title: GSM8767156: MDA_with_inhibitor; Homo sapiens; RNA-Seq
library_name: GSM8767156
library_construction_protocol: Fresh tissue samples were directly placed in MACS tissue storage buffer (Miltenyi Biotec, Germany) on ice before transportation to the processing laboratory. Given the potential need to transport tissue from different institutions we also tested the performance of tissue storage buffer (results in Supplementary Table 4). Once received in the laboratory, tissue cores were morselized into small pieces under sterile conditions using sterile scalpels, triturated using pipettes then transferred to a 50ml falcon tube (Corning, USA), then incubated with papain (37°C, 25 min)(Worthington, USA) according to manufacturer instructions. After incubation HBSS without magnesium or calcium was then added to make up to 50ml and the sample centrifuged (300×g, 5 min, 4◦C), supernatant aspirated before being passed through a 70μm cell strainer (BD, USA). Erythrocytes were removed by resuspending and incubating the obtained pellet in 5 ml of ACK-lysis buffer (eBioscience, USA) for 10 min, followed by centrifugation (300×g, 5 min,4°C). Residual myelin and extracellular debris were eliminated using the Debris Removal Kit (Miltenyi Biotech, Germany). Cell counts were quantified using an automated cell counter (Nexcelom, USA) after resuspending the pellet in PBS. Cells were stored in Bambanker (Wako, Japan). Cell suspensions were immediately placed in a storage container (Mr. FrostyTM, Thermo Fisher Scientific, USA) and stored in LN2. Freshly isolated cells were droplet-separated using the Chromium Single Cell 3′ v.3 Reagent Kit with the 10x Genomics microfluidic system creating cDNA library with individual barcodes for individual cells. scRNA-seq was performed using the 10x Genomics Chromium Single Cell Controller
identifiers.namespace: GEO
identifiers.id: GSM8767156_r1