FIG. 3.

Analysis of cytokine production by ELISA. (A) IFN-γ ELISA from the supernatant of splenocytes (10⁵) of mice infected orally with Y. enterocolitica. Splenocytes received the indicated treatment: no further treatment, 10 μg of heat-killed Y. enterocolitica/ml, or 5 μg of ConA/ml for 3 days. The concentration of IFN-γ secreted into the culture supernatant was determined by sandwich ELISA according to the manufacturer's instructions. Splenocytes from C57BL/6j mice are represented by black bars, and those from IL-6^−/− mice are represented by white bars. (B) TGF-β1 ELISA from the supernatant of splenocytes (10⁵ cells) of mice orally infected with Y. enterocolitica. The concentration of TGF-β1 secreted into the culture supernatant was determined by sandwich ELISA according to the manufacturer's instructions. (C) IFN-γ ELISA. Splenocytes from infected mice (C57BL/6j mice, IL-6^−/− mice, and IL-6^−/− mice treated with 1 μg of rIL-6 SC every day for 5 days) were used for ELISA. Splenocytes (10⁵ cells) received the indicated treatment, i.e., no further treatment or 5 μg of ConA/ml. The concentration of IFN-γ secreted into the culture supernatant was determined by sandwich ELISA according to the manufacturer's instructions. The asterisks indicate comparisons that were not statistically significant. (D) TGF-β1 ELISA. Mice were treated as described for panel C except that ConA was not added. Subsequently, splenocytes (10⁵) from these mice were cultured for 3 days. The concentration of TGF-β1 secreted into the culture supernatant was determined by sandwich ELISA according to the manufacturer's instructions. The results are the average of four assays done in duplicate. The error bars represent the standard deviation, and statistically significant comparisons (as determined by a Mann-Whitney nonparametric two-tailed ANOVA) are indicated by brackets.