Fitc mouse anti human cd62p

SARS-CoV-2 (2019-nCoV) spike protein was purchased from Sino Biological. Coenzyme Q10 (CoQ10), 2′,7′-Dichlorofluorescin diacetate (DCFH-DA), ADP, thrombin from human plasma, and N-acetylcysteine (NAC) were obtained from sigma-Aldrich (St. Louis, MO, USA). Collagen was obtained from Chrono-Log Corp. (Havertown, PA, USA). FITC mouse anti-human CD62P and FITC Mouse Anti-Human PAC-1 were purchased from BD bioscience (San Jose, CA, USA). Alexa Fluor^TM 488-conjugated fibrinogen was purchased from Thermo Fisher Scientific (Waltham, MA, USA). TMRM assay kit (mitochondrial membrane potential) was purchased from Abcam (Cambridge, UK). Prostaglandin E1 (PGE1) was obtained from MedChemExpress (Monmouth Junction, NJ, USA). MDA, SOD, and T-AOC assay kits were purchased from Nanjing Jiancheng (Nanjing, China) and the 8-iso-PGF-2α assay kit was obtained from Andy gene (Beijing, China).

Immediately after isolation, 2 µl mouse platelets were incubated with antibodies against IgG1-FITC (Milteny Biotec, Bergisch Gladbach, Germany, order no.: 130-098-847, 1:25) or CD62P-FITC (BD Biosciences, Heidelberg, Germany, Cat: 561923, 1:25). Human platelets were incubated with antibodies against FITC Mouse IgG1 (BD Biosciences, Cat: 555748, 1:25) or FITC Mouse Anti-Human CD62P (BD Biosciences, Cat: 555523, 1:25). Subsequently, 46 µl of FACs buffer (2 mM EDTA, 0.5 % FCS ad 100 ml PBS, pH 7.1) was added and the samples incubated for 30 min at 4 °C in dark. To study apoptosis, 2 µl of platelets were incubated with 5 µl Annexin V-FITC (Milteny Biotec, Order no.: 130-097-928) in 100 µl Annexin V Binding Buffer (Miltenyi Biotec, 20x Stock Solution) at RT for 30 min in dark. Immediately before analysis, 2 µl of 7-AAD (Miltenyi Biotec) was added. All samples were centrifuged at 300g for 10 min and the pellets were resuspended in 100 µl of FACs flow (BD FACSFlow). FACs analysis was instantly performed with a BD FACScan.