PCNA Staining of Retinal Cryosections to Assess Microglial/Macrophage Proliferation

Detection of the protein PCNA (proliferating cell nuclear antigen) is used to identify cells in the S phase of the cell cycle to indicate cellular proliferation. Here we describe our method to detect PCNA expression by microglia and macrophages in retinal cryosections. We have used this procedure with zebrafish tissue, but this procedure could be applied to cryosections from any organism. Retinal cryosections are subjected to a heat-mediated antigen retrieval step in Citrate Buffer, then immunostained with antibodies to label PCNA and microglia/macrophages, and counterstained for cell nuclei. After fluorescent microscopy, the number of total and PCNA+ microglia/macrophages can be quantified and normalized to compare across samples and groups.