Rat erythrocyte plasma membranes have been extracted exhaustively with digitonin at low temperature, and the residual, detergent-extracted membrane cytoskeletal material is compared to that prepared with Triton X-100 with respect to protein, glycoprotein, phospholipid, and cholesterol content. Digitonin, a weaker detergent than Triton X-100, solubilizes only 26% of the phospholipids and none of the cholesterol. SDS-polyacrylamide gel electrophoresis reveals that differences between the proteins extracted by the two detergents are primarily quantitative. In terms of functional preservation, digitonin retains in the cytoskeleton 28% of the β-adrenergic receptor binding activity (with the balance accounted for in the supernatant),>90% of the adenylate cyclase and>90% of the 45,000 mol wt polypeptide cholera toxin substrate. The cytoskeletal-associated β-adrenergic receptor retains binding properties for antagonist and agonist which are identical to those of the native membrane receptor. The digitonin-extracted cytoskeleton containing the β-adrenergic receptor may provide a useful vehicle for the reconstitution of a hormone-sensitive adenylate cycla