GSM6422823: Patient2 (P2); Homo sapiens; RNA-Seq
Carotid tissue was extensively washed in RPMI, finely minced and tissue fragments were incubated in collagenase type I, elastase type III , and DNase, with trypsin inhibitor, polymixin B , andCaCl2, in RPMI medium 1640 with 5% FBS in a shaker at 37°C for 45 min. Cell suspension was filtered through an 80-μm Nylon mesh and washed in media. Collected cells were incubated with live/dead stain at 1:1,000 in PBS for 10 min at 4 °C. Cells were then washed with FACS buffer (PBS with 2% FBS), stained for CD45 antibody for 30 min at RT then washed twice with FACS buffer, resuspended in a final FACS buffer volume of 500 μl, filtered through an Easy Strain 100-μm cell strainer and sorted for live CD45+ cells using FACS ARIAIII in FBS.