We have used ruthenium red, a cationic dye, to detect at the electron microscopic level the presence of anionic sites in various murine basement membranes, with particular emphasis on those of the microvasculature. We have observed anionic sites in all continuous and fenestrated capillaries examined. Terminal lymphatics, which have a discontinuous basement membrane, have sites only where the basement membrane is present. Anionic sites are not present beneath sinusoidal lining cells of the liver which lack a basement membrane. Basement membranes of epithelial cells and those surrounding striated and smooth muscle cells, pericytes, fat cells, and Schwann cells also exhibit anionic sites. We compared the electrostatic properties of anionic sites in basement membranes of continuous and fenestrated capillaries by determining the salt concentration (critical electrolyte concentration, Scott and Dorling, 1965) required to displace ruthenium red from the sites. A concentration of 0.5 M Na+ was required to displace ruthenium red from the basement membrane of continuous capillaries of muscle whereas 1.3 M Na+ was required to displace ruthenium red from the basement membrane of fenestrated peritubular capillaries of the renal cortex. Our results suggest that anionic sites in the basement membrane of fenestrated peritubular capillaries are more strongly negatively charged than those in the basement membrane of continuous capillaries of muscle. We conclude from this study, first, that anionic sites are a general property of vascular, epithelial, and pericellular basement membranes and, second, that the electrostatic properties of the sites differ in different vascular basement membranes. We speculate that the anionic sites in vascular basement membranes and the variation in their electrostatic properties in different types of capillaries may have important implications for exchange of substances across the capillary wall.