Cholestyramine, a well-known bile-salt sequestrant, can be used effectively to remove cholate or deoxycholate from a solution of phosphatidylcholine-bile salt mixed micelle. Upon removal of the bile salt, unilamellar phospholipid vesicles form essentially instantaneously. Cholestyramine resin could be pelleted and removed from the vesicle solution after a low speed centrifugation. Based on phosphate analyses, the recovery of vesicles was approximately 60% of the starting material. The average diameter of these vesicles, as estimated by gel exclusion chromatography on sephacryl S-1000 beads and by trapped volume measurement using [3H]sucrose, ranged between 85 to 121 nm. Phosphatidylethanolamine, cholesterol, or n-alkane such as tetradecane can be incorporated into the vesicles without any selective loss; however, selective loss was experienced when negatively charged phospholipid species such as phosphatidylglycerol or phosphatidylserine was included in vesicle formation.