Abstract: [Abstract] Objective To establish a flow-cytometric method for detection of microvesicles (MVs) in rat peripheral blood, detect platelet-derived MVs (PMVs) and endothelial cell-derived MVs (EMVs) in blood from ischemic preconditioning (IPC) treated rats. Methods Blood was withdrawn from rat abdominal aorta and anticoagulated with sodium citrate. Platelet-free plasma (PFP) was obtained after two centrifugations at room temperature. PFP was incubated with FITC-conjugated mouse anti-rat CD61 or PE-conjugated mouse anti-rat CD144. Standard beads in diameter 1 and 2 μm were used for size calibration and absolute counting, respectively. Analysis was performed on flow cytometer. Results When 3.5% sodium citrate was blent with blood according to the volume ratio of 1:4, clear supernatant was obtained after centrifugation. Signals of particles smaller than 1 μm accounted for more than 99% of all. PMVs and EMVs were CD61 positive and CD144 positive, respectively. Their diameters were smaller than 1 μm. The concentration of PMVs and EMVs in peripheral blood from IPC treated rats was 4053±1987/μL and 4870±825/μL, respectively. Conclusion The method for MVs detection by flow cytometry was successfully established and optimized, and verified through detecting PMVs and EMVs in peripheral blood from IPC treated rats.

Key words: blood platelets, endothelial cells, flow cytometry, rats, Wistar, microvesicels, ischemic preconditioning