Anti cd4 apc ef780

Mouse peripheral blood was obtained by retro-orbital venipuncture, collected in sodium citrate solution, red blood cells lysed and the remaining cells were washed in staining buffer (2% BSA, 0.02% azide in PBS (PAB)) as previously described (Bustamante et al 2008, 2014, 2020). Whole blood was incubated with a major histocompatibility complex I (MHC I) tetramer containing the specific TSKB20 peptide (ANYKFTLV/Kb) labeled with BV421 (Tetramer Core Facility at Emory University, Atlanta, GA) and the following labeled antibodies: anti-CD8 FITC, anti-CD4 APC EF780, anti-CD127 PE (BD Bioscience, San Jose, CA). For whole blood, we lysed red blood cells in a hypotonic ammonium chloride solution after washing twice in PAB. We stained cells for 45 min at 4°C in the dark, washed them twice in PAB and fixed them in 2% formaldehyde. At least 500,000 cells were acquired using a CyAn ADP flow cytometer (Beckman Coulter, Hialeah, Florida) and analyzed with FlowJo software v10.6.1 (Treestar, Inc., Ashland, OR).