A sample of whole blood, anticoagulated with EDTA, from each patient was used for flow cytometry analysis. One hundred microliters of whole blood was incubated with a rabbit anti-human RAGE antibody (Chemicon, Billerica, MA) for 15 min at room temperature. A Goat anti-Rabbit IgG FITC conjugate (Chemicon) was used for flow cytometry detection. Samples were incubated for 15 min at room temperature in darkness. Lysis solution was then added to eliminate erythrocytes and two washes with PBS (0.1 M, pH 7.2) were done centrifuging at 220-240 × g for10 min in each time. Leukocytes were recovered by centrifugation in the same condition and the samples were resuspended in 1 ml of FACS flow (BD Biosciences, Pharmingen) for cytofluorometric analysis (FACS SortCalibur, BD, San Jose, CA). Then 10, 000 cells, in which mean fluorescence intensity (MFI) was obtained and nonspecific fluorescence was deleted, were analyzed.