Autogenous endothelial seeding (AES) of vascular prostheses (VP) using venous endothelial cells (EC) reduces platelet-VP interactions and improves patency rates in small caliber VP in dogs. To conserve patients' veins for use in coronary or limb bypass surgery, human trials of AES should require proof that adequate numbers of EC with the growth capacity to cover VP can be harvested from acceptably small pieces of peripheral vein. EC were isolated from excess saphenous vein segments remaining after coronary bypass surgery by filling veins with 0.1% CLS II collagenase at 37°C for 15 min and removing EC by flushing the veins with culture medium. EC were cultured on fibronectin-coated dishes in medium 199 with 30% human serum and 300 μg/ml of endothelial cell growth factor. These cells grew to form confluent monolayers, and were identified as EC by tests for factor VIII antigen. Veins from 53 patients with a mean age of 55.8 ± 9.8 (SD) years yielded vein segments with an average area of 1.9 ± 0.6 cm2, from which an average of 5.3 ± 2.8 × 104 cells were removed per cm2 of vein area. EC in culture underwent 14.3 ± 1.4 population doublings with an average population doubling time of 1.8 ± 0.3 days (N = 14 cultures), which allowed an 100-fold increase in cell number to occur in 11 to 12 days. These data suggest that the EC available from small vein segments in adult humans have the growth capacity to cover areas comparable in size to the luminal areas of VP commonly used in arterial surgery.