title: GSM5172569: liver, S2; Homo sapiens; RNA-Seq
library_construction_protocol: 10X Genomics Single cell 3' gene expression V2 Tissue was placed in PBS on ice and transported from the hospital directly to the laboratory for immediate tissue dissociation. To enable paired histological assessment, a section of the sample was fixed in 10% Neutral Buffered Formalin (VWR) for 24 hours and then placed into 70% ethanol for long term storage before paraffin embedding and staining. For human liver scRNA-seq, liver tissue was mechanically dissociated with scissors and placed into a digestion buffer of 15mL PBS +/+ (Invitrogen), DNAse I (250mg/l, Akron Biotech), and Collagenase (500 mg/l, Worthington Biochemical) at 37 °C for 20 minutes with one agitation halfway through digestion. Samples were strained through a 250um metal strainer. Cells were centrifuged (450 x g for 5 minutes; cell were washed twice with 10mL PBS -/- (Invitrogen).  Red blood cell lysis was performed (Gibco ACK Lysing Buffer), followed by centrifugation (450 x g, 5 minutes), and resuspension in stromal growth media (ALPHA MEM  + L-Glutamine, -Deoxyribo, -Ribonucleodite (Life Technologies, Cat #12561072), 30% heat inactivated fetal bovine serum (Sigma), 1 % penicillin (100 U ),streptomycin (100ug/mL) (Invitrogen). Cells were passed through 70um filter. Single cell suspensions were used for primary cell line development and scRNA-seq analysis.