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INFLAMMATION, CYTOKINES, AND TEMPERATURE REGULATION
Innovative Methodology

Interleukin-10 concentration determined by sandwich enzyme-linked immunosorbent assay is unrepresentative of bioactivity in murine blood

Department of Human Biology and Nutritional Sciences, University of Guelph, Guelph, Ontario, Canada N1G 2W1

Submitted 9 July 2003 ; accepted in final form 8 August 2003

Two experiments were performed, each using six male and six female C57BL/6J mice collectively ranging from 4 wk to 17 mo of age. Blood was obtained following CO₂ anesthesia, and the IL-10 concentration of each serum sample was determined both by sandwich enzyme-linked immunosorbent assay (ELISA) and by bioassay. In the first experiment, mean serum IL-10 immunoactivity was 9.3 pg/ml while the mean bioactivity was 700 times greater, i.e., 6.5 ng/ml. However, the bioassay required sample dilution, which might have released bound cytokine that the ELISA could also detect. In the second experiment, therefore, the ELISA was applied to samples diluted to 20% as for the bioassay. Nevertheless, the immunoassay continued to detect only a small fraction of the serum IL-10 identified by the bioassay (mean values: 32.4 pg/ml vs. 2.6 ng/ml). Although currently the preferred method, the sandwich ELISA is inappropriate for quantification of blood IL-10 concentrations. Moreover, studies of the actions of IL-10 are needed at the concentrations revealed in the blood by bioassay and currently considered supraphysiological.

bioassay; cytokine; immunoactivity; mouse

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