Fitc conjugated mouse anti human cd62p

Lysed WB and PRP samples were resuspended in 150 μL Tyrode’s buffer. Samples were labelled with APC-conjugated mouse anti-human CD41a (BD Pharmingen; 559777), FITC-conjugated mouse anti-human CD62P (BD Pharmingen; 555523) and PE-Cy7 mouse anti-human CD63 (BD Pharmingen; 353010) at a concentration of 1:20 as determined using titration. Following compensation, experiments were conducted on an LSR Fortessa Flow Cytometer (BD Biosciences, South Africa). Events were recorded at 100,000 events per sample using FACSDiva Software (version 6.2) (BD Biosciences)⁸⁶ . The platelet population was initially gated using a forward scatter (FSC) voltage of 300 V and a side scatter (SSC) voltage of 275 V followed by gating a singlet population using FSC-height and FSC-area. The parent platelet population was identified and further gated based on expression of the CD41a (APC, 565 V). CD62P (FITC, 469 V) and CD63 (PE-CY7, 645 V). Interval gates^{30 (link),50} were drawn to classify a graded level of each platelet activation marker determined using Geometric Mean Fluorescence Intensity (gMFI) of the platelet population (Fig. 2). Data acquired was exported into Microsoft Excel.