• 专利附录
  • 专利说明
  • 权利要求
  • 类似技术
  • 同族专利
  • 引用文献
  • 法律状态
  • 优先权
    • 专利摘要:
    Die Erfindung betrifft ein Verfahren zur Herstellung von a-und y-Interferon durch Trennung der Leukozytenfraktion (buffy coat) des Blutes, Entfernung der Erythrozyten, Suspendieren der Leukozyten in einer geeigneten Nährlösung und Behandlung mit einem a-Interferon-Induktor und einem y-Interferon-Induktor, dadurch gekennzeichnet,daß man das a-Interferon und y-Interteron in nacheinanderfolgenden Stufen in derselben Leukozytenkultur erzeugt, indem man die nach Trennung der Leukozytenfraktion des Blutes, Entfernung der Erythrozyten und Suspendieren der Leukozyten in einer geeigneten Nährlösung erhaltene Suspension mit a- oder β-Interteron vorbehandelt, mit einem a-Interferon-Induktor in Berührung bringt, die a-interferonhaltige Flüssigkeit von den Zellen abtrennt, das a-Interferon erwünschtenfalls gewinnt, die Zellen wäscht und in einer geeigneten Nährlösung suspendiert, mit einem mitogenen Mittel behandelt, die y-interferonhaltige Flüssigkeit von den Zellen abtrennt und erwünschtenfalls das y-Interferon gewinnt und erwünschtenfalls a-interteronfrei macht. Der Vorteil des erfindungsgemäßen Verfahrens liegt darin, daß a- und β-Interferonproduktion in derselben Zellenkultur in zwei nacheinanderfolgenden Stufen durchgeführt werden kann. Das Verfahren wird dadurch viel wirtschaftlicher und die Erfindung ermöglicht eine wesentliche Erhöhung der interferonproduzierenden Kapazität der nur begrenzt zur Verfügung stehenden Leukozyten.
    公开号:SU1713591A1
    申请号:SU847773679
    申请日:1984-12-11
    公开日:1992-02-23
    发明作者:Тот Миклош；Эндрес Валериа；Белади Илона；Тот Шандор
    申请人:Эдьт Дьедьсерведьесети Дьяр (Инопредприятие)；
    IPC主号:
    专利说明:

    The invention relates to medicine, namely to the production of biological products, and relates to a method for producing human leukocyte interferon.
    Closest to the proposed one is a method for producing human leukocyte a- or y-interferon, which consists in separating the leukocyte fraction, removing red blood cells from it, cultivating a suspension of leukocytes in a nutrient medium and treating it with an appropriate inducer to induce a- or y-interferon.
    However, the known method allows to obtain on the same culture only a-or y-interferon.
    The aim of the invention is to obtain on the same culture sequentially a- and y-interferon.
    Example·! The collected blood placed in an ACI solution (aqueous solution containing citric acid and dextrose) is stored at 4 ° C for 3 hours. One volume part of the leukocyte concentrate thus obtained is mixed with five volume parts of a 0.83% aqueous solution of ammonium chloride while cooling in an ice bath. The suspension is kept in an ice bath until erythrocyte lysis occurs (5-10 min). After which the suspension of leukocytes is cultured in a nutrient solution of the following composition, mg / l:
    Calcium Chloride 175-350
    Potassium chloride 300-500
    Magnesium sulfate or an equivalent amount of magnesium chloride 175-500
    SU., ·, 1713591 A1
    Sodium chloride 5000-7000 Bicarbonate of soda 200-3500 Monosubstitutedsodium phosphate 30-150 Glucose 500-5500 Iron (III) nitrate 0,0-0,2 In this case, you can use a feeder
    needle environment, modified MEM Dulbekko, modified MEM Glazgova, 10
    The indicated erythrocyte leading is repeated in such a way that by 1 vol. including suspension of leukocytes, the use of 10 vol. including 0.83% aqueous solution of ammonium chloride. A 15 suspension of leukocytes in a nutrient solution is formed, after which the number of cells is set equal to 10 7 cells / ml. Nutrient u solution contains 2 mg / ml of globulin-free human blood serum (about 20. Lo 6%). The cells are treated at 37 ° C. with constant stirring of 200 IU / ml of concentrated inducer-free human interferon-free inducer. After 2 hours, the cells induce 400 hemagglutinated 25 units of Sendai virus. Incubation is completed 18 hours after induction. Suspension liquid antiviral titer, i.e. crude a-interferon is
    54,200 El / ml. The cells used to obtain α-interferon once were washed with the indicated nutrient solution, after which a suspension of leukocytes in the nutrient solution was formed at a concentration of 2.5 × 10 7 cells / ml. The nutrient solution contains 1 mg / ml of globulin-free human blood serum (about 3%). Then the cells are stimulated with 15 μg / ml of concanavalin A. The incubation is carried out with constant stirring 40 at 37 ° C and 16 hours after induction, the supernatant is separated from the cells. The antiviral titer of the supernatant that contains the / -interferon is determined on human amnial 45 WISH cells. In addition, the titers are expressed in comparison with the standard human α-interferon in the form of international units (ml / E / ml). There is currently no recognized international 50 interferon standard. The titer of the suspension liquid is 10.600 IU / ml and is a potentiated value, since the preparations of the / -interferon also contain “interferon. Using a 55 grading curve and based on the titer of the substance that was initially processed at pH 2, the actual content of β-interferon is 1.840 IU / ml. For comparison, the above method is carried out with the change that the cells are incubated for one day without the production of Sendai viruses before the production of / -interferon. The titer of β-interferon obtained under the influence of concanavalin A is 340 IU / ml.
    For further comparison, exclude the first, i.e. the α-interferon-producing period and isolated white blood cells are directly induced by conconavalin A. The titer of β-interferon is 450 IU / ml.
    Example 2. Act analogously to example 1 with the difference that as a nutrient solution apply the nutrient medium Needle. Thus obtained titer of α-interferon is ’56,000 IU / ml. In the second stage of the method, the titer of / -interferon equal to 1,680 IU / ml is achieved in the same nutrient solution. If in this method the production of α-interferon is excluded, the titer of / -interferon will be 280 IU / ml.
    The proposed method allows to obtain a - and / -interferons with a high titer on the same leukocyte culture.
    权利要求:
    Claims (1)
    [1]
    Claim
    A method of producing human leukocyte interferon by dividing the blood leukocyte fraction, removing red blood cells from it, cultivating a suspension of leukocytes in a nutrient medium, treating them with an interferon inducer and separating the supernatant containing interferon, characterized in that, in order to obtain sequentially on the same culture α- and γ-interferon, cultivation of a suspension of leukocytes is carried out in the presence of 200 IU / ml of α-interferon for two hours, the Sendai virus is used as an indicator entratsii 400 GE / ml and cultured for 18 h., separated supernatant containing α-interferon, precipitated leukocyte cells are washed, resuspended in culture medium treated konkonavalinom A, were cultured for 16 hours and separating the supernatant containing / interferon.
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    同族专利:
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    HUT38399A|1986-05-28|
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    引用文献:
    公开号 | 申请日 | 公开日 | 申请人 | 专利标题
    JPS6154040B2|1979-05-24|1986-11-20|Hayashibara Takeshi|
    FR2513124B1|1981-07-21|1989-11-17|Hayashibara Biochem Lab|PRODUCTION AND APPLICATIONS OF THE TARGET CELL LYSE FACTOR|
    DE3136166A1|1981-09-12|1983-04-14|Blutspendedienst der Landesverbände des Deutschen Roten Kreuzes Niedersachsen, Oldenburg und Bremen Gemeinnützige GmbH, 3257 Springe|Process for obtaining interferon- alphaand interferon- gamma|
    HU184972B|1981-12-01|1984-11-28|Egyt Gyogyszervegyeszeti Gyar|Process for preparing human gamma interferone|HU201100B|1988-03-04|1990-09-28|Egyt Gyogyszervegyeszeti Gyar|Process for large-scale production of high purity human leukocyte alpha interferon with reduced endotoxin content and with improved therapeutic effect|
    US5676942A|1992-02-10|1997-10-14|Interferon Sciences, Inc.|Composition containing human alpha interferon species proteins and method for use thereof|
    WO1993016107A1|1992-02-10|1993-08-19|Interferon Sciences, Inc.|Improved alpha interferon composition and method for its production from human peripheral blood leukocytes|
    CA2078805C|1992-09-22|1997-02-25|Intelcor Biotech Enterprises Inc.|Cytokine preparation|
    HU222980B1|1998-03-13|2004-01-28|Acapi, Alpha-Chem Advanced Pharmaceutical Industries S.A.E.|Process for producing human alpha-interferon|
    SE519827C2|1998-03-30|2003-04-15|Viranative Ab|Nutritional medium containing methionine and its use|
    WO1999050390A1|1998-03-30|1999-10-07|Bionative Ab|Methionin containing animal cell culture medium and its use|
    US6433144B1|1999-01-12|2002-08-13|Viragen, Inc.|Compositions of highly-purified natural mixtures of type I Interferon derived from leukocytes and methods|
    WO2005069778A2|2004-01-09|2005-08-04|Electric Mobility Corporation|Vehicle with improved turning|
    法律状态:
    优先权:
    申请号 | 申请日 | 专利标题
    HU834237A|HU192254B|1983-12-13|1983-12-13|Process for producing human leucocite and human gamma interferons in consecutive steps|
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