PD1_Mut
This is single-cell RNA sequencing data. The frozen PBMCs were rapidly thawed at 37C and gently resuspended by serial additions of DMEM containing 10% FBS to a final volume of 14 mL. Cells were centrifuged at 300 x g for 5 minutes and were washed twice with 5 mL of medium to remove debris. Viability was assessed with the LIVE/DEAD Aqua viability kit. The preparation was enriched in viable cells by resuspending 1106 cells in 200 L of 2% FBS and 1 mM calcium chloride in PBS and passing the suspension through a strainer with 40 m pores, before removing dead cells with the EasySep Dead Cell Removal Kit. Cells were then centrifuged at 300 x g for 5 minutes and resuspended at a concentration of 1103 cells/L in 0.04% BSA in PBS. The initial viability of ~50% was increased to 79% and 75% for the samples of the patient and his brother, respectively. The samples were loaded onto a 10X Genomics Chromium chip for single-cell capture. Libraries were prepared with the Chromium Single Cell 3 Reagent Kit. Library quality was assessed with a Bioanalyzer DNA chip. The libraries were sequenced on by Illumina NovaSeq 6000 sequencer.