Interleukin-6 (IL-6) expression and secretion， induced by inflammatory processes， stimulate the acute phase response cascade. The overexpression of IL-6 contributes to a variety of inflammatory diseases， e.g. rheumatoid arthritis， Castleman's disease， multiple myeloma， and prostate cancer. Screening for high amounts of IL-6 in the patients' blood serum can be crucial for an adequate treatment. In this study， five novel murine monoclonal antibodies (mAbs) reactive to human IL-6 were generated. The mAbs were characterized for potential diagnostic purposes and recombinant antibodies were derived thereof. Initial epitope mapping using a combination of blocking experiments and Hyper-IL-6， a fusion pr... More
Interleukin-6 (IL-6) expression and secretion， induced by inflammatory processes， stimulate the acute phase response cascade. The overexpression of IL-6 contributes to a variety of inflammatory diseases， e.g. rheumatoid arthritis， Castleman's disease， multiple myeloma， and prostate cancer. Screening for high amounts of IL-6 in the patients' blood serum can be crucial for an adequate treatment. In this study， five novel murine monoclonal antibodies (mAbs) reactive to human IL-6 were generated. The mAbs were characterized for potential diagnostic purposes and recombinant antibodies were derived thereof. Initial epitope mapping using a combination of blocking experiments and Hyper-IL-6， a fusion protein consisting of IL-6 and the soluble IL-6 receptor revealed distinct but overlapping binding sites. At least one of the mAbs was found to interact with the region of IL-6/ IL-R complex formation. Three mAbs were applied successfully in intracellular staining by flow cytometry， whereas one of the mAbs showed comparable binding as a reference reagent. Furthermore， the mAbs were tested for applications in various immunological assays such as ELISA， Western blot and surface plasmon resonance spectroscopy (SPR)， using IL-6 from commercial sources as well as in-house produced protein (IL-6_IME). The limit of detection was determined by sandwich ELISA (0.5 ng/mL， SD ±0.005). Our results also demonstrated that the recombinant IL-6 produced was functional and correctly folded. These findings support the use of the generated mAb clones as promising candidates for application in various immunological assays for diagnostic and scientific purposes.
