The co-localization of PCNA with Rab7 in RANKL-induced RAW264.7 cells. (A) IF assay was performed to localize PCNA in the 4-day RANKL-treated RAW264.7 cells using appropriate PCNA antibody. Arrows: PCNA located in the cytoplasm. Scar bar: 20μm for the top panel images and 5μm for the bottom three panels. (B) Co-localization of PCNA with Rab7 in RANKL-induced osteoclast. RAW264.7 cells were treated with RANKL (100 ng/mL) for 3 days and subjected to IF assays using PCNA and Rab7 antibodies. The Z-stack scan model of the LSM microscope was used to observe the cells from the abdominal end to dorsal end. All images were presented in order. Arrows: co-localization of PCNA with Rab7. Scar bar: 5μm for negative control (NC) cells and 10μm for RANKL treated cells. (C) Co-staining of PCNA with LAMP1 in RANKL-induced osteoclast as described in (B). Scar bar: 2μm. (D) Co-staining of PCNA with LC3 in RANKL-induced osteoclast as described in (B). Scar bar: 5μm. (E) Z-stack scan mode in confocal microscopy was used to detect the subcellular localization of PCNA and F-actin in the 3-day RANKL treated RAW264.7 cells. Upper panels: untreated negative control RAW264.7 cells; lower panels: 3-day RANKL treated RAW264.7 cells. Scar bar: 5μm.