Isolation of Podocyte Cell Fractions From Mouse Kidney Using Magnetic Activated Cell Sorting (MACS).

Abstract

Glomerular diseases characterized by injury to post-mitotic epithelial cells called podocytes are a leading cause of chronic kidney disease. Yet, isolating podocytes from the kidney for transcriptomic, proteomic, and metabolomic studies has been a major technical challenge. Protocols utilizing glomerular sieving and laser capture methods are of limited use because they are not podocyte-specific but instead capture all four glomerular cell types. Here, we present a magnetic-activated cell sorting (MACS) method where podocytes are isolated from digested whole kidneys using antibodies specific to extracellular antigens on podocytes. Using microbeaded secondary antibodies binding to the podocyte-specific primary antibodies allows sorting of the podocytes using a magnet. This podocyte-only cell fraction is a unique source of in vivo-derived cells for molecular and cellular experiments. Key features • The protocol isolates a podocyte-only cell population from kidneys that is readily available for molecular and cellular studies. • The non-podocyte fraction serves as a matching negative control. • High cell yields are obtained. • The method can be applied to separately isolate podocytes from the outer cortex and juxtamedullary regions of the kidney.