CS277142B6 - Mouse hybridoma IFNA1-48 producing a monoclonal antibody - Google Patents

Mouse hybridoma IFNA1-48 producing a monoclonal antibody Download PDF

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CS277142B6
CS277142B6 CS905554A CS555490A CS277142B6 CS 277142 B6 CS277142 B6 CS 277142B6 CS 905554 A CS905554 A CS 905554A CS 555490 A CS555490 A CS 555490A CS 277142 B6 CS277142 B6 CS 277142B6
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Czechoslovakia
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human interferon
interferon alpha
ifna1
monoclonal antibody
mouse hybridoma
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CS905554A
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Czech (cs)
Slovak (sk)
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CS555490A3 (en
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Peter Rndr Csc Kontsek
Michal Mvdr Csc Novak
Ladislav Akademik Borecky
Eva Rndr Kontsekova
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Virologicky Ustav Sav
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Priority to CS905554A priority Critical patent/CS277142B6/en
Publication of CS555490A3 publication Critical patent/CS555490A3/en
Publication of CS277142B6 publication Critical patent/CS277142B6/en

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Abstract

Příprava novej homogénnej protilátky, ktorá neutralizuje a viaže ludský interferon alfa 1 a nerozlišuje iné subtypy ludského interferonu alfa. Uvedeného účelu sa dosiahne použitím nového myšieho hybridómu IFNA1-48, produkujúceho monoklonálnu protilátku imunoglobulínovej podtriedy Gl voči rekombinantnému ludskému interferonu alfa 1. Myší hybridóm IFNA1-48 sa využívá v imunodiagnostike a pri čistění ludského interferonu alfa 1.Preparation of a novel homogeneous antibody that neutralizes and binds human interferon alpha 1 and does not discriminate between other subtypes of human interferon alpha. The above purpose is achieved by using a novel mouse hybridoma IFNA1-48, producing a monoclonal antibody of the immunoglobulin subclass G1 against recombinant human interferon alpha 1. The mouse hybridoma IFNA1-48 is used in immunodiagnostics and in the purification of human interferon alpha 1.

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1 CS 277142 B61 CS 277142 B6

VIN

Vynález sa týká nového myšieho hybridómu IFNA1-48 produkujú-ceho monoklonálnu protilátku, ktorá neutralizuje a viaže ludskýinterferon alfa 1.The present invention relates to a novel mouse hybridoma IFNA1-48 producing a monoclonal antibody that neutralizes and binds human interferon alpha 1.

Protilátky voči ludskému interferonu alfa i sa připravujú voformě antisér alebo monoklonálnych protilátok. Antiséra sa získa-vajú imunizáciou pokusných zvierat purifikovaným antigénom (lud-ským interferonom alfa 1). Takto připravené protilátky sú nestan-dardně a jednotlivé šarže antisér majú rozdielne vázobnéa neutralizačně schopnosti voči ludskému interferonu alfa 1.Antiséra vykazujú aj krížovú reaktivitu s příbuznými subtypmiÍudského interferonu alfa. Přípravu homogénnych protilátoks neutralizačnou a súčasne s vázobnou aktivitou voči ludskémuinterferonu alfa 1 umožňuje iba hybridómová technika přípravymonoklonálnych protilátok. Takéto monoklonálne protilátky umožňu-jú dokaž přítomnosti íudského interferonu alfa i v róznych biolo-gických materiáloch pomocou citlivých neutralizačných a imunoche-mických stanovení. Hybridómy sa získavajú známým spósobom opísa-ným v literatúre [G. Kóhler, C. Milstein: Nátuře, 256 (1975)].Hybridóm IFNA1-48 nebol doteraz připravený.Antibodies to human interferon alpha 1 are prepared in antisera or monoclonal antibodies. Antisera are obtained by immunizing experimental animals with purified antigen (human interferon alpha 1). The antibodies thus prepared are non-standard and individual batches of antisera have different binding and neutralizing capabilities to human interferon alpha 1. The antisera also exhibits cross-reactivity with related human interferon alpha subtypes. Only the hybridoma technique of preparymonoclonal antibodies allows the preparation of homogeneous antibodies by neutralizing and simultaneously binding to human interferon alpha 1. Such monoclonal antibodies allow the presence of human interferon alpha to be detected in various biological materials by sensitive neutralization and immunochemical assays. Hybridomas are obtained in a manner known in the literature [G. Köhler, C. Milstein: Nature, 256 (1975)] The IFNA1-48 hybridoma has not yet been prepared.

Nevýhody konvenčně připravených antisér voči ludskému inter-feronu alfa i v podstatnej miere odstraňuje vynález, ktoréhopodstatou je myší hybridóm IFNA1-48, produkujúci monoklonálnuprotilátku imunoglobulínovej podtriedy G1 voči rekombinantnémuludskému interferonu alfa 1. Hybridóm IFNA1-48 je uložený voVirologickom ústave SAV, Dúbravská cesta, 842 46 Bratislava. Výhodou hybridómu IFNA1-48 je, že produkuje homogénnu proti-látku, ktorá je schopná špecificky neutralizovat a viazat ludskýinterferon alfa 1 a nerozoznáva iné subtypy Íudského interferonualfa. Po rozmražení buňky hybridómu IFNA1-48 pokračujú v produk-ci! protilátky i bez áalšej imunizácie ludským interferonomalfa 1. PříkladThe disadvantages of conventionally prepared antisera to human alpha interferon are substantially eliminated by the invention, which is essentially the murine hybridoma IFNA1-48, producing a monoclonal antibody of the G1 immunoglobulin subclass against recombinant human interferon alpha 1. The IFNA1-48 hybridoma is deposited in the SAS Institute of Virology, Dubravska cesta, 842 46 Bratislava. The advantage of the IFNA1-48 hybridoma is that it produces a homogeneous anti-substance that is able to specifically neutralize and bind human interferon alpha 1 and does not recognize other subtypes of human interferonualpha. After thawing, the IFNA1-48 hybridoma continues to produce! antibodies even without further immunization with human interferon 1. Example

Hybridómy sa získajú fúziou myších myelómových buniek NSOa buniek získaných zo sleziny myší kmeňa BALB/c imunizovanýchrekombinantným ludským interferonom alfa 1 (Ernst Boehringer--Institute fur Arzneimittelforschung, Viedeň). Po fúzii bolvybratý hybridóm IFNA1-48, ktorý produkuje monoklonálnu protilát-ku imunoglobulínovej podtriedy Gl, selektívne neutrálizujúcua viažúcu ludský interferon alfa 1. Hybridóm IFNA1-48 sa pěstujein vitro v kultivačných médiách alebo in vivo v peritoneálnejdutině myší kmeňa BALB/c. Buňky sa zmrazujú a uchovávajú v teku-tom dusíku a po rozmražení pokračujú v produkcii protilátky bezčfalšej imunizácie ludským interferonom alfa 1. Buňky hybridómuIFNA1-48 rastů in vitro ako suspenzná kultúra a produkujú do kul-tivačného média přibližné 1 μg/ml špecifickej protilátky. Kulti-vačným médiom je Dulbeccova modifikácia Eaglovho minimálnehoesenciálneho média [R. Dulbecco, G. Freeman: Virology 8, 396(1959)]. Toto médium je pre kultiváciu hybridómov doplněné inak-tivovaným koňským sérom (ELÁN CLONĚ). Za účelom získania váčšiehomnožstva monoklonálnej protilátky voči ludskému interferonualfa 1 sa buňky IFNA1-48 aplikovali v množstve 5.106 do perito-neálnej dutiny myši BALB/c. Myš sa desat dní před touto apliká-Hybridomas were obtained by fusing murine NSO mouse myeloma cells and spleen cells from BALB / c mice immunized with recombinant human alpha 1 interferon (Ernst Boehringer-Institute fur Arzneimittelforschung, Vienna). After fusion, the IFNA1-48 hybridoma, which produces the monoclonal antibody immunoglobulin G1 subclass, selectively neutralizing human interferon alpha 1, was selected. The IFNA1-48 hybridoma was grown in vitro in culture media or in vivo in the peritoneal cavity of BALB / c mice. Cells are frozen and stored in liquid nitrogen and, after thawing, continue to produce antibody to blunder immunization with human interferon alpha 1. HybridomaIFNA1-48 growth cells in vitro as a suspension culture and produce approximately 1 µg / ml specific antibody to culture medium. The culture medium is Dulbecco's modification of Eagle's minimal essential medium [R. Dulbecco, G. Freeman: Virology 8, 396 (1959)]. This medium is supplemented with inactivated horse serum for the cultivation of hybridomas (ELAN CLONA). In order to obtain a higher amount of monoclonal antibody to human interferon 1, IFNA1-48 cells were applied at 5 x 10 6 to the BALB / c mouse peritoneal cavity. The mouse will be ten days prior to this

Claims (2)

CS 277142 B6CS 277142 B6 2 Λ ciou premedikuje parafínovým olejom (0,5 ml intraperitoneálne namyš). Po 10 až 15 dňoch od injekcie buniek IFNA1-48 bola myšiodobratá ascitická tekutina, ktorá obsahovala požadovaná monoklo-nálnu protilátku v koncentrácii 1,1 mg/ml. Myší hybridóm IFNA1-48 sa používá ako zdroj homogénnejprotilátky imunoglobulínovej podtriedy Gl, ktorá špecificky viažea neutralizuje Iudský interferon alfa 1, pričom nereaguje s inýmisubtypmi ludského interferonu alfa. Připravená protilátka sapoužívá na detekciu ludského interferonu alfa i v biologických(neutralizačných) a imunochemických (RIA, ELISA, imunoblot)testoch. Protilátka je vhodná aj na imunoafinitnú purifikáciuludského interferonu alfa 1. PATENTOVÉ NÁROKY Myší hybridóm IFNA1-48 produkujúci monoklonálnu protilátkuimunoglobulínovej podtriedy Gl voči rekombinantnému ludskémuinterferonu alfa 1. Konec dokumentuIt is pretreated with paraffin oil (0.5 ml intraperitoneally). 10-15 days after injection of IFNA1-48 cells, the ascites fluid was collected from the mouse, containing the desired monoclonal antibody at 1.1 mg / ml. Mouse hybridoma IFNA1-48 is used as a source of a homogeneous antibody of the immunoglobulin G1 subclass that specifically binds to neutralize human interferon alpha 1 without reacting with other human interferon alpha subtypes. The prepared antibody is used for the detection of human interferon alpha in biological (neutralizing) and immunochemical (RIA, ELISA, immunoblot) assays. The antibody is also suitable for immunoaffinity purification of human interferon alpha 1. PATENT REQUIREMENTS Mouse hybridoma IFNA1-48 producing a monoclonal antibody immunoglobulin G1 subclass against recombinant human interferon alpha 1. End of document
CS905554A 1990-11-12 1990-11-12 Mouse hybridoma IFNA1-48 producing a monoclonal antibody CS277142B6 (en)

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