Preparing a Single Cell Suspension from Zebrafish Retinal Tissue for Flow Cytometric Cell Sorting of Muller Glia

Preparation of a single cell suspension from solid tissue is vital for a successful flow cytometry experiment. We report a detailed and reproducible method to produce a quality cell suspension from the zebrafish retina. Zebrafish retinas, especially their Muller glia cells, are of particular interest for their inherent regenerative capacity, making them a useful model for regenerative medicine and cell therapy research. Here, we detail a papain-based dissociation that is gentle enough to keep cells intact, but strong enough to disrupt cell-cell and cell-matrix interactions to yield a cell suspension that produces clean and reliable flow cytometric cell sorting results. This procedure consistently results in over 90% viability and three populations of cells based on GFP expression. The dissociation procedure described herein has been optimized for the collection of Muller glia fromTg(apoe:gfp)zebrafish retinas; however, the overall process may be applicable to other cell types in the fish retina, additional flow cytometric techniques, or preparing cell suspensions from similar tissues. (c) 2019 International Society for Advancement of Cytometry