Collagen IV is a major component of vertebrate basal laminae (BLs). Studies in humans have revealed a family of genes encoding α1-α6 collagen IV chains and implicated α3-α6 in disease processes (Goodpasture and Alport syndromes and diffuse leiomyomatosis). To extend studies of these components to an experimentally accessible animal, we cloned cDNAs encoding partial collagen α3, α4, and α5(IV) chains from the mouse. Ribonuclease protection assays showed that all three genes were expressed at highest levels in kidney and lung; α5(IV) was also expressed at high levels in heart. We then made antibodies specific for each collagen IV chain. Immunohistochemical studies of several tissues revealed many combinations of collagen IV chains; however, α3 and α4 (IV) were always coexpressed, and only appeared in BLs that were α5(IV) positive. The α3-α5(IV) chains were frequently but not exclusively associated with the S (β2) chain of laminin, as were the α1, 2 (IV) collagen chains with laminin B1 (β1). An analysis of developing rat kidney BLs showed that newly formed (S-shaped) nephrons harbored collagen α1 and α2(IV) and laminin B1; maturing (capillary loop stage) BLs contained collagen α1-α5(IV) and laminin B1 and S-laminin; and mature glomerular BLs contained mainly collagen α3-α5(IV) and S-laminin. Thus, collagen α1 and α2(IV) and laminin B1 appear to be fetal components of the glomerular BL, and there is a developmental switch to collagen α3-α5(IV) and S-laminin expression.