GSM9597726: Visium standard 11-0418 FFPE specimen; Homo sapiens; OTHER
Frozen tissue samples were obtained from brain resections. Tissue was thawed and minced on ice prior to transfer to 1mL of lysis buffer (10µL 1M Tris-HCl, pH 7.5, 1µL 5M NaCl, 3µL 1M MgCl2, 10µL 10% NP-40, 1µL RNasin Plus (Promega N2615) in sterile water). The sample was incubated on ice for 5 minutes and then triturated 10-15 times every 30 seconds for 10 minutes. The sample was strained through a 70µm filter and centrifuged at 300g for 5 minutes at 4oC. The cell pellet was resuspended into 1.5mL Optiprep solution (Sigma D1556) and centrifuged at 10,000g for 10 minutes at 4oC. The cell pellet was resuspended in 1mL resuspension buffer (1X PBS, 1% BSA, RNase Plus inhibitor (Promega N2615)) and filtered using a 40µm filter. Finally, the suspension was centrifuged again at 10,000g for 10 minutes at 4oC and resuspended in resuspension buffer. Single nuclei were prepared for cDNA amplification and library preparation using Single Cell 3' Reagent Kits v2 (10x Genomics)