IL-20R2 knockout mice remain proficient in IL-24 ligand binding in the epidermis. (a) Expression and characterization of mouse IL-24-AP (mIL-24-AP) fusion protein. Ten microliters of 1 unit per milliliter of either mIL-24-AP or AP control protein were detected by western blot analysis using antibody specific to AP. (b) Expression and characterization of mouse IL-24-Fc (mIL-24-Fc) fusion protein. Two micrograms of either IL-24-Fc or TNFR2-Fc as a negative control were separated on a SDS-PAGE and either stained by Coomasie Blue or detected by in situ affinity staining with human IL-20R2-AP fusion protein7 after transferred to a PVDF membrane. (c) Paraffin-embedded skin sections of WT (top) and IL-20R2−/− (bottom) mice were analyzed by either hematoxylin and eosin staining or in situ ligand-receptor binding to either mIL-24-AP or AP control to detect the expression of IL-24 receptors. Note the intense staining of the epidermis by mIL-24-AP of both WT and IL-20R2 knockout mice. The specificity of mIL-24-AP staining was confirmed by competition with 10-fold excess of mIL-24-Fc, which essentially completely blocked the mIL-24-AP binding to the epidermis. The same amount of TNFR2-Fc served as a negative control for the specificity for the competition experiment.