Development of polyclonal and monoclonal antibodies for immunoassay and neutralization of human interleukin-4
| Autor: | John S. Abrams, I Chrétien, A Van Kimmenade, Jacques Banchereau, M K Pearce |
|---|---|
| Rok vydání: | 1989 |
| Předmět: |
Male
medicine.drug_class Molecular Sequence Data Immunology Monoclonal antibody law.invention Immunoenzyme Techniques Neutralization Tests law medicine Animals Humans Immunology and Allergy Avidity Amino Acid Sequence Antiserum medicine.diagnostic_test biology Immune Sera Interleukins Antibodies Monoclonal Molecular biology Recombinant Proteins Rats Rats Inbred Lew Cell culture Polyclonal antibodies Immunoassay Recombinant DNA biology.protein Interleukin-4 Rabbits Antibody |
| Zdroj: | Journal of Immunological Methods. 117:67-81 |
| ISSN: | 0022-1759 |
| DOI: | 10.1016/0022-1759(89)90120-8 |
| Popis: | A rabbit antiserum to partly purified recombinant E. coli-expressed human interleukin-4 (IL-4) has been produced which neutralizes the T cell growth factor, B cell growth factor, and Fc epsilon R2/CD23 inducing activities of IL-4. The antiserum demonstrated sufficient avidity to immunoprecipitate labelled COS7-expressed recombinant human IL-4. In contrast, rabbits immunized with conjugates of various synthetic IL-4 oligopeptides produced antisera which recognized IL-4 in both enzyme-linked immunosorbent assay (ELISA) and Western blotting formats, but failed to immunoprecipitate IL-4 from solution, or to neutralize bioactivity. Two rat monoclonal antibodies, 11B4, 22C10 were produced from a rat immunized with purified COS7 cell-expressed IL-4. These IgG2a antibodies recognized both E. coli-expressed and mammalian cell-expressed (COS7 and L cell) recombinant human IL-4 in solution (immunoprecipitation), as well as on solid phase (indirect ELISA and dot-blotting). The 11B4 antibody inhibited IL-4 bioactivity at an IC50 which was 25-50-fold in molar excess of factor. Both antibodies also recognized IL-4 bound to an immobilized rabbit IgG fraction of anti-IL-4. The 11B4 antibody was used to develop an immunoenzymetric assay capable of detecting less than 100 pg of analyte/ml. Supernatants from PBL, activated under varying conditions were tested for IL-4 levels. PHA and ConA were found to induce a relatively low degree of IL-4 production by these PBL. An approximately ten-fold greater level of IL-4 production was observed when they were stimulated with A23187 in combination with PMA. Various patient sera and cell line supernatants were also tested. These IL-4 immunoreagents are important tools for further studies of IL-4 immunobiology. |
| Databáze: | OpenAIRE |
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