Establishment of an interleukin 6 (IL 6)/B cell stimulatory factor 2‐dependent cell line and preparation of anti‐IL 6 monoclonal antibodies

Abstract

Murine hybridomas producing monoclonal antibodies (mAb) specific to human interleukin 6 (IL6/BSF‐2) were established. One of these hybridomas (MH60.BSF2) was found to be dependent on IL6 for its in vitro growth. None of the known biological factors tested, such as recombinant (r) human (Hu) Illα, rHuILlα, rHuIL2, rHuIL3, rHuIL4, rHu interferon (1FN)‐γ, HuIFN‐β, rHuG‐CSF, or recombinant murine (Mu) IL3, MuIL4, rMuIL5, could induce the in vitro growth of MH60.BSF2 cells. The half‐maximum tritiated thymidine uptake by MH60.BSF2 cells could be achieved by picogram amounts of rIL 6, making this hybridoma clone an indicator cell for specific and sensitive detection of the IL6 activity in test samples. The MH166.BSF2 clone was found to produce IgG l,χ type mAb (αBSF2‐166) capable of neutralizing IL6 activity. The other clone, MH60.BSF2, produced IgM,x type mAb (aBSF2‐60) unable to neutralize IL6 activity. Both mAb specifically reacted with IL6 as demonstrated by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis and Western blotting analysis. An enzyme‐linked immunosorbent assay (ELISA) utilizing αBSF2‐166 and rabbit anti‐IL 6 antibodies was established which could detect as low as 50 pg/ml of IL6. Since both the ELISA and MH60.BSF2 hybridoma could detect small amounts of IL6 in biological fluids, they constitute powerful tools in exploring the presence or the role of IL6 in various immunological disorders.