Background. Injury to endothelium can compromise the patency of bypass grafts harvested during coronary artery bypass graft (CABG) surgery. Maintaining structural and functional viability of endothelium in grafts may lead to improved long-term patency. The information gained from the application of multi-photon microscopy in transmission and epifluorescence mode was used to assess the structural and functional integrity of human saphenous vein segments stored in multiple preservation solutions, and to design a superior storage solution. Methods. Multi-photon microscopy was used to image deep within saphenous vein tissue harvested from patients undergoing CABG for analysis of endothelial structure and function. Endothelial cell structural viability, calcium mobilization, and nitric oxide generation were determined using specific fluorescence markers. Results. Within 60 minutes of harvest and storage in standard preservation solutions, calcium mobilization and nitric oxide generation were markedly diminished with more than 90% of endothelial cells no longer viable in the vein. In contrast, veins could be stored for 24 hours without substantial loss in cell viability in a newly formulated heparinized physiologic buffered salt solution containing glutathione, ascorbic acid, and L-arginine (GALA). Conclusions. Standard solutions in clinical use today led to a profound decline in saphenous vein endothelial cell viability, whereas the newly designed physiologic salt solution (GALA) maintained endothelial function and structural viability for up to 24 hours. The improvements seen from using GALA as a vessel storage medium may lead to greater long-term vein graft patency following CABG surgery.