Blood vessels are critical for delivering oxygen and nutrients to all tissues in the body. This is especially important in the central nervous system, which is extremely sensitive to hypoxia and ischemia. Blood vessels are made of two main cell types: endothelial cells and mural cells. Endothelial cells form the walls of the blood vessels that generate a lumen through which blood flows. Mural cells are support cells thought to be involved in vessel contractility, vascular remodeling, and regulation of endothelial permeability. On large vessels, including arteries and veins, mural cells are termed vascular smooth muscle cells. On the small vessels of the capillary bed, they are called pericytes. Here, we provide a brief introduction to the methods for purification of endothelial cells, including an immunopanning method that we developed for isolating these cells from the rodent brain and optic nerve.