title: GSM8072837: OA1-1_Condition_NWB_Rep1; Homo sapiens; RNA-Seq
library_construction_protocol: Cartilage was harvested within an hour after surgery, washed twice with phosphate-buffered saline (PBS) buffer containing penicillin and streptomycin, and cut into 1 mm3 pieces. Samples were then subjected to pretreatment with 0.25% Trypsin (Thermo Fisher Scientific, Waltham, MA, USA) at 37°C in an incubator with 5% CO2 for 30 minutes. The cartilage fragment was then centrifuged at 350×g for 5 minutes and the supernatant was discarded completely, followed by digestion in basal Dulbecco's modified Eagle's medium/F12 media (Gibico, Grand Island, NY, USA) supplemented with 0.2% type II collagenase (Gibico, Grand Island, NY, USA) at 37 °C for 10-12 hours. Cell count and viability were estimated using a fluorescence Cell Analyzer (Countstar® Rigel S2) with AO/PI reagent after the removal of erythrocytes (Miltenyi 130-094-183). Debris and dead cells removal was performed according to the recommended manuals (Miltenyi 130-109-398/130-090-101). For the 10X Genomics protocol, isolated chondrocytes were filtered through a 70 μm cell strainer, washed twice with PBS buffer, and directly subjected to cDNA library construction. Articular cartilage tissue was dissociated into single cells by enzymatic methods according to standard procedures. Briefly, the cells were washed with PBS and resuspended in 500 μL PBS, and single-cell RNA-seq libraries were prepared using the Chromium Single Cell 3' Library v3 Reagent Kit (10x Genomics, Pleasanton, CA, USA). Single-cell RNA-seq libraries were prepared using the Chromium Single Cell 3' Library v3 Reagent Kit (10x Genomics, Pleasanton, CA, USA). Sequencing was performed on an by Illumina NovaSeq 6000.