title: GSM5176928: Sarcoidosis patient scRNA-seq 5; Homo sapiens; RNA-Seq
library_construction_protocol: 4 mm punch biopsies of skin were obtained on separate days from 3 patients with GA and 3 healthy volunteers.. Biopsies were immediately bisected and placed in dispase (10 mg/mL) (Dispase II Sigma Cat# D4693) in RPMI with 2% FBS for 45 minutes at 37°C with shaking at 125 RPM. The tissue was removed from the dispase solution, minced in a sterile plate with a scalpel, and then subsequently incubated with liberase (0.5 mg/mL) (Liberase TH Sigma Cat# 5401135001) in RPMI with 2% FBS for 45 minutes at 37°C with shaking at 125 RPM. The dispase solution was placed on ice. After incubation in the liberase, the dispase solution was combined (to recover any additional cells from the dispase) and the mixture was triturated and filtered through a 100 mM sterile filter. Cells were washed with RPMI + 2% FBS and RBCs were lysed with ACK Lysing Buffer (Lonza Cat# 10-548E) and washed again in RPMI + 2% FBS. Cells were stained with LIVE/DEAD Red viability dye (ThermoFisher Cat# L23102), FC block (Biolegend Cat#422301) and CD45 Alexa Fluor 488 (Biolegend Cat# 304017) in RPMI + 0.1% FBS. Viable cells were isolated using a BD FACS Aria cell sorter and sorted into cold RPMI + 2% FBS. CD45 positivity was recorded but not used as a criterion for sorting. Up to 10,000 cells (depending on yield) were loaded onto a 10X chromium controller for each sample according to the manufacturer's instructions. Single cell libraries were created according to the manufacturer's instructions by the Yale Center for Genome Analysis (YCGA).
attributes.tag: GEO Accession
attributes.value: GSM5176928