title: GSM8653197: Murine Splenic NK cells; sample 2; Mus musculus; RNA-Seq
library_construction_protocol: All animal handling procedures were in accordance with National Institutes of Health guidelines and were approved by the Institutional Animal Care and Use Committees of New York University School of Medicine. Isolated splenocytes were washed 2x with media and passed again through 70 µm cell strainer. In other experiments, spleens were dissociated with the gentleMACS spleen disassociation kit (Miltenyi Biotec). Cells were then filtered, erythrocytes were depleted via isotonic hemolysis, and NK cells were isolated using a negative selection method (NK Cell Isolation Kit, Miltenyi Biotec) following the manufacture's guidelines. In this negative selection method, non-NK cells, i.e. T cells, dendritic cells, B cells, granulocytes, macrophages, and erythroid cells are magnetically labelled with a cocktail of biotin-conjugated antibodies and anti-biotin MicroBeads, and then removed from isolated splenic cells using magnetic columns. We used the Chromium Next GEM Single Cell 3ʹ Reagent Kit v3.1 (10x Genomics) to perform gel beads-in-emulsion (GEM) generation and barcoding, Post GEM-reverse transcriptase (RT) cleanup and cDNA amplification, and 3' gene expression library construction.