• 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:
    Process for the production of human gamma-interferon by separating the leukocyte fraction (buffy coat) of blood, removing the erythrocytes, suspending the leukocytes in a suitable nutrient medium, treating the same with a mitogenic agent and separating the liquid containing interferon from the cells which comprises pre-treating the leukocytes with 200-20000 IU/ml. of alpha- or beta-interferon prior to treatment with the mitogenic agent. The advantage of the process is that the gamma-interferon production is increased by 500-1000 %.
    公开号:SU1405691A3
    申请号:SU833624508
    申请日:1983-07-26
    公开日:1988-06-23
    发明作者:Белади Илона；Тот Миклош；Ростоци Иштван；Тот Шандор；Эндрес Валериа
    申请人:Эдьт Дьедьсерведьесети Дьяр (Инопредприятие)；
    IPC主号:
    专利说明:

    (21) (86) (22) (31) (32) (33) (46) (71) (72)
    3624508 / 28-14 RST / Ni 82/00064 07.26.83 3594/81 12.1.8.8
    neither
    06/23/88.
    (11/30/82)
    Bul No. 23
    Edt Dyodvervedenseseti Gyur (HU) Ilona Beladi, Miklos Toth, Ishtvan Rostotsi, Shandor Toth and Valeria Endres (ni)
    (53) 615.45 (088.8) (56) J. Immunol, 1980, 10, 877. Virology, T961, 14, 359.
    (54) METHOD FOR PRODUCING HUMAN GAMMA-SHTERFERON
    (57) The invention relates to medicine, not more precisely to the pharmaceutical industry, namely to the production of v-interferon. The purpose of the invention is to increase the yield of the target product by additional treatment of leukocytes or interferon in the amount of 500-5000 IE / ml for 1-12 hours. Then mitogen-leukocytes are treated with. ticking agent and centrifuged. Erythrocytes are removed with a 0.83-0.89% aqueous solution of ammonium chloride. Leukocytes are suspended in a nutrient medium containing calcium chloride, ferric nitrate, potassium chloride, magnesium sulfate, heptahydrate, sodium chloride, sodium hydrogencarbonate, sodium dihydrate monosubstituted sodium phosphate and glucose.
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    ; The invention relates to the field of medicine, in particular to the pharmaceutical industry of gamma-interferon. I The aim of the invention is to increase the yield of the target product due to the additional treatment of leucogs, cytes with alpha or beta interferon.
    The method is carried out as follows.
    Example 1 Blood obtained in an LCD solution (water solution containing lemon cylot and dextrose) is stored for 3 hours at a temperature. The leukocyte fraction is separated by centrifugation and left standing; . Then one removable part of the concentration of leukocytes thus obtained is mixed from May 5, h,), with a 89% aqueous solution of ammonium chloride at 4 ° C, the suspension is kept until complete; red blood cells are dissolved (usually this process ends in 5-10 min 1 after which the leukocytes are separated by centrifuging () by bending the IC is suspended in a nutrient medium of the following composition, mg / l:
    Calcium chloride 265 ferrous nitrate, 1
    Potassium chloride 400 magnesium sulfate,
    heptahydrate 200
    Sodium Chloride 6400 1 Sour Carbonate I Sodium2750
    Monosodium phosphate dihydrate
    rub 140
    Glucosl4500
    After this, the erythrocyte is removed i | iHe again, using | 0. 0.83% aqueous solution: | silver ammonium. Leukocytes. Suspended; (Schyut in a nutrient solution, bringing the concentration of cells in it to 0 klefok / ml, In the nutrient solution, except for} | ingredients, add i mg / ml human agamic serum and: And µg / ml neomycin. The si is treated for 4 hours at 37 ° C and continuously stirring with alpha interferon in the amount of 1500 IE, pretreated at pH 2.0 to remove the Sendai virus from it. Then the interferon is removed by doubling the cells with a solution Xnxx and concavaline is added to them in the amount of 15 µg / ml, keeping them at a temperature of 12 hours. Thereafter, the cells are separated from the supernatant layer by centrifugation. The gamma-interferon supernatant in the crude supernatant layer is 1666 IE / MP.
    For comparison, the process was performed in the same way, but before adding the inducer, the leukocytes were not pretreated with alpha interferon. In this case, the output of gamma interferon is 216 lE / ivui,
    Example 2 The process is carried out as in Example 1 with the only difference that MEM of the Hasgow type (Virology, J4, 359, 1961) is used as the nutrient solution, while the yield of gamma interferon in crude form is 1600 IE / ml
    The process described is repeated with the exception of the stage of treatment with alpha interferon. The content of active substance in untreated interferon, thus obtained, is only 206 IE / ml,
    Example 3. The process is carried out as in example 1 with the difference that the alpha interferon used for the additional treatment is not removed. The cells treated with alpha-interferon are incubated in the presence of concavalin A (concentration 15 µg / mp) at 37 ° C for 12 hours with constant stirring. The content of interferon in the supernatant is 1733 IE / ml.
    The resulting product, containing alpha- and gamma-interferon, is separated into individual components using xp chromatography. For this, the crude product is introduced into a column filled with a nozzle made of CPG glass. Alpha interferon and impurities are passed through the column, and gamma interferon is delayed. Elution of gamma interferon is carried out with an aqueous solution containing 20 vol, ethylene glycol, 150 mmol sodium chloride and 20 mmol phosphate buffer.
    Example 4 The process is carried out analogously to example 1 with the difference that 2500 IE / ml beta interferon is used for pretreatment instead of alpha interferon. The content of gamma-interferon in the irradiated
    the raw product is 1866 IE / MP.
    The described process is repeated with the exception of the pre-treatment stage with beta-interferon. The content of active substance in the crude interferon thus obtained is 226 IE / ml.
    Example 5. The preparation of gamma-interferon is carried out as in example 1 with the only difference that the preliminary treatment is carried out with alpha-interferon at a concentration of 500 1E / mp, while gamma-interferon was obtained in an amount of 700 IE / ml.
    Example 6. The process is carried out as in Example 1, only for pretreatment treatment using beta interferon at a concentration of 500 IE / ml, and the yield of gamma interferon is 1000 IE / ml.
    Example 7. The process is carried out as in Example 1 with the only difference that alpha interferon at a concentration of 5000 IE / MP is used to process leukocytes, and the output of gamma interferon is 300 IE / ml.
    Example 8. The preparation of gamma-interferon is carried out as in example 1, while leukocyte treatment is carried out with interferon-beta interferon with a concentration of 5000 IE / ml. The output of gamma interferon in this case is 1000 IE / m.
    During the processing of alpha- or beta-interferon leukocytes for 30 minutes rather than 4 hours as in example 1, the production of gamma-interferon decreases from 1,500 to
    1A05691
    250 IE / MP, and holding it for 15 hours does not exceed the output of gamma-inter-. feron, equal to 1550 IE / ml.
    The advantage of the proposed method is that due to the pretreatment. or beta interferon, the yield of gamma interferon increases 5-10 times. IC In its physicochemical properties (sensitivity at pH 2.0; stability determined at 56 and) and biological activity (antiviral and anti-cell effect), obtained by the proposed method, gamma-interferon is completely equivalent to gamma-interferon, obtained in a known manner.
    权利要求:
    Claims (1)
    [1]
    Invention Formula
    The method of obtaining human gamma-interferon by separating the blood leukocyte fraction from the blood, removing the red blood cells, suspending the white blood cells in a nutrient medium, followed by treating them with a mitogenic agent and separating the cells by centrifugation, in order to increase the yield of the target product, removal of red blood cells is carried out with 0.83 - 0.89% aqueous solution of ammonium chloride, and before being treated with a mitogenic agent, leukocytes are treated with alpha or beta interferon in an amount of 500-5000 IE / ml for 1 -12 hours
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    同族专利:
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    WO1983001899A1|1983-06-09|
    FI832508A0|1983-07-08|
    AU1017483A|1983-06-17|
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    GB2123007B|1985-01-30|
    JPS58502032A|1983-12-01|
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    引用文献:
    公开号 | 申请日 | 公开日 | 申请人 | 专利标题
    US3970749A|1973-04-03|1976-07-20|Baugh Clarence L|Interferon production|
    FR2244543B1|1973-07-27|1977-07-01|Inst Nl Sante Rech Medica|
    GB1464939A|1974-01-11|1977-02-16|Anvar|Process for the production of interferon|
    CS215108B2|1977-07-15|1982-07-30|Wellcome Found|Method of making the interferrone|
    SU839547A1|1979-04-06|1981-06-23|Всесоюзный Научно-Исследовательскийинститут Биологического Приборострое-Ния|Method of obtaining leukocytic interferon|EP0145775B1|1983-06-13|1989-02-08|Ragnvald Erik Lindblom|A METHOD OF TREATING INTERFERON SENSITIVE DISEASES, AND A METHOD AND A DEVICE FOR PREPARING A $g-INTERFERON CONTAINING PREPARATION|
    HU192254B|1983-12-13|1987-05-28|Egyt Gyogyszervegyeszeti Gyar|Process for producing human leucocite and human gamma interferons in consecutive steps|
    EP0197149B1|1984-09-21|1990-05-02|Vsesojuzny Nauchno-Issledovatelsky Institut Osobo Chistykh Biopreparatov|Method of obtaining human leukocytic interferon|
    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|
    JP2969461B2|1988-07-23|1999-11-02|株式会社林原生物化学研究所|Human γ-interferon, its production method and use|
    DE102012209673A1|2012-06-08|2013-12-24|Artcline Gmbh|A method for producing a leukocyte preparation and leukocyte preparation|
    法律状态:
    优先权:
    申请号 | 申请日 | 专利标题
    HU813594A|HU184972B|1981-12-01|1981-12-01|Process for preparing human gamma interferone|
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