GSM7885773: scSeq_CAR5_Seq2; Homo sapiens; RNA-Seq
Human carotid arteries were harvested during carotid endarterectomy or open repair in our Department of Vascular and Endovascular Surgery. The tissue was transported in PBS on ice to our laboratory. The two distinct pieces of each tissues (except CAR1) were minced and digested using the Multi Tissue Dissociation Kit 2 (Miltenyi Biotech, 130-110-203), GentleMACS Dissociator (Miltenyi Biotech, 130-093-235), GentleMACS C tubes (Miltenyi Biotech, 130-096-334) and the 37C_Mulit_G program, all according to the manufacturer´s instructions. The cell suspension was strained (70 µm, 40 µm) and Dead Cell Removal (Miltenyi Biotech, 130-090-101) using MS Columns (Miltenyi Biotech, 130-042-201) was performed. Cells were resuspended in PBS + 0,04% BSA. Cells were loaded into a 10x Genomics microfluidics Chip G and encapsulated with barcoded oligo-dT-containing gel beads using the 10x Genomics Chromium Controller. Gel Beads-in-emulsion (GEM) cleanup, cDNA Amplification and 3´Gene Expression Dual Index Library Construction was performed according to the manufacturer´s instructions (CG000315 Rev C). Libraries from individual samples were multiplexed into one lane before sequencing on an Illumina NovaSeq6000 instrument Chromium next GEM Single Cell 3´ Reagent Kit v2 (CAR 1), v3.1 (CAR 2, CAR 3, CAR 4), v3.1 Dual Index (CAR 5, CAR 6, CAR 7, CAR 8, CAR 9, CAR 10)