GSM7017458: Spatial CLAD sample 2; Homo sapiens; RNA-Seq
A 2cm x 2cm segment of lung tissue was isolated at least 1cm from the pleura and minced using a razor blade. Tissue was digested at 37C using Miltenyi Multi Tissue Dissociation Kit 1 (Miltenyi Biotech, 130-110-201) for 30 minutes. Mechanical digestion was performed three times using Miltenyi gentleMACS dissociator (Miltenyi, 130-093-235). The cell suspension was spun down at 300g for 10 minutes, and cells were treated with RBC lysis buffer (Sigma, R7757). The cell suspension was filtered using a 70μm filter, spun down at 300g for 10 minutes, and resuspended in PBS with 0.04% BSA. For spatial transcriptomics, 10µm sections were cut from frozen OCT blocks and mounted onto Visium spatial gene expression slides and processed according to the manufacturer's protocol (10x Genomics). Tissue sections were permeabilized immediately to capture RNA molecules onto barcoded spots on the slide. Single cells were prepared for cDNA amplification and library preparation using Single Cell 3' Reagent Kits v2 (10x Genomics) following the manufacturer's protocol