GSE183456
Spatial localization with Spatial Transcriptomics for an atlas of healthy and injured cell states and niches in the human kidney [Visium ST]
To spatially localize cell types within the tissue, snCv3 subclasses were used to predict the corresponding identities in Visium transcriptomic data. This allowed for recapitulated renal corpuscle, tubular, vascular, and interstitial cell types having proportions, marker profiles, and spatial organizations consistent observed histopathology. Overall design: Human kidney tissue was prepared and imaged according to Visium Spatial Gene Expression protocols (10x Genomics) according to the manufacturer protocol (CG000240 protocol, Visium Tissue Preparation Guide) and as previously described 89. Tissue was sectioned at 10 µm thickness from Optimal Cutting Temperature (OCT) compound embedded blocks. A Keyence BZ-X810 microscope equipped with a Nikon 10X CFI Plan Fluor objective was used to acquire hematoxylin and eosin (H&E) stained brightfield mosaics which were subsequently stitched. mRNA was isolated, libraries prepared, and sequencing was performed on an Illumina NovaSeq 6000. mRNA was isolated from stained tissue sections after permeabilization for 12 minutes. Released mRNA was bound to oligonucleotides in the fiducial capture areas. mRNA was then reverse transcribed and underwent second strand synthesis, denaturation, cDNA amplification, and SPRIselect cDNA cleanup (Visium CG000239 protocol). Space Ranger (v1.0.0) with the reference genome GRCh38 3.0.0 was used to perform expression analysis, mapping, counting, and clustering. Normalization was performed with SCTransform. Final data processing was done in Seurat (v3.2.3).  As of the design of this study, QC metrics were not established by the lab or the manufacturer. Currently, three CKD biopsies would fall below the established threshold of fraction of reads mapped to exons above 0.3. These samples were not included in the main analysis of the paper.