70kPa and -50kPa, between -50kPa and -20kPa, or between -20kPa and -1 kPa.
[00143] As mentioned above, in one embodiment, two vacuum reservoirs can be provided. The pump can be activated in such a way that the first reservoir can be pressurized to about -60kPa. A regulator positioned between the reservoirs can ensure that the second reservoir can only be pressurized up to a different pressure, for example, around -30kPa. This regulator can maintain the reservoir pressure at -30kPa (or at another appropriate pressure) as long as the reservoir remains within a certain pressure range, for example, between -60kPa and -30kPa. Pressure sensors can monitor the pressure within each reservoir. If the pressure in the first reservoir reaches a stipulated point (for example, about -40kPa), the pump can be activated to decrease the pressure in the first reservoir. The second reservoir can be configured to detect any leaks in the total vacuum system. Optionally, the vacuum system 40 can include a filter coupled to the reservoirs. A solenoid valve can serve as a breathing valve connected through the distributor to the port.
[00144] In certain modalities, the cassette is positioned
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66/101 inside a analyzer, a fluid flow source that is part of the analyzer can be attached to the cassette to ensure an impermeable connection to fluids. For example, cassette 20 may include a port configured to couple a channel of the cassette with the fluid source. In one embodiment, seals, or O-rings are positioned around the door and a linear solenoid can be positioned above the O-rings to press and seal the O-rings against the cassette body. A distributor adapter can be positioned between the linear solenoid and the distributor, and passive return springs can be provided around the distributor to move the distributor away from the cassette body when the solenoid is not loaded. In one embodiment, multiple ports on the cassette can form an interface with the distributor. For example, in addition to a port for inserting and / or removing reagents, the cassette can include one or more exhaust ports and / or a mixing port. The interface between each port and the distributor can be independent (for example, there may be no fluid connection inside the distributor).
[00145] In one embodiment, when the fluid flow source is activated, one or more channels in the cassette can be pressurized (for example, up to about -30kPa) which will direct the fluids within the channel (for example, both fluid samples as well as reagents) to the outlet. In an embodiment that includes an exhaust port and a mixing port, a breathing valve connected to the exhaust port via a distributor may be initially open, which can allow all reagents downstream of a mixing port to move to the outlet, but it will not cause the reagents upstream of the mixing port to move. Once the breathing valve is closed, the reagents upstream of the mixing port can move to a mixing port and then to the outlet. For example, fluids can be stored
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67/101 in series in a channel upstream of the mixing port, and after a breathing valve positioned along the channel has been closed, fluids can flow sequentially to the outlet of the channel. In some cases, fluids can be stored in separate intercepting channels and, after a breathing valve has been closed, fluids will flow together to an intersection point. This set of modalities can be used, for example, to mix the fluids in a controllable way as they flow together. The timing of the distribution and the volume of fluid delivered can be controlled, for example, by the timing of activation of the breathing valve.
[00146] Advantageously, the breathing valves can be operated without constricting the cross section of the microfluidic channel in which they operate, as it should occur with certain valves in the prior art. Such a mode of operation can be effective in preventing leaks through the valve. In addition, due to the fact that breathing valves can be used, some systems and methods described here do not require the use of certain internal valves, which can be problematic due, for example, to their high cost, complexity in manufacturing, fragility, limited compatibility with mixed gas and liquid systems, and / or non-reliability in microfluidic systems.
[00147] It should be understood that, although breathing valves are described, other types of valve mechanisms can be used with the systems and methods described herein. Non-limiting examples of a valve mechanism that can be operatively associated with a valve include a diaphragm valve, a ball valve, a gate valve, a butterfly valve, a globe valve, a needle valve, a pinch valve, a trigger valve, or a pinch valve. THE
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68/101 valve mechanism can be activated by any appropriate means, including a solenoid, a motor, by hand, by electronic activation, or by hydraulic / pneumatic pressure.
[00148] As previously mentioned, all liquids in the cassette (eg, sample and reagents) can move into the liquid containment area which may include an absorbent material. In one embodiment, the absorbent material absorbs only liquids, so that gases can flow out of the cassette through the outlet.
[00149] A variety of determination techniques (for example, measurement, quantification, detection and qualification) can be used, for example, to analyze a component of the sample or another component or condition associated with a microfluidic system or cassette described herein. . Determination techniques can include optically based techniques such as light transmission, light absorbance, light scattering, light reflection and visual techniques. Determination techniques can also include luminescence techniques such as photoluminescence (for example, fluorescence, chemiluminescence, bioluminescence, and / or electrochemiluminescence. In other embodiments, the determination techniques can measure conductivity or resistance. , an analyzer can be configured to include such appropriate detection systems and others.
[00150] Different optical detection techniques provide a number of options for determining reaction results (eg, assay). In some embodiments, measurement of transmission or absorbance means that light can be detected at the same wavelength as it is emitted from a light source. Although the light source can be a narrow band source that emits a single wavelength it can also be a wide
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69/101 spectrum, emitting in a range of wavelengths, since many opaque materials can effectively block a wide range of wavelengths. In some embodiments, a system can be operated with a minimum of optical devices (for example, a simplified optical detector). For example, the determination device can be free of a photomultiplier, it can be free of a wavelength selector such as a grid, a prism or a filter, it can be free of a device to direct or collimate light such as a collimator, or it can be free of an optical magnification system (for example, lenses). Eliminating or reducing these characteristics can result in a less expensive and more robust device.
[00151] In a set of modalities, an exemplary optical system is positioned in the housing of an analyzer. As shown illustratively in Figure 7, the optical system includes at least a first light source 80 and a detector 84 spaced from the first light source. The first light source 82 can be configured to pass light through a first measuring zone of the cassette 20 when the cassette is inserted into the analyzer 301. The first detector 84 can be positioned opposite the first light source 82 to detect the amount of light that passes through the first measuring zone of the cassette. In a particular embodiment, the optical system includes ten light sources and ten detectors. It should be appreciated that, in other embodiments, the number of light sources and detectors may vary, since the invention is not so limited. As described herein, the cassette can include a plurality of measurement zones and the cassette 20 can be positioned within the analyzer in such a way that each measurement zone aligns with a light source and a corresponding detector. In some embodiments, the light source includes an optical aperture that can help direct light from the light source to a region
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70/101 particular within a measuring zone of the cassette.
[00152] In one embodiment, the light sources are light-emitting diodes (LEDs) or laser diodes. For example, a red semiconductor laser diode of InGaAlP that emits at 654 nm can be used. Other light sources can also be used. The light source can be positioned inside a nest or enclosure. The nest or shell may include a narrow opening or a thin tube that can help to collimate the light. The light sources can be positioned above where the cassette is inserted in the analyzer in such a way that the light source shines down on the upper surface of the cassette. Other appropriate configurations of the light source with respect to the cassette are also possible.
[00153] It should be appreciated that the wavelength of the light sources can vary, since the invention is not so limited. For example, in one embodiment, the wavelength of the light source is about 670 nm, and in another embodiment, the wavelength of the light source is about 650 nm. It should be appreciated that, in one embodiment, the wavelength of each light source can be different in such a way that each measuring zone of the cassette receives a different wavelength of light. In a particular mode when measuring hematocrit or hemoglobin, an isobesic wavelength range between about 590 nm and about 805 nm can be used for at least one of the measurement zones.
[00154] As mentioned, a detector 84 can be spaced and positioned below a light source to detect the amount of light that passes through the cassette. In one embodiment, one or more of the detectors are photodetectors (for example, photodiodes). In certain embodiments, the photodetector can be any appropriate device with the ability to detect the transmission of light that is
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71/101 emitted by the light source. One type of photodetector is an optical integrated circuit (IC) that includes a photodiode that has a peak sensitivity at 700 nm, an amplifier and a voltage regulator. The detector can be positioned inside a nest or enclosure that can include a narrow opening or a thin tube to ensure that only the light from the center of the measurement zone is measured in the detector. As described in more detail below, if the light source is pulse-modulated, the photodetector can include a filter to remove the effect of light that is not in the selected frequency. When multiple signals and neighbors are detected at the same time, the light source used for each measurement zone (for example, the detection region) can be modulated at a frequency sufficiently different from that of its neighboring light source. In this configuration, each detector can be configured (for example, when using software) to select its assigned light source, thereby avoiding light interference from neighboring optical pairs.
[00155] As described herein, a cassette may include a measurement zone that includes a sinuous channel configured and arranged to align with a detector in such a way that, with alignment, the detector can measure a single signal through more than one adjacent segment of the sinuous channel. In some embodiments, the detector can detect at least one signal within a part of the sinuous channel area and across more than one segment of the sinuous channel in such a way that a first part of the signal, measured from a first channel segment sinuous, is similar to a second part of the signal, measured from a second segment of the sinuous channel. In such modalities, due to the fact that the signal is present as a part of more than one segment of the sinuous channel, there is no need for precise alignment between a detector and a measurement zone.
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[00156] Positioning the detector over the measurement zone (for example, a winding region) without the need for precision is an advantage, since external (and possibly expensive) equipment, such as microscopes, lenses and stages alignment are not required (although they can be used in certain modalities). Instead, alignment can be performed by low-cost methods that do not necessarily require an active or separate step from alignment by the user. For example, in one embodiment, a cassette comprising a winding region can be placed in a notch on an analyzer described here (for example, in a cavity that is identical or similar in shape to the cassette), and the measurement zone can automatically be located in a beam of light from the detector. The possible causes of misalignment caused, for example, by variations from cassette to cassette, the exact position of the cassette in the notch, and the normal use of the cassette, can be insignificant compared to the dimensions of the measurement zone. As a result, the sinuous region can remain within the light beam and the detection is not interrupted due to these variations.
[00157] The detector can detect a signal within all or part of a measurement zone (for example, which includes a winding region). In other words, different amounts of the sinuous region can be used as an optical detection pathway. For example, the detector can detect a signal within at least 15% of the measurement zone, at least 20% of the measurement zone, at least 25% of the measurement zone, within at least 50% of the measurement zone, or within at least 75% of the measurement zone (but less than 100% of the measurement zone). The area in which the measurement zone is used as an optical detection passage can also depend, for example, on the opacity of the material in which the channel is
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73/101 manufactured (for example, if all or part of the channel is transparent), the amount of a non-transparent material that can cover a part of the channel (for example, through the use of a protective cover), and / or the size of the detector and the measurement zone.
[00158] In one mode, a signal produced by a reaction performed on the cassette is homogeneous throughout the measurement zone (for example, over an entire region of the sinuous channel). That is, the measurement zone (for example, the region of the sinuous channel) can allow the production and / or detection of a single homogeneous signal in said region with the execution of a chemical and / or biological reaction (for example, and detection by a detector). Before executing a reaction in the region of the sinuous channel, the sinuous channel can include, for example, a single species (and concentration of the species) to be detected / determined. The species can be adsorbed onto a winding channel surface. In another embodiment, the signal can be homogeneous only in parts of the winding region, and one or more detectors can detect different signals within each part. In certain examples, more than one measurement zone can be connected in series and each measurement zone can be used to detect / determine a different species. It should be understood that, although winding regions are described, measurement zones that do not include winding regions can also be used.
[00159] The applicant has recognized that the amount of light transmitted through a measuring zone of the cassette can be used to determine information about not only the sample, but also information about specific processes that occur in the fluidic system of the cassette (for example , mixing of reagents, flow, etc.). In some cases, light measurement across a region can be used as feedback to control the flow of fluid in the system, as described here.
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[00160] In some cases, the optical density of a fluid is determined.
[00161] It must be recognized that a transparent liquid (such as water) can allow a large amount of light to be transmitted from the light source, through a measurement zone and towards a detector. The air within the measurement zone can lead to less light transmitted through the measurement zone because more light can be dispersed within the channel compared to when a clear liquid is present. When a blood sample is in a measurement zone, significantly less light can pass through the same path to the detector due to light scattering out of the blood cell and also due to absorbance. In one embodiment, silver is associated with a sample component attached to a surface within the measurement zone and as the silver accumulates within the zone less and less light is transmitted through the measurement zone.
[00162] It must be recognized that measuring the amount of light that is detected in each detector allows a user to determine which reagents are in a particular measurement zone at a particular point in time. It must also be recognized that, by measuring the amount of light that is detected with each detector, it is possible to measure the amount of silver deposited in each measurement zone. This amount can correspond to the amount of analyte captured during a reaction, which can thus provide a measure of the analyte concentration in the sample.
[00163] As described herein, the applicant has recognized that an optical system can be used for a variety of quality control reasons. First, the time it takes for a sample to reach a measurement zone where the optics detect the light that passes through the measurement zone can be used to determine whether
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75/101 there is a leak or an obstruction in the system. In addition, when the sample is expected to be at some volume, for example, about 10 microliters, there is a predicted flow time with which it must be associated for the sample to pass through the channels and measurement zones. If the sample falls outside this predicted flow time, it may be an indication that there is not enough sample to perform the analysis and / or that the wrong type of sample has been loaded into the analyzer. In addition, a predicted range of results can be determined based on the type of sample (eg, serum, blood, urine, etc.), and if the sample is outside the predicted range it can be an indication of an error.
[00164] In one embodiment, an optical system includes a plurality of light sources 82 and a plurality of corresponding detectors. In one embodiment, a first light source is adjacent to a second light source, where the first light source is configured to pass light through a first measuring zone on a cassette and the second light source is configured to pass the light through a second measuring zone of the cassette. In one embodiment, the light sources are configured in such a way that the second light source is not activated unless the first light source is disabled. The applicant has recognized that some light from a light source may spread to an adjacent detector and may affect the amount of light detected in the adjacent detector. In a set of modes, if the adjacent light source is activated at the same time as the first light source, then both detectors are also measuring the amount of light that passes through the first and second measuring zones of the cassette at the same time. , which can lead to inaccurate measurements.
[00165] Thus, in a set of modalities, the plurality of light sources is configured to activate sequentially with
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76/101 only one light source activated at a time. The corresponding detector for the activated light source is thus detecting only the amount of light that passes through the corresponding measurement zone. In a particular mode, each light source is configured to activate for a short period of time (for example, at least about 500, 250, 100, or 50 microseconds, or, in some modes, less than or equal to about 500, 250, 100 or 50 microseconds), and then an adjacent light source is configured to activate for a similar time frame. Activation for 100 microseconds corresponds to a rate of 10 kHz. In one embodiment, a multiplexed analog to digital converter is used to pulse the light and measure the amount of light detected in each detector corresponding to every 500, 250, 100 or 50 microseconds. Pulsing the light in this way can help prevent scattered light passing through a measurement zone from altering the amount of detected light that passes through an adjacent measurement zone.
[00166] Although there may be some benefits associated with the pulsation of the light sources in the manner described above, it must be recognized that the invention is not so limited and that other arrangements may be possible, such as where multiple light sources can be activated by Same time. For example, in one embodiment, light sources that are not directly adjacent to each other can be activated substantially simultaneously.
[00167] In one embodiment, an analyzer includes a temperature regulator system positioned inside the enclosure that can be configured to regulate the temperature inside the analyzer. For a given sample analysis, the sample may have to be kept within some temperature range. For example, in one embodiment, it is desirable to maintain the temperature inside the analyzer at about 37 ° C. Therefore, in one embodiment, the regulatory system
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77/101 temperature includes a heater configured to heat the cassette. In one embodiment, the heater is a resistive heater that can be positioned on the underside of where the cassette is placed in the analyzer. In one embodiment, the temperature regulator system also includes a thermistor to measure the temperature of the cassette and a control circuit can be provided to control the temperature. [00168] In one embodiment, the passive air flow inside the analyzer can act to cool the air inside the analyzer if necessary. A fan (not shown) can optionally be provided on the analyzer to lower the temperature inside the analyzer. In some embodiments, the temperature regulating system may include Peltier thermoelectric heaters and / or coolers within the analyzer.
[00169] In certain embodiments, an identification system that includes one or more identifiers is used and associated with one or more components or materials associated with a cassette and / or an analyzer. Identifiers, as described in more detail below, can themselves be encoded with information (that is, carry or contain information, such as by using an information carrier, storage, generator or conductor device such as a tag or code radio frequency identification (RFID) bars on the component that include the identifier, or may themselves not be encoded with information about the component, but instead can be associated only with information that can be contained, for example, in a database on a computer or in a medium that can be read by computer (for example, information about a user and / or the sample to be analyzed). In the latter case, the detection of such an identifier can activate the retrieval and use of the associated information from the database.
[00170] The coded identifiers with information about a
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78/101 component need not necessarily be encoded with a complete set of information about the component. For example, in certain embodiments, an identifier can be encoded with information merely sufficient to allow a unique identification of the cassette (for example, in relation to a serial number, part number, etc.), while information additional items that are related to the cassette (for example, type, use (for example, trial type), property, location, position, connectivity, content, etc.) can be stored remotely and be associated with the identifier only.
[00171] Information about or information associated with a cassette, material or component, etc., is information regarding the identity, positioning, or position of the cassette, material or component or identity, positioning, or position of the contents of a cassette, material or component and may additionally include information regarding the nature, state or composition of the cassette, material, component or content. Information about or information associated with a cassette, material or component or its contents may include information that identifies the cassette, material or component or its content and distinguishes the cassette, material, component or its contents from others. For example, information about or information associated with a cassette, material or component or its contents may refer to information that indicates the type or what the cassette, material or component or its contents are, where it is or should be. found, how it is or should be positioned, the function or purpose of the cassette, the material or the component or its contents, such as the cassette, the material or the component or its contents must be connected with other system components, the batch number , the origin, the calibration information, the date of
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79/101 validity, destination, manufacturer or ownership of the cassette, material or component or its contents, the type of analysis / test to be performed on the cassette, information on whether the cassette was used / analyzed, etc.
[00172] In a set of modalities, an identifier is associated with a cassette and / or an analyzer described here. In general, as used herein, the term identifier refers to an item with the ability to provide information about the cassette and / or the analyzer (for example, information that includes one or more of the identity, location, or the position / positioning of the cassette and / or the analyzer or a component thereof) with which the identifier is associated or in which it is installed, or which can be identified or detected and the identification or detection event is associated with information about the cassette and / or the analyzer with which the identifier is associated. Non-limiting examples of identifiers that can be used in the context of the invention include radio frequency identification (RFID) tags, bar codes, serial numbers, colored tags, fluorescent or optical tags (for example, when using quantum dots) , chemical compounds, radio tags, magnetic tags, among others.
[00173] In one embodiment, an analyzer may include an identification reader 60 positioned within the housing configured to read information about the cassette. Any appropriate identification reader that can be used to read information from an identifier. Non-limiting examples of identification readers include RFID readers, barcode scanners, chemical detectors, cameras, radiation detectors, magnetic or electrical field detectors, among others. The detection / reading method and the appropriate type of identification detector depend on the particular identifier used and may include, for example, optical imaging,
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80/101 excitation and fluorescence detection, mass spectrometry, nuclear magnetic resonance, sequential arrangement, hybridization, electrophoresis, spectroscopy, microscopy, etc. In some embodiments, identification readers can be mounted or pre-embedded in specific locations (for example, on or inside a cassette and / or an analyzer).
[00174] In one embodiment, the identification reader is an RFID reader configured to read an RFID identifier associated with the cassette. For example, in one embodiment, the analyzer includes an RFID module and an antenna that are configured to read information from the cassette inserted in the analyzer. In another embodiment, the identification reader is a barcode reader configured to read a barcode associated with the cassette. Once the cassette is inserted into the analyzer, the identification reader can read information from the cassette. The identifier on the cassette can include one or more types of information such as the type of the cassette, the type of analysis / test to be performed, the batch number, the information on whether the cassette was used / analyzed, and other information described here. The reader can also be configured to read information provided with a group of cassettes, such as in a cassette box, such as, but not limited to, calibration information, expiration date, and any additional information specific to that batch. The identified information can be optionally displayed to a user, for example, to confirm that a correct cassette and / or test type is being performed.
[00175] In some cases, the identification reader can be integrated with a control system via communication channels. Communication between the ID readers and the control system can occur over a wired network or can be transmitted wirelessly. In one mode, the control system can be programmed
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81/101 to recognize a specific identifier (for example, a cassette associated with information that is related to a type of cassette, a manufacturer, a test to be performed, etc.) as indicating that the cassette is properly connected or inserted into a particular type of analyzer.
[00176] In one embodiment, the identifier of a cassette is associated with the predetermined or programmed information contained in a database regarding the use of the system or the cassette for a particular purpose, user or product, or with the conditions reaction methods, sample types, reagents, users, and more. If an incorrect combination is detected or an identifier is deactivated, the process may be interrupted or the system may become inoperable until the user is notified, or with acknowledgment by a user.
[00177] The information of or associated with an identifier can, in some modalities, be stored, for example, in a computer memory or in a computer-readable medium, for the purposes of future reference and record keeping. For example, certain control systems may use information from or associated with identifiers to identify which components (for example, cassettes) or the type of cassettes that were used in a particular analysis, the date, time and duration of use, conditions of use, etc. Such information can be used, for example, to determine whether one or more components of the analyzer should be eliminated or replaced. Optionally, a control system or any other appropriate system can generate a report from the information collected, including information encoded by or associated with identifiers, which can be used to provide evidence of compliance with regulatory standards or the verification of quality control.
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[00178] The information encoded in or associated with an identifier can also be used, for example, to determine whether the component associated with the identifier (for example, a cassette) is authentic or false. In some embodiments, determining the presence of a false component causes the system to shut down. In one example, the identifier can contain a unique identity code. In this example, the software or the process control analyzer should not allow the system to start (for example, the system can be disabled) if a foreign or incompatible identity code (or no identity code) is detected.
[00179] In certain modalities, the information obtained from or associated with an identifier can be used to verify the identity of a customer to whom the cassette and / or the analyzer are sold or to whom a biological, chemical or pharmaceutical process must be performed . In some cases, information obtained from or associated with an identifier is used as part of a data collection process to detect defects in a system. The identifier can also contain or be associated with information such as group histories, assembly process and instrumentation diagrams (P and IDs), defect detection histories, among others. The detection of defects in a system can be carried out, in some cases, through remote access or include the use of diagnostic software.
[00180] In one embodiment, an analyzer includes a user interface, which can be positioned inside the enclosure and configured for a user to enter information in the sample analyzer. In one embodiment, the user interface includes a touch screen. The touch screen can guide a user through the operation of the analyzer, providing text and / or graphical instructions for using the analyzer. The user interface can guide the user to enter the patient's name or other source / patient identification number in the analysis.
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83/101 sador. Any appropriate patient information, such as the name, date of birth, and / or the patient ID number can be entered in the touch screen user interface to identify the patient. The user interface can indicate the amount of time remaining to complete the sample analysis.
[00181] In another mode, the user interface can be configured differently, such as with an LCD display and a one-button scroll menu. In another mode, the user interface can simply include a Start Copy button to activate the analyzer. In other embodiments, the user interface of separate standalone devices (such as a smart phone or mobile computer) can be used to interface with the analyzer.
[00182] Figure 8 is a block diagram 300 that illustrates how a control system 305 (see figure 7) can be operatively associated with a variety of different components according to a modality. The control systems described here can be implemented in numerous ways, such as with dedicated hardware or firmware, when using a processor that is programmed when using a microcode or software to perform the functions recited above or any appropriate combination of the elements above. A control system can control one or more operations from a single analysis (for example, for a biological, biochemical or chemical reaction), or from multiple analyzes (separate or interconnected). As shown illustratively in figure 7, the control system 305 can be positioned inside the analyzer enclosure 101 and can be configured to communicate with the identification reader 60, the user interface 200, the fluid flow source 40, the optical system 80 and / or the temperature regulating system for analyzing a sample in the cassette.
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[00183] In one embodiment, the control system includes at least two processors, including a real-time processor that controls and monitors all subsystems that directly form an interface with the cassette. In one embodiment, at a particular time interval (for example, every 0.1 second), that processor communicates with a second higher level processor that communicates with the user through the user interface and / or the communication subsystem (discussed below) and directs the analyzer operation (for example, determines when to start analyzing a sample and interprets the results). In one embodiment, communication between these two processors takes place via a serial communication bus. It should be appreciated that in another embodiment the analyzer can only include one processor, or more than two processors, since the invention is not so limited.
[00184] In one embodiment, the analyzer can make the connection with external devices and can, for example, include ports for the connection with one or more external communication units. External communication can be carried out, for example, via USB communication. For example, as shown illustratively in Figure 8, the analyzer can send the results of a sample analysis to a USB 400 printer, or to a 402 computer. In addition, the data stream produced by the processor in real time can be sent to a computer or to a USB 404 memory stick. In some embodiments, a computer may also be able to directly control the analyzer via a USB connection. In addition, other types of communication options are available, since the present invention is not limited in this regard. For example, Ethernet, Bluetooth and / or WI-FI 406 communication with the analyzer can be established through the processor.
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[00185] The calculation methods, steps, simulations, algorithms, systems and system elements described here can be implemented by using a computer-implemented control system, such as the various types of computer-implemented systems described Next. The methods, steps, systems and system elements described here are not limited in their implementation to any specific computer system described here, since many other different machines can be used.
[00186] The control system implemented by computer can be part of or be coupled in operative association with a sample analyzer, and, in some modalities, configured and / or programmed to control and adjust operational parameters of the sample analyzer, as well as analyze and calculate values, as described above. In some embodiments, the computer-implemented control system can send and receive reference signals to adjust and / or control operational parameters of the sample analyzer and, optionally, of another device in the system. In other embodiments, the computer-implemented system can be separated from and / or located remotely with respect to the sample analyzer and can be configured to receive data from one or more devices of the remote sample analyzer via an indirect and / or portable medium , such as through portable electronic data storage devices, such as magnetic disks, or through communication over a computer network, such as the Internet or a local intranet.
[00187] A control system implemented by a computer may include several known components and circuits, including a processing unit (ie processor), a memory system, input and output devices and interfaces (for example, a
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86/101 interconnection mechanism), as well as other components, such as transport circuits (for example, one or more buses), a video / audio data input / output (I / O) subsystem, special purpose hardware , as well as other components and circuits, as described in more detail below. In addition, the computerized system can be a multiprocessor computerized system or can include multiple computers connected to a computer network.
[00188] The control system implemented by computer may include a processor, for example, a commercially available processor such as one in the x86 series, Celeron and Pentium processors, available from Intel, AMD and Cirix similar devices, microprocessors in the series 680X0 available from Motorola, and the IBM PowerPC microprocessor. Many other processors are available, and the computer system is not limited to a particular processor.
[00189] A processor typically runs a program called the operating system, examples of which are: WindowsNT, Windows 95 or 98, UNIX, Linux, DOS, VMS, MacOS and OS8, which controls the execution of other computer programs and provides the programming, debugging, input / output control, counting, compiling, storage allocation, data management and memory management, communication control and related services. The processor and the operating system together define a computer platform for which application programs in high-level programming languages are recorded. The computer-implemented control system is not limited to a particular computer platform.
[00190] The control system implemented by computer may include a memory system, which typically includes a medium
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87/101 non-volatile activity that can be read and written by computer, examples of which are a magnetic disk, an optical disk, a flash memory and a tape. Such recording medium can be removable, for example, a floppy disk, read / write CD or memory bar, or it can be permanent, for example, a hard disk.
[00191] Such recording medium stores signals, typically in binary form (that is, a form interpreted as a sequence of ones and zeros). A disk (for example, magnetic or optical) has a number of tracks, on which such signals can be stored, typically in binary form, that is, a form interpreted as a sequence of ones and zeros. Such signals can define a software program, for example, an application program, to be executed by the microprocessor, or information to be processed by the application program.
[00192] The computer implemented control system memory system may also include an integrated circuit memory element, which is typically a volatile random access memory such as a dynamic random access memory (DRAM) or a static memory ( SRAM). Typically, in operation, the processor causes programs and data to be read from the non-volatile recording medium on the integrated circuit memory element, which typically allows faster access to instructions and program data by the processor than in the case of non-volatile recording medium.
[00193] The processor in general manipulates the data inside the integrated circuit memory element according to the program instructions and then copies the manipulated data in the middle of the non-volatile recording after the processing is finished. A variety of mechanisms are known to manage the movement of data between the non-volatile recording medium and the cir memory element.
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88/101 integrated cuito, and the computer-implemented control system that executes the methods, steps, systems and system elements described above with respect to figure 8 is not limited to this. The computer-implemented control system is not limited to a particular memory system.
[00194] At least a part of such a memory system described above can be used to store one or more data structures (for example, lookup tables) or equations described above. For example, at least a portion of the non-volatile recording medium can store at least a portion of a database that includes one or more of such data structures. Such a database can be any one of a variety of database types, for example, a file system that includes one or more flat file data structures where data is organized into data units separated by delimiters, one relational database where data is organized into data units stored in tables, an object-oriented database where data is organized into data units stored as objects, another type of database, or any combination of these.
[00195] The control system implemented by computer may include an input / output (I / O) subsystem of video and audio data. An audio portion of the subsystem can include an analog to digital (A / D) converter, which receives analog audio information and converts it to digital information. Digital information can be compressed by using known compression systems for hard disk storage for use on another occasion. A typical video portion of the I / O subsystem may include a video image compressor / decompressor, many examples of which are known in the prior art. Such compression
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89/101 res / decompressors convert analog video information into compressed digital information, and vice versa. The compressed digital information can be stored on the hard drive for use at a later time.
[00196] The control system implemented by computer can include one or more output devices. Exemplary output devices include a cathode ray tube (CRT) display, a liquid crystal display (LCD) and other video output devices, printers, communication devices such as a modem and a network interface, storage devices such as disc or tape, and audio output devices such as a speaker.
[00197] The control system implemented by computer can also include one or more input devices. Exemplary input devices include a keyboard, keypad, trackball, mouse, pen and tablet, communication devices as described above, and data input devices such as audio and video capture devices and sensors. The computer-implemented control system is not limited to the particular input or output devices described here.
[00198] It should be appreciated that one or more of any type of computer-implemented control system can be used to implement the various modalities described here. Aspects of the invention can be implemented in software, hardware or firmware, or any combination thereof. The control system implemented by computer can include specially programmed hardware, special purpose hardware, for example, an application specific integrated circuit (ASIC). Such special purpose hardware can be configured to perform one or more of the methods, steps, simulations, algorithms, systems, and system elements described above as part of the system.
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90/101 computer-implemented control theme described above or as an independent component.
[00199] The computer-implemented control system and its components can be programmable using any of a variety of one or more appropriate computer programming languages. Such languages can include procedural programming languages, for example, C, Pascal, Fortran and BASIC, object-oriented languages, for example, C ++, Java and Eiffel and other languages, such as a cryptography language or even assembly language.
[00200] Methods, steps, simulations, algorithms, systems, and system elements can be implemented by using any of a variety of appropriate programming languages, including procedural programming languages, programming languages oriented to objects, other languages and combinations of these, which can be implemented by such a computerized system. Such methods, steps, simulations, algorithms, systems, and system elements can be implemented as separate modules of a computer program, or they can be implemented individually as separate computer programs. Such modules and programs can be run on separate computers.
[00201] Such methods, steps, simulations, algorithms, systems, and system elements, individually or in combination, can be implemented as a computer program product incorporated in a tangible way as signals that can be read by a computer in a medium that it can be read by computer, for example, on a non-volatile recording medium, an integrated circuit memory element, or a combination of these. For each of such a method, step, simulation, algorithm, system, or system element, such
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91/101 computer program product can comprise computer-readable signals tangibly incorporated into the computer-readable medium that define instructions, for example, as part of one or more programs, which, as a result of the fact to be executed by a computer, it instructs the computer to execute the method, step, simulation, algorithm, system, or system element.
[00202] It should be appreciated that several modalities can be formed with one or more of the characteristics described above. The above aspects and features can be employed in any appropriate combination since the present invention is not limited in this regard. It should also be appreciated that the drawings illustrate the various components and features that can be incorporated into various modalities. For the sake of simplicity, some of the drawings may illustrate more than one feature or optional components. However, the invention is not limited to the specific embodiments shown in the drawings. It should be recognized that the invention encompasses modalities that can include only a part of the components illustrated in any figure in the drawing, and / or can also encompass the modalities that combine the components illustrated in multiple different figures in the drawing. EXAMPLES
[00203] The following example is intended to illustrate certain embodiments of the present invention, but does not exemplify the full scope of the invention.
Example 1
[00204] This example describes the use of a cassette and an analyzer to perform an assay to detect PSA in a sample by depositing non-electrolytically silver on gold particles that are associated with the sample. Figure 9 includes an illustration
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92/101 qualification of a 500 microfluidic cassette system used in this example. The cassette had a shape similar to cassette 20 shown in figure 3. The microfluidic system used in this example is described in a general manner in international patent publication no. WO2005 / 066613 (international patent application serial number PCT / US2004 / 043585), filed on December 20, 2004 and entitled Device and Test Method, which is incorporated herein by reference in its entirety for all purposes .
[00205] The microfluidic system included measurement zones 510A510D, the waste containment region 512, and an outlet 514. The measurement zones included a microfluidic channel 50 microns deep and 120 microns wide, with a total length of 175 mm. The microfluidic system also included microfluidic channel 516 and ramifications 518 and 520 of the channel (with entries 519 and 521, respectively). Branches 518 and 520 of the channel were 350 microns deep and 500 microns wide. Channel 516 was formed from subchannels 515, which were 350 microns deep and 500 microns wide located on alternate sides of the cassette, connected by through holes 517 having a diameter of about 500 microns. Although figure 9 shows that the reagents were stored on one side of the cassette, in other embodiments the reagents were stored on both sides of the cassette. Channel 516 had a total length of 390 mm, and each of branches 518 and 520 were each 360 mm in length. Before sealing the channels, anti-PSA antibodies were attached to a surface of the microfluidic system in a segment of the 510 measurement zone.
[00206] Before the first use, the microfluidic system was loaded with liquid reagents that were stored in the cassette. A series of 7 523-529 wash buffers (both water and buffer, about 2 microliters each) was loaded when using a pipette in the subca
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93/101 channels 515 of channel 516 when using through holes. Each wash buffer was separated by air buffers. Fluid 528, containing a silver salt solution, was loaded into the branch channel through port 519 using a pipette. Fluid 530, containing a reducing solution, was loaded into branch channel 520 through port 521. Each of the liquids shown in figure 9 was separated from the other liquids by air plugs. Doors 514, 519, 521, 536, 539 and 540 were sealed with adhesive tape that could be easily removed or punctured. In this way, the liquids were stored in the microfluidic system before the first use.
[00207] In the first use, doors 514, 519, 521, 536, 539 and 540 were detached by a user when pulling out an adhesive tape that covered the opening of the doors. A tube 544 containing lyophilized antiPSA antibodies labeled with colloidal gold and to which 10 microliters of blood sample (522) were added, was connected to ports 539 and 540. The tube was part of a fluid connector that has a shape and a configuration shown in figure 3. This created a fluid connection between measurement zone 510 and channel 516, which were not connected and not in fluid communication with each other before the first use.
[00208] The cassette that includes the microfluidic system 500 was inserted into an opening of an analyzer (for example, as shown in figure 7). The analyzer casing included an arm positioned inside the casing that was configured to couple the cam surface to the cassette. The arm extended at least partially into the opening in the housing in such a way that, when the cassette was inserted into the opening, the arm was pushed away from the opening to a second position allowing the cassette to enter the opening. Once the arm engages the cam surface internally of the cassette, the cassette is positioned and retained within the housing of the cassette.
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94/101 analyzer, and the spring push prevented the cassette from sliding out of the analyzer. The analyzer detects the insertion of the cassette by means of a position sensor.
[00209] An identification reader (RFID reader) positioned inside the analyzer housing was used to read an RFID tag on the cassette that includes the lot identification information. The analyzer used this identifier to combine the batch information (for example, calibration information, the cassette expiration date, the verification that the cassette is new, and the type of analysis / test to be performed on the cassette) stored in the cassette. analyzer. The user was prompted to enter information about the patient (from which the sample was acquired) on the analyzer when using the touch screen. After the information on the cassette was verified by the user, the control system started the analysis.
[00210] The control system included instructions programmed to perform the analysis. To start the analysis, a signal was sent to the electronic system that controls a vacuum system, which was a part of the analyzer and used to provide the flow of fluid. A distributor with O-rings was pressed against the surface of the cassette by a solenoid. A port in the dispenser was sealed (by an O-ring) to port 536 of the cassette's microfluidic system. This port on the distributor was connected by a tube to a simple solenoid valve (SMC V124A-6Q-M5, not shown) that was opened to the atmosphere. A separate vacuum port on the dispenser was sealed (by an O-ring) to port 514 of the cassette's microfluidic system. A vacuum of about -30 kPa was applied to port 514. Throughout the analysis, the channel including measurement zone 510 positioned between ports 540 and 514 had a substantially constant non-zero pressure drop of around -30 kPa. Sample 522 was flowed in the direction of arrow 538 in each of the 510A-510D measurement zones.
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When the fluid passed through the measurement zones, the PSA proteins in sample 522 were captured by the anti-PSA antibodies immobilized on the walls of the measurement zone, as described in more detail below. The sample took about 7 to 8 minutes to pass through the measurement zone, after which it was captured in the waste containment region 512.
[00211] The initiation of the analysis also involved the emission of a signal from the control system to the optical detectors, which were positioned adjacent to each of the measurement zones 510, to start the detection. Each of the detectors associated with the measurement zones recorded the transmission of light through the channels of the measurement zones, as shown in a graph 600 illustrated in figure 10. When the sample passed through each of the measurement zones, the 610A peaks -610D were produced. The peaks (and valleys) measured by the detectors are signals (or are converted into signals) that are sent to the control system that compared the measured signals to the reference signals or pre-programmed values in the control system. The control system included a pre-programmed set of instructions to provide feedback to the microfluidic system based at least in part on the comparison of signals / values.
[00212] In a first measurement zone 510-A of the device 500 of figure 9, the channel walls of that measurement zone were blocked with a blocking protein (bovine serum albumin) before the first use (for example, before seal the device). Little or no protein in the blood sample has been attached to the walls of the 510-A measurement zone (with the exception perhaps of some non-specific binding that can be eliminated). This first measurement zone acted as a negative control.
[00213] In a second measurement zone 510-B, the channel walls of that measurement zone were covered with a large amount of
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96/101 predetermined age of a prostate specific antigen (PSA) before first use (for example, before sealing the device) to act as an elevated or positive control. Due to the fact that the blood sample passed through the second 510-B measurement zone, little or no PSA protein in the blood was attached to the channel walls. Antibodies to gold-conjugated signals in the sample may not yet be bound to the PSA in the sample, and thus may bind to the PSA on the channel walls to act as an elevated or positive control.
[00214] In a third measurement zone 510-C, the channel walls of that measurement zone were coated with a small predetermined amount of PSA before first use (for example, before sealing the device) to act as a low control . Due to the fact that the blood sample flowed through this measurement zone, little or no PSA protein in the sample was attached to the channel wall. The gold-conjugated signal antibodies in the sample can bind to PSA on the channel walls to act as a low control.
[00215] In a fourth measurement zone 510-D, the channel walls of that measurement zone were coated with the capture antibody, an anti-PSA antibody, which binds to a different epitope in the PSA protein of the signal antibody conjugated with gold. The walls were covered before the first use (for example, before sealing the device). Due to the fact that the blood sample flowed through the fourth measurement zone during use, the PSA proteins in the blood sample bound to the anti-PSA antibody in a way that is proportional to the concentration of these proteins in the blood. Since the sample, which included PSA, also included gold-labeled anti-PSA antibodies coupled with PSA, the PSA captured on the walls of the measurement zone formed a sandwich immune complex.
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[00216] The washing fluids 523-529 followed the sample through the measurement zones 510 towards the waste containment region 512 in the direction of the arrow 538. When the washing fluids were passed through the measurement zones, they removed the components Unbound sample samples remaining. Each wash buffer cleaned the channels of the measurement zones, providing progressively more thorough cleaning. The last washing fluid 529 (water) removed the salts that could react with the silver salts (for example, chloride, phosphate, azide).
[00217] As shown in the graph illustrated in figure 10, when washing fluids were flowing through the measurement zones, each of the detectors associated with the measurement zones measured a 620 peak and valley pattern. The valleys corresponded to the washing plugs (which are transparent liquids and thus provide maximum light transmission). The peaks between each buffer represent the air between each buffer of clear liquid. Since the assay included 7 wash buffers, 7 valleys and 7 peaks are shown in Graph 600. The first valley 622 is generally not as deep as the other 624 valleys, since the first wash buffer frequently captures the cells of the blood that stays in the channel and thus is not completely transparent.
[00218] The final air peak 628 is much longer than the previous peaks because there was no wash buffer to follow. Due to the fact that a detector detects the length of this air spike, one or more signals are sent to the control system that compares the time span of that peak to a previously set reference signal or input value that has a particular length. If the time span of the measured peak is long enough compared to the reference signal, the control system sends a signal to the electronics that controls the breather valve.
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98/101 tion 536 to activate the valve and start mixing fluids 528 and 530. (It should be noted that the air peak signal 628 can be combined with a signal that indicates any of: 1) the peak intensity; 2) where that peak is positioned as a function of time, and / or 3) one or more signals that indicate that a series of peaks 620 of particular intensity have passed. In this way, the control system distinguishes air peak 628 from other long-duration peaks such as peak 610 in the sample, for example, when using a signal pattern).
[00219] To start mixing, the solenoid connected by the distributor to the exhaust port 536 is closed. Since the vacuum remains activated and no air can enter through the breathing valve 536, air enters the device through ports 519 and 521 (which are open). This forces the two fluids 528 and 530 in the storage channels upstream of the breathing valve 536 to move substantially simultaneously to outlet 514. These reagents mix at the intersection of the channels to form an amplifying reagent (a solution reactive silver) which has a viscosity of about 1x10 ^-3 Pa-s. The volume ratio of fluids 528 and 530 was about 1: 1. The amplification reagent continued through the downstream storage channel, through tube 544, through measurement zones 510, and then to the waste containment region 512. After a stipulated amount of time (12 seconds), the analyzer reopened the breathing valve 536 in such a way that air flowed through the breathing valve 536 (instead of the exhaust ports). This left some reagent behind in the upstream storage channel 518 and 520 on the device. This also results in a single mixed amplification reagent buffer. The 12 seconds of closing the exhaust valve results in an amplification buffer of about 50 pl. (Instead of
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99/101 simple timing, another way to activate the reopening of the breathing valve is to detect the amplification reagent when it first enters the measurement zones).
[00220] Due to the fact that the mixed amplification reagent is stable for only a few minutes (usually less than 10 minutes), mixing was performed less than one minute before use in measurement zone 510. The amplification reagent is a liquid transparent, so that when it enters the measurement zones, the optical density is at its lowest level. When the amplification reagent passed through the measurement zones, silver was deposited on the captured gold particles to increase the size of the colloids in order to amplify the signal (as noted above, the gold particles were present in the measurement zones High and Low positive control and, to the extent that PSA was present in the sample, in the test measurement zone). The silver can then be deposited on top of the silver already deposited, reacting more and more silver deposited in the measurement zones. Eventually, the silver deposited reduces the transmission of light through the measurement zones. The reduction in transmitted light is proportional to the amount of silver deposited and may be related to the amount of gold colloids captured on the canal walls. In a measurement zone where no silver has been deposited (the negative control, for example, or the test area when the sample contains no target proteins, such as PSA), there will be no (or minimal) increase in optical density. In a measurement zone with significant silver deposition, the slope and the final level of the optical density increase pattern will be high. The analyzer monitors the pattern of this optical density during amplification in the test area to determine the concentration of analyte in the sample. In a test version, the pattern is monitored within the first three minutes of amplification. The density
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100/101 optics in each of the measurement zones as a function of time has been recorded and is shown as curves 640, 644, 642 and 646 in the figure
10. These curves corresponded to the signals that were produced in the measurement zones 510-A, 510-B, 510-C and 510-D, respectively.
[00221] After three minutes of amplification, the analyzer stops the test. No more optical measurements are recorded and the distributor is decoupled from the device. The test result is indicated on the analyzer screen and communicated to a printer, computer, or whatever output the user has selected. The user can remove the device from the analyzer and discard it. The sample and all reagents used in the assay remain in the device. The analyzer is ready for another test.
[00222] It should be noted that controlling the flow rates of fluids within channel 516 and measurement zone 510 was important when fluids were flowing through the system. Due to the relatively small cross-sectional area of the measurement zone, it served as a bottleneck, controlling the total flow in the system. When the measurement zone contained liquids, the linear flow rates of liquids in channel 516 were about 0.5 mm s ^-1 . Fluids flowing from branch channels 518 and 520 to main channel 516 may not have reproducibly mixed at this rate, since one fluid may flow more quickly than the other, causing uneven parts of the fluids 528 and 530 were mixed. On the other hand, when the measurement zone contained air, the linear flow rates of fluids in channel 516 and in branch channels 518 and 520 were about 15 mms s ^-1 . At this higher flow, the flow in branch channels 518 and 520 was equal and reproducible (when the breathing valve 536 was closed), producing a reproducible mixture. For this reason, the valve connected to port 536 was not closed until fluid 542 had passed through the measurement zone to the
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101/101 waste containment region. As noted above, the determination of when fluid 542 had left measurement area 510 was made by using an optical detector in order to measure the transmission of light through a portion of measurement area 510 in combination with a feedback system .
[00223] The microfluidic system shown in figure 9 was designed in such a way that the volume of the channel between the breathing valve 536 and the measurement zone 510 was greater than the expected volume of the mixed activated silver solution (ie, the combined portion of fluids 528 and 530 that moved into channel 516 when breathing valve 536 was closed). This ensured that substantially all of the mixing took place at a relatively high linear flow (since no liquid, and only air, was present in the measurement zone 510 at that time), and before the activated solution reached the measurement zone. This configuration helped to promote a reproducible and identical mix. For the test described in this example, it was important to sustain a flow of the activated silver mixture within the measurement zone for a few minutes (for example, 2 to 10 minutes).
[00224] This example shows that the analysis of a sample in a microfluidic system on a cassette can be performed by using an analyzer that controls the flow of fluid in the cassette, and by using feedback from one or more measured signals to modulate the flow of fluid.
[00225] Although various modalities of the present invention have been described and illustrated herein, the elements skilled in the art will immediately provide for a variety of other means and / or structures to perform the functions and / or obtain the results and / or one or more of the advantages described herein, and each such variation and / or modification is considered to be within the scope of the present invention.

权利要求:
Claims (14)
[1]
1. Method of conducting quality control to determine abnormalities in the operation of a microfluidic system characterized by the fact that it comprises the steps of:
detecting a first fluid and a second fluid in a first measurement zone of the microfluidic system and forming a first signal corresponding to the first fluid and a second signal corresponding to the second fluid;
transmitting at least one signal between the first signal and the second signal to a control system;
comparing at least one signal between the first signal and the second signal to a reference signal, thereby determining the presence of abnormalities in the operation of the microfluidic system; and determine whether to modulate the fluid flow in the microfluidic system and / or alert a user to an abnormality in an analysis being conducted in the microfluidic system based at least in part on the results of the compare step, in which to determine whether to modulate the flow of fluid fluid in the microfluidic system comprises determining whether for an analysis being conducted in the microfluidic system.
[2]
2. Method according to claim 1, characterized by the fact that detecting the first fluid and the second fluid in the first measurement zone of the microfluidic system comprises detecting at least two of:
a) an opacity of the first fluid;
b) a volume of the first fluid;
c) a flow of the first fluid;
d) a position of the detection of the first fluid in time in relation to a second position in time; and
e) an average period of time between the detection of the first and second liquids.
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[3]
3. Method, according to claim 1 or 2, characterized by the fact that it comprises:
transmitting a first signal pattern to a control system, the first signal pattern comprising at least two of:
a) an intensity of the first signal;
b) a duration of the first signal;
c) a position of the first signal in time in relation to a second position in time; and
d) an average period of time between the first and second signals; and determine whether to modulate fluid flow in the microfluidic system and / or alert a user to an abnormality in an analysis being conducted in the microfluidic system based at least in part on the first signal pattern, in which to determine whether to modulate fluid flow in the microfluidic system comprises determining whether for an analysis being conducted in the microfluidic system.
[4]
4. Method according to any of the claims
1 to 3, characterized by the fact that:
a) further comprises counting a series of signals each having an intensity above or below a limit intensity, and determining whether to modulate the fluid flow in the microfluidic system and / or alert a user of an abnormality in an analysis being conducted in the microfluidic system based at least in part on the number of signals that have an intensity above or below the limit intensity; or
b) it also comprises counting a series of signals, in which the series of signals is generated by the passage of a series of washing fluids along the first measurement zone.
[5]
5. Method according to any of the claims
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1 to 4, characterized by the fact that:
a) the first and / or the second fluid is a washing fluid; or
b) the first and second fluids are immiscible with each other; or
c) the first fluid is a liquid and the second fluid is a gas; or
d) the first and second fluids are miscible with each other; or
e) the first and second fluids are separated by a third immiscible fluid; or
f) the first and second fluids are separated by air; or
g) the first fluid is a sample; or
h) the first fluid comprises whole blood; or
i) the first fluid is an amplification reagent; or
j) the first fluid is air; or
k) the first and second fluids do not contain a component of a chemical and / or biological reaction; or
l) the first and / or the second fluid is aqueous; or
m) the first fluid is serum or plasma.
[6]
6. Method according to any of the claims
1 to 5, characterized by the fact that:
a) comprises the continuous or periodic detection of the passage of any fluids through the first measurement zone;
b) it also comprises the continuous or periodic detection of the passage of fluids through a second measurement zone of the microfluidic system; or
c) comprising detecting a third fluid in the first measurement zone; or
d) comprising detecting a third fluid, fourth fluid and a fifth fluid in the first measurement zone;
e) further comprising the detection of a second fluid
Petition 870200007448, of 1/16/2020, p. 111/124
4/11 in the first measurement zone of the microfluidic system and the formation of a second signal corresponding to the second fluid; or
f) comprising passing a sample through the first measurement zone; or
g) comprising depositing a sample component from the sample in the first measurement zone; or
h) further comprising detecting a signal indicating a metal deposited on a surface of the first measurement zone; or
i) wherein the first washing fluid and the second washing fluid are liquid, the method further comprising passing a gaseous buffer along the first measurement zone after the passage of the second washing fluid; or
j) in which the first and second measurement zones are positioned in series with respect to each other; or
k) in which the microfluidic system comprises a first detector statically positioned adjacent to the first measurement zone during the detection step.
[7]
7. Method according to any of the claims
1 to 6, characterized by the fact that:
a) further comprises the transmission of an electrical signal from the control system to a component of the microfluidic system that can modulate the fluid flow as a result of the transmission step; or
b) it also comprises the transmission of an electrical signal from the control system to a component of the microfluidic system, where the component of the microfluidic system is a pump, a valve or a vacuum; or
c) still comprises the transmission of the first signal pattern to a control system, the comparison of at least the first
Petition 870200007448, of 1/16/2020, p. 112/124
5/11 first signal pattern to a pre-programmed signal or value control pattern, and determining whether the application of the fluid flow source to the microfluidic system should be stopped based at least in part on the results of the comparison step ; or
d) comprises the transmission of the first signal pattern to a control system, the first signal pattern comprising at least three of:
i) an intensity of the first signal;
ii) a duration of the first signal;
iii) a position of the first signal in time in relation to a second position in time; and iv) an average period of time between the first and second signals; or
e) the transmission comprises the first signal pattern to a control system, the first signal pattern comprising at least two of:
i) an intensity of the first signal;
ii) a duration of the first signal;
iii) a position of the first signal in time in relation to a second position in time;
iv) an average period of time between the first and second signals;
v) a second signal strength;
vi) a duration of the second signal; and vii) a position of the second signal in time in relation to a second position in time.
[8]
8. Method according to any one of the claims
1 to 7, characterized by the fact that:
a) also includes the comparison of the first signal pattern to a pre-programmed signal or value control pattern
Petition 870200007448, of 1/16/2020, p. 113/124
6/11 in the control system; or
b) wherein the first signal pattern comprises an intensity of the first signal and a duration of the first signal; or
c) wherein the first signal pattern comprises an intensity of the first signal and a position of the first signal in time with respect to a time of the initiation step;
d) in which the first signal pattern is produced without a label; or
e) the first signal pattern comprises an intensity of the first signal; or
f) in which the first signal pattern comprises an intensity of the first signal and an average period of time between the first and second signals.
[9]
9. Method according to any of the claims
1 to 8, characterized by the fact that:
a) in which the intensity of the first signal comprises an average or maximum intensity; or
b) where the first signal is indicative of the passage of the first fluid through the first measurement zone; or
c) where the first signal is indicative of a component deposited from the first fluid in the first measurement zone; or
d) where the first signal is indicative of a metal deposited from the first fluid in the first measurement zone; or
e) in which the first signal comprises an intensity as a function of time; or
f) where the intensity of the first signal is indicative of a fluid type of the first fluid and the duration of the first signal is indicative of a flow rate of the first fluid; or
g) where the first signal is indicative of the first fluid that passes through the first measurement zone and the second signal is
Petition 870200007448, of 1/16/2020, p. 114/124
7/11 indicative of the second fluid passing through the first measurement zone, where the first and second signals are separated by a period of time.
[10]
10. Method according to any one of claims 1 to 9, characterized by the fact that:
a) comprises the formation of signals corresponding to the passage of each fluid passage through the first measurement zone; or
b) in which the intensity of each of the signals is indicative of the concentration of a component in a fluid and / or the amount of a component in a fluid that passes through the first measurement zone; or
c) where the intensity of each signal is indicative of the type of fluid that passes through the first measurement zone; or
d) where the intensity is determined by the opacity of a fluid or a component of the fluid;
e) comprises determining the opacity of the sample component or a material associated with the sample component in the first measurement zone as a function of time; or
f) comparing a signal strength with a programmed threshold strength within a control system.
[11]
11. Method according to any one of claims 1 to 10, characterized by the fact that:
a) comprises the application of a fluid flow source to a microfluidic system, wherein the determination step comprises determining whether the application of the fluid flow source to the microfluidic system should be stopped based at least in part on the results of the comparison stage; or
b) also includes the passage of a second fluid through the first measurement zone of the microfluidic system, that of
Petition 870200007448, of 1/16/2020, p. 115/124
8/11 protection of the passage of the second fluid through the first measurement zone, and the formation of a first signal pattern as a result of the detection step, wherein the first signal pattern includes a first signal indicative of the passage of the first fluid through the first measurement zone and a second signal indicating the passage of the second fluid through the first measurement zone, where the first and second signals are separated by a period of time; or
c) comprises the passage through the first measurement zone of first and second fluids in sequence, in which the first and second fluids are immiscible with each other, the detection of a property of the first fluid and the formation of a first sign indicating the property of the first fluid, the transmission of the first signal to a control system, the transmission of a signal from the control system to a component of the microfluidic system that can modulate the flow of fluid, the activation of the component of the microfluidic system that can modulate fluid flow, and modulation of fluid flow upstream of the first measurement zone; or
d) comprises the application of a substantially constant vacuum to an output in fluid communication with the first measurement zone while the first and second fluids flow into the measurement zone.
[12]
12. Method according to any one of claims 1 to 11, characterized by the fact that:
a) in which the detection step is performed by measuring the transmission of light or absorption light through the first and second fluids; or
b) in which the detection comprises the measurement of light transmission through the measurement zone; or
c) in which the transmission of light or absorption of light through
Petition 870200007448, of 1/16/2020, p. 116/124
9/11 of the first and second fluids is measured as a function of time; or
d) comprising the measurement of light transmission or light absorption through a sample fluid in the first measurement zone; or
e) measure light transmission or light absorption through a series of washing fluids in the first measurement zone.
[13]
13. Method according to any one of claims 1 to 12, characterized in that it comprises:
a) detecting each fluid that passes through the first measurement zone during analysis, forming a signal for each fluid to produce an analysis signal pattern, and determining analysis information based on at least part about the analysis signal pattern; or
b) providing the user with information about the analysis based on the detection of an abnormality during the analysis; or
c) providing the user with information, in which providing the user with information comprises alerting the user through a user interface; or
d) alerting the user through a user interface, where the user interface comprises a screen that is part of a sample analyzer, and where the sample analyzer comprises the control system and at least one detector to detect the first and second fluids in the first measurement zone; or
e) the provision of information to the user, in which the information provided to the user comprises information that the results of the analysis should not be trusted, that the analysis must be carried out again, that the analysis may take longer to be performed, or that the user must take action; or
f) providing the user with information, in which the information
Petition 870200007448, of 1/16/2020, p. 117/124
10/11 training provided to the user comprises information that the analysis is canceled and / or the results must be disregarded; or
g) providing the user with information and seeking additional input from the user after providing the user with the information or using the information on the analysis to provide feedback to the microfluidic system and / or conduct quality control; or
h) providing feedback to the microfluidic system and / or conduct quality control, with information about the analysis.
[14]
14. Method according to any one of claims 1 to 13, characterized by the fact that:
a) comprises determining whether to modulate fluid flow in the microfluidic system and / or alert a user of an anomaly in an analysis being conducted in the microfluidic system based at least in part on information derived from the intensity and / or duration of a signal obtained from the passage of the gaseous buffer through the first measurement zone; or
b) comprises determining whether to modulate the fluid flow in the microfluidic system and / or alert a user of an anomaly in an analysis being conducted in the microfluidic system based at least in part on information derived from the intensity and / or duration of a signal obtained from the passage of the first washing fluid and the second washing fluid through the first measurement zone; or
c) comprises determining whether to modulate the fluid flow in the microfluidic system and / or alert a user of an anomaly in an analysis being conducted in the microfluidic system based at least in part on the number of washing liquids passing through
Petition 870200007448, of 1/16/2020, p. 118/124
11/11 of the first measurement zone; or
d) comprises determining whether to modulate the fluid flow in the microfluidic system and / or alert a user of an anomaly in an analysis being conducted in the microfluidic system based at least in part on information derived from the intensity and / or duration of a signal obtained from passing the sample through the first measurement zone.

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法律状态:
2018-04-10| B06F| Objections, documents and/or translations needed after an examination request according [chapter 6.6 patent gazette]|

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2019-02-12| B06T| Formal requirements before examination [chapter 6.20 patent gazette]|

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2019-10-22| B07A| Technical examination (opinion): publication of technical examination (opinion) [chapter 7.1 patent gazette]|

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2020-04-07| B09A| Decision: intention to grant [chapter 9.1 patent gazette]|

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2020-06-09| B16A| Patent or certificate of addition of invention granted|Free format text: PRAZO DE VALIDADE: 20 (VINTE) ANOS CONTADOS A PARTIR DE 15/04/2011, OBSERVADAS AS CONDICOES LEGAIS. |

优先权:

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申请号 | 申请日 | 专利标题

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US32504410P| true| 2010-04-16|2010-04-16|

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US32502310P| true| 2010-04-16|2010-04-16|

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US36300210P| true| 2010-07-09|2010-07-09|

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PCT/US2011/032680|WO2011130625A1|2010-04-16|2011-04-15|Feedback control in microfluidic systems|

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