Development of polyclonal and monoclonal antibodies for immunoassay and neutralization of human interleukin-4.
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.
Chrétien
I
I
UNICET, Laboratory for Immunological Research, Dardilly, France.
Van Kimmenade
A
A
Pearce
M K
MK
Banchereau
J
J
Abrams
J S
JS
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000032:analysis / Q000096:biosynthesis*; D006801:Humans; D007106:Immune Sera / Q000032:analysis / Q000494:pharmacology; D007124:Immunoenzyme Techniques; D015847:Interleukin-4; D007378:Interleukins / Q000138:chemical synthesis / Q000276:immunology* / Q000302:isolation & purification; D008297:Male; D008969:Molecular Sequence Data; D009500:Neutralization Tests; D011817:Rabbits; D051381:Rats; D011917:Rats, Inbred Lew; D011994:Recombinant Proteins / Q000138:chemical synthesis / Q000276:immunology / Q000302:isolation & purification
D000911:Antibodies, Monoclonal; D007106:Immune Sera; D007378:Interleukins; D011994:Recombinant Proteins; D015847:Interleukin-4