GSM7589541: Brain slice, naïve, 1; Mus musculus; RNA-Seq
At 12 weeks post-immunization, animals were euthanized in CO2 chamber and perfused with cold PBS. Brains were dissected and one hemisphere drop fixed in isopentane cooled on dry ice. Fresh frozen brain samples were then cut coronally at a thickness of 10μm and placed on the capture area of Visium Gene Expression slides (v1; 10x Genomics). Each slide contained four 6.5 mm x 6.5 mm capture areas. Sample preparation was carried out according to manufacturer's instructions. After fixation with methanol at -20C, Hematoxylin and Eosin (H&E) staining was performed for morphological analysis and spatial alignment of the sequencing data. After the enzymatic permeabilization, mRNA was captured by probes and cDNA generated. Barcoded cDNA was isolated using SPRIselect-cleanup (Beckman Coulter) and amplified. Amplified cDNA was fragmented and subjected to end-repair, poly-A-tailing, adaptor ligation, and 10x-specific sample indexing as per the manufacturer's instructions. Following assessment of RNA quality, sequencing was performed using a Novaseq S2 100.