DETERMINATION OF INTERLEUKIN-6 IN HUMAN URINE AND EPITHELIAL-CELL SUPERNATANTS

Interleukin 6 (IL-6) is a cytokine with many biological functions. It is produced by different tissues in response to infection and is secreted into the local body fluids. The aim of this study was to find a suitable assay to measure IL-6 in human urine. IL-6 was quantitated by a bioassay and by immunoassays based on neutralizing or nonneutralizing antibodies. The effect of human urine on the quantitation of IL-6 by these assays was analyzed using pooled human urine with added recombinant or natural human IL-6. Urine was found to disturb the growth of the B9 cells. When fractions from gel-filtered human urine were tested, a fraction corresponding to a protein molecular weight range of 10,000-1,000 was found to have a strong inhibitory effect in the B9 assay. In contrast, the low molecular weight fractions containing salts and pigments were not found to disturb the assay. The inhibitory effect of urine was avoided by diluting the samples > 80 times (final dilution in the test plate) or by dialysis. Furthermore, we analyzed IL-6 in urine samples from patients with urinary tract infection and supernatants from epithelial cells stimulated with bacteria in vitro. The B9 assay and the immunoasay based on non-neutralizing anti-IL-6 antibodies were more sensitive than the immunoassays based on neutralizing antibodies. While most of the B9 activity in the urine samples and supernatants could be neutralized by anti-IL-6 antibodies, some samples contained unneutralizable activity. These components remain to be defined. The results demonstrate considerable variation between assays used to quantitate natural IL-6.