Aim: Saphenous vein (SV) is the most commonly used conduit in bypass procedures but has a one-year occlusion rate of 15-30%. A new ‘no-touch’ technique where the SV is harvested with a cushion of surrounding tissue with no distension has led to improved early patency rates of 5% at 18-months. Nitric oxide (NO), synthesised by nitric oxide synthase (NOS) has properties beneficial to graft patency. Our aim was to study the distribution of NOS in SV harvested by this technique and the effect of distension and removal of perivascular tissue on NOS content of SV. Methods: Following ethical committee approval and patients' informed consent, SVs were harvested from ten patients undergoing coronary artery bypass grafting. A segment of vein was harvested by the conventional technique (surrounding tissue stripped and vein distended with saline); another part was stripped but not distended (‘control’) and the remaining parts harvested by the ‘no-touch’ technique. Samples of each segment were taken and transverse sections prepared for NOS identification using 3[H]L-NG nitroarginine (NO Arg) autoradiography and NADPH-diaphorase histochemistry. NOS isoforms were studied using standard immunohistochemistry. Endothelial cells and nerves were also identified using immunohistochemistry with CD31 and NF200 respecitvely, to confirm sources of NOS. Morphometric analysis of NADPH-diaphorase staining was carried out to study tissue NOS content. Results: NO Arg binding representing NOS was preserved on the lumen of ‘no-touch’ vessels whilst that on conventional and control vessels was reduced. NOS was also localised to the medial smooth muscle cells of all vein segments and to the intact adventitia of ‘no-touch’ segments. This was confirmed by NADPH-diaphorase staining, which revealed a mean reduction of NOS by 19.5% (p < 0.05, ANOVA) in control segments due to stripping of surrounding tissue alone and a reduction of 35.5% (p < 0.01, AVNOVA) in conventional segments due to stripping and distension, compared to ‘no-touch’ segments. Adventitial NOS sources in ‘no-touch’ vessels corresponded to vasa vasorum and paravascular nerves. All three NOS isoforms contributed to the preserved NOS in ‘no-touch’ vessels. Conclusions: Apart from preserved lumenal NOS, NOS sources are also located in the media and adventitia of SV grafts. These are reduced by both adventitial damage and vein distension during conventional vein harvesting. The ‘no-touch’ technique avoids these procedures, preserving NOS sources. This may result in improved NO availability in SV harvested by this technique, contributing to the improved patency rates reported.