Centrifugation of citrated whole blood (cit-WB) for platelet rich plasma (PRP) preparation resulted in a significantly diminished platelet yield compared to when blood was collected in citrate + aspirin (acetylsalicylic acid; ASA) or EDTA anticoagulants in both apparently healthy young subjects and peripheral vascular disease (PVD) patients. The loss in platelets was significantly greater in PVD patients than in young apparently healthy subjects when using cit-WB. Median platelet volume (MePV) of PRP preparations was not different between PVD patients and apparently healthy subjects. The MePV of EDTA-PRP preparations was significantly bigger than cit-PRP and cit + ASA-PRP in both PVD patients and apparently healthy subjects. Citrate and citrate + ASA-PRP of patients was significantly more aggregable to adrenaline. In EDTA-PRP, adrenaline-induced aggregation did not occur. These findings indicate that platelet hyperactivity occurs in PVD even in the presence of ASA. Furthermore, studies involving the use of PRP may not be representative in PVD due to the significant loss of platelets during centrifugation. The methodology described here can be used by those who do not have facilities designed specifically for platelet research.