Bioassay for interleukin-1, interleukin-6, and tumor necrosis factor-like activities in canine sera

To measure interleukin (IL)-1, IL-6, and tumor necrosis factor (TNF)-like activities in canine serum, bioassays were conducted using human melanoma A375S1, IL-6 dependent murine hybridoma MH60.BSF2, and WEHI 164 murine sarcoma subclone 28-4. Clinically normal adult beagles were experimentally induced endotoxic shock by an intravenous injection of lipopolysaccharide or local inflammation by an intramuscular injection of turpentine oil. IL-1-like activity was detected in sera from dogs with endotoxic shock. IL-6 and TNF-like activities were detected in sera from both dogs with endotoxic shock and local inflammation. IL-1-like activity in sera from the dogs with endotoxic shock declined after dilution with either medium or serum obtained before treatment (pre-serum), but the IL-1-like activity was maintained to a greater extent in samples diluted with pre-serum compared to those diluted with medium. TNF-like activity declined equally after dilution with either medium or pre-serum. On the other hand, IL-6-like activity was inhibited at low dilution. It was, therefore, necessary to dilute the serum samples to 1:180 from dogs with endotoxic shock or 1:60 from dogs with local inflammation, in order to minimize the effect of inhibitory factors on IL-6-like activity. IL-6-like activity was neutralized by monoclonal antibody against murine IL-6 receptors. TNF-like activity was neutralized by anti-mouse TNF alpha rabbit serum. However IL-1-like activity was not neutralized by either anti-mouse or anti-human IL-1 rabbit serum.