A method for the preparation of cardiac sarcoplasmic reticulum (SR) from rat heart is described. SR isolated from control and diabetic rats was characterized to determine if differences in calcium transport activity could originate as an artifact of preparation. Electron micrographs of cardiac SR preparations isolated from control and diabetic rats were indistinguishable. The yeild of SR and the mitochondrial membrane contamination were similar in both preparations. These results suggest that depression in diabetic rat cardiac SR microsomal function is not due to nonspecific alterations in the membrane preparations used.