KSY neither enhances the secretion of anticancer-associated cytokines, nor increases the recruitment of tumor-infiltrating leukocytes in tumor-bearing mice. The anticancer immunity in the bladder cancer model (C3-B model in Figure 1) was evaluated. (a) The expressions of indicated cytokines in tumor tissues were measured by ELISA. (b) The splenocytes (3 × 106/ml) from each treatment group were cultured in the presence of PHA (5 μg/ml) for 72 h, and the amount of cytokines in the culture medium were measured by ELISA. (c) The culture medium collected from the splenocytes of drug-treated mice (splenocyte-conditioned medium; SCM) was incubated with MBT-2 cells for 48 h. The death of MBT-2 cells was detected by PI stain and analyzed by flow cytometry. # indicates undetectable. *indicates p < 0.05 versus the vehicle group. n = 18-21. (d) Tumor tissues from each treatment group in C3-B model were fixed, sectioned and stained with hematoxylin and eosin (H&E stain) for microscopic examination. (e) The expression of various immune cell maker genes in tumor tissues of C3-B model mice was measured using qRT-PCR. CD4: T helper cells; CD8: cytotoxic T cells; CD11b: macrophages; CD11c: dendritic cells; NK1.1: NK cells. Three independent experiments were performed (n = 3 for each group). *indicates p < 0.05 versus the vehicle group.