• 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:
    The invention relates to a process for the industrial production of a product for the diagnosis of carcinoma consisting of a antigen obtained by the infection with Herpes Simplex Virus of cells of guinea-pig kidney the process comprising three main section in the first of which the antigen is produced in the second section a biochemical purification of the antigen is carried out, while in the third section the product is prepared in preservable condition and confection.
    公开号:SU1213974A3
    申请号:SU802918702
    申请日:1980-05-13
    公开日:1986-02-23
    发明作者:Тарро Джулио
    申请人:Депа С.П.А. (Фирма)；
    IPC主号:
    专利说明:

    37 ° C
    This invention relates to medicine, in particular to immunology.
    The aim of the invention is to provide an industrial method for producing the herpes simplex virus antigen used in the diagnosis of tumors.
    Example. Kidneys are removed from guinea pigs at the age of two weeks. The kidneys are freed from the membranes and the loose layer and ground, then added from trypsin in an amount of approximately 200 ml for every 10 tablets in the presence of lactalbumin and bovine serum.
    This suction is kept for 1 hour under the conditions of first mixing. Then the suspension is centrifuged for 10 minutes at a speed of 1500. The supernatant is removed, and the settled cells are washed with the addition of nutrient medium (lactalbush) into the same tube, then another centrifugation is performed. The supernatant is again removed, and the cells are suspended in 10 micronutrient medium in the same tube. Thereafter, the suspension is ground into a smaller centrifugation tube, which is conducted at a speed of 1000 rpm.
    A dilution of the cells is prepared in a ratio using the nutrient medium of the bodies (whose serum is taken in a concentration of 10%. Then the suspension is transferred into rotating flasks, which are placed in a thermostat with a temperature of 37 ° C for 2/3 days. During this period, the nutrient medium of the TM ™ medium is added once. Then the suspension is centrifuged at a speed of 2000 rpm for 10 in, until a precipitate forming the cell cake is obtained at the bottom of the tube.
    placed in a rotating flask, then herpes simplex virus (HSV) is added, mixing the contents at a rate of 50-60 for 3 hours at 37 ° C. During this period, an antigen is formed, which will then be referred to as TAP, at the end of this stage a cell cake is obtained,
    VNIIPI Order 788/63 Circulation 659 Subscription Branch PIP Patent, Uzhgorod, Project Design St., A
    ii
    0
    five
    0
    five
    Cell leptosh is stored in a spike at -80 ° C, from which
    containing TAG in its raw form. 1.5 ml of cells can be obtained from 10 guinea pigs. To these cells, 1.5 ml of a tris buffer was added to obtain a 50% cell pellet.
    sea boy
    can be obtained in order to start the purification of TAP contained in 3 ml
    0 mixture from impurities. To do this, cells are disintegrated by freezing and thawing, repeating these procedures three times, and sonication (0.9 total capocycles, 3 times
    5 to 3 min. The liquid phase is separated by centrifugation at 27,000 rpm, 20 mM three with -HCf buffer solution, pH 72, is added to it, until the bulk volume is mixed, again
    0 is centrifuged at 50,000 rpm,
    the supernatant is saturated to 80% with powdered ammonium sulfate. The resulting mixture was centrifuged at 15,000 rpm. Tothe buffer is added to the formed precipitate, the suspension is passed through a column, filled with DEAE-Sephadex D 50, at room temperature, eluted with Tris-HCF buffer. Then it is passed through a column filled with Sephadex G-100 5 and eluted with a 7ris -HCf buffer mixture (20 1 1 / l) j, then chromatographed on DEAE-sepha-dex A 60, washed with a tris-HN solution (20 mmol / l ), eluted with a solution of NaCj in Tris-HH (30 Yol / L), NaC5 was removed from the eluted powder using dialysis CHCTiaMbi DS-2, non-glycated proteins were removed using affinity chromatography on a column filled with Sepharose resin, and bound with concanavalin A by metalloprotein in the presence of a 3% aqueous solution of oL -tip-O-mannose and dialyzed to remove pos- .lednego vschel dissolved and the desired product ..
    At the end of the process, 288 μg of TAF with mol. m, 70,000 daltons, distilled water is added and the resulting solution is lyophilized.
    权利要求:
    Claims (1)
    [1]
    METHOD FOR OBTAINING HERPES SIMPLEX VIRUS ANTIGEN THE DIAGNOSIS OF TUMORS, which consists in treating kidneys of guinea pigs with trypsin in the presence of lactalbumin and calf serum, the obtained cells are cultured in a dynamic state in a nutrient medium, then they are infected with Herpes Simplex virus supplemented with and cultured with vigorous stirring at a speed of 50-60 rpm for 3 hours, then the cells are disintegrated by sequential freezing and thawing, and ultrasonic treatment, the liquid phase is separated add to it 20 mm Tris-HN buffer solution, pH 7.2 to 40% volume saturation, the resulting mixture was centrifuged at 50,000 rpm, the supernatant was saturated with 80% powdered ammonium sulfate, the mixture was centrifuged at 15,000 rpm, the same buffer was added to the resulting precipitate, passed through column filled with DEAE-Sephadex A 50, at room temperature, elute Tris-HC with a buffer, then passed through a column filled with Sephadex G-100, and eluted with the same buffer with NaCl added, the resulting eluate was dialyzed, non-glycosylated proteins were removed by affinity chromatography on a column filled with Sepharose resin bound to concaavalin A via metalloprotein B the presence of a 3% C aqueous solution of α-methyl-D-mannoside with successive removal of the latter and the glycoprotein mol. m. 70,000 daltons.
    12131>
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    同族专利:
    公开号 | 公开日
    ZA802935B|1981-06-24|
    CA1172562A|1984-08-14|
    JPS55158559A|1980-12-10|
    AT10102T|1984-11-15|
    US4391911A|1983-07-05|
    IT7949082D0|1979-05-18|
    ES8104404A1|1981-04-16|
    EP0016735A3|1981-04-15|
    EP0016735B1|1984-10-31|
    IL59539D0|1980-06-30|
    BR8003165A|1980-12-30|
    IT1117218B|1986-02-17|
    ES491553A0|1981-04-16|
    IL59539A|1983-10-31|
    EP0016735A2|1980-10-01|
    DE3069536D1|1984-12-06|
    EG14898A|1985-06-30|
    引用文献:
    公开号 | 申请日 | 公开日 | 申请人 | 专利标题
    GB1535317A|1976-02-20|1978-12-13|Mun Hon Ng W|Anti-cancer vaccines|
    IT1117218B|1979-05-18|1986-02-17|Depa Sas|PROCEDURE FOR THE PRODUCTION OF A PRODUCT FOR THE DIAGNOSIS OF TUMORS OF THE SOFT PARTS OR CARCINOMES AND PRODUCT OBTAINED|US4452734A|1980-02-11|1984-06-05|Merck & Co., Inc.|Herpes subunit vaccine|
    IT1117218B|1979-05-18|1986-02-17|Depa Sas|PROCEDURE FOR THE PRODUCTION OF A PRODUCT FOR THE DIAGNOSIS OF TUMORS OF THE SOFT PARTS OR CARCINOMES AND PRODUCT OBTAINED|
    US4572896A|1980-08-27|1986-02-25|The United States Of America As Represented By The Department Of Health And Human Services|Monoclonal antibodies to herpes simplex virus type I polypeptides|
    JPS6051120A|1983-08-31|1985-03-22|Chemo Sero Therapeut Res Inst|Herpes simplex subunit vaccine|
    AT106565T|1984-10-31|1994-06-15|Alcon Lab Inc|SANDWICH IMMUNOTEST PROCEDURE FOR ANTIGENS IN OPHTHALMIC DISORDERS.|
    US4786590A|1985-01-15|1988-11-22|California Institute Of Technology|Diagnostic and therapeutic aspects of receptor-mediated leukemogenesis|
    ES2196026T3|1993-04-30|2003-12-16|Wellstat Biologics Corp|USE OF NDV IN THE MANUFACTURE OF A MEDICINAL PRODUCT TO TREAT CANCER.|
    法律状态:
    优先权:
    申请号 | 申请日 | 专利标题
    IT4908279A|IT1117218B|1979-05-18|1979-05-18|PROCEDURE FOR THE PRODUCTION OF A PRODUCT FOR THE DIAGNOSIS OF TUMORS OF THE SOFT PARTS OR CARCINOMES AND PRODUCT OBTAINED|
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