This study tests the hypothesis that denucleating vascular graft material and binding cell adhesion molecules increases endothelial attachment. Removal of gas nuclei (denucleation) increases the available surface area of biomaterials for modification and/or cell adhesion, while adhesion molecules provide specific attachment sites. Microvascular endothelial cells (MVEC) were isolated from fat, fluorescently labeled, and allowed to settle onto expanded polytetrafluoroethylene (ePTFE) vascular patches. Patch treatments included fibronectin alone (F), gas denucleation followed by fibronectin (D/F), denucleation followed by the surfactant tridodecylmethylammonium chloride (TDMAC) (D/T), denucleation followed by TDMAC followed by fibronectin (D/T/F), or denucleation followed by TDMAC followed by a synthetic polymer with numerous arginine-glycine-aspartic acid sequences (D/T/R). After 1 h of incubation, the 45 mm2 patch area covered with microvascular endothelial cells was assessed using computer-aided fluorescence microscopy. Initial graft coverage with D/T (26.2 +/- 2.4 mm2) and D/T/F (25.9 +/- 2.1 mm2) was better than with F (16.8 +/- 2.5 mm2) (P < .05). Patches were then exposed to a detachment stress and coverage was again measured. Following stress, coverage was greater with D/T (20.7 +/- 3.4 mm2) and D/T/F (20.7 +/- 2.0 mm2) than with D/T/R (8.4 +/- 1.8 mm2) or F (3.6 +/- 0.9 mm2) (P < .001). Percent retention of cells following stress was better with D/T and D/T/F than with D/T/R, D/F, or F (P < .0001). Scanning electron micrographs were consistent with the qualitative findings. The results indicate that TDMAC alone or with fibronectin increases adhesion of human microvascular endothelial cells to denucleated ePTFE.