It is generally believed that sphingomyelin- and cholesterol-enriched microdomains can be isolated as detergent-resistant membranes (DRMs) from plasma membrane and organelle membranes. Here the isolation of microdomains from porcine cardiac sarcoplasmic reticulum (SR) membranes is described. These SR-derived detergent-resistant membranes (SR-DRMs) enriched in sphingomyelin and cholesterol have a low buoyant density. Both ganglioside GM1 and caveolin-3 in the SR, known as a marker protein of caveolae at plasma membrane are present in the SR-DRMs. It was demonstrated that significant amount of SERCAs together with caveolin-3 associates with SR-DRMs, and are fully functional. The results suggested that SR membranes like other biomembranes, such as the plasma membrane, contained microdomains.