Assays were performed as previously reported [20 (link)]. In brief, for the IF experiment, cells were incubated with primary antibodies against CCL20 (1:200, #ab9829) (Abcam) at 4 °C overnight, then incubated with the fluorescent secondary antibody Alexa Fluor 594-conjugated goat anti-mouse IgG (Cell Signaling Technology) and imaged using a fluorescence microscope (DM5000B, Leica). For IHC, the paraffin sections were incubated with antibodies against CCL20 (1:200, #ab9829) (Abcam). Intensity scores were recorded as: 0 (no staining), 1 (weakly staining, light yellow), 2 (moderately staining, yellowish brown), and 3 (strongly staining, brown). In addition, we also detected the expression of ki67 (1:1000, #9449), CD68 (1:200, #97778), CD163 (1:400, #93498), CD206 (1:200, #24595) and CD31 (1:100, #77699) (Cell Signaling Technology) in PCa tissue or xenograft tissue. Images were observed under an Olympus multifunction microscope (Olympus BX51, Tokyo, Japan). All evaluations were performed by three independent senior pathologists using the same microscope.
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