Technologies

Transcriptomics

Single-nucleus, single-cell, regional, and spatial transcriptomics: goals, protocols, and metadata standards.

KPMP Transcriptomics technologies

Single-nucleus RNA-seq (snRNA-seq)

Goal: Identify cell types and states associated with normal and injured kidney functions using gene expression profiling. Identify marker genes for cell type/states and any proportion shifts underlying pathology.

Single-nucleus RNA-seq workflow: a kidney biopsy from AKI, CKD or reference tissue is homogenized into single nuclei, run on 10X Chromium, and clustered into epithelial, endothelial, immune and stromal cell types.

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Single-cell RNA-seq (scRNA-seq)

Goal: Empirically derive cell subtypes and cell-type-specific gene expression profiles​.

Single-cell RNA-seq workflow from sample collection, cryopreservation, thawing and tissue dissociation to 10X Genomics sequencing, with a UMAP of kidney cell clusters and a bar chart of cluster distributions across AKI, DKD, hypertension-associated CKD and living donor samples.

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Protocol(s)

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Regional transcriptomics

Goal: Generate deep transcriptomic signatures from nephron segments defined spatially by antibody staining using laser microdissection.

Regional transcriptomics workflow: kidney biopsy tissue is cryosectioned, laser-microdissected, RNA-sequenced and analyzed, and the data is uploaded to KPMP.

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Spatial transcriptomics - Visium

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Goal: Whole transcriptome mRNA expression with localization to kidney cells and structures

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snRNAseq clusters mapped onto a nephrectomy specimen. In each spot, the proportion of signature arising from each cell type is determined
snRNAseqclusters mapped onto a nephrectomy specimen. In each spot, the proportion ofsignature arising from each cell type is determined

Protocol(s)

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Spatial transcriptomics - Xenium

Spatial transcriptomics data generated using the 10x Genomics Xenium platform allows researchers to see where genes are active within a kidney tissue sample. By mapping gene activity directly to individual cells and their locations, Xenium provides a detailed view of how cells are organized and interact within the kidney.

This information helps researchers better understand how kidney diseases develop, identify which cells are involved in injury and repair, and discover new targets for treatments. By combining molecular data with tissue structure, Xenium offers a powerful tool for advancing kidney research and precision medicine.

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Protocol(s):

Formalin-Fixed Paraffin-Embedded (FFPE)

Liquid Nitrogen (LN2) & Carboxymethyl Cellulose (CMC)-Embedded Fresh-Frozen Samples 

  • 10X Xenium In Situ Gene Expression for Fresh-Frozen Kidney (VU_hKidney_300g) This is a protocol used by the Vanderbilt University Tissue Interrogation Site for preparing fresh-frozen kidney tissue for in situ spatial transcriptomics using a 300 gene custom probe panel with the 10X Xenium Analyzer platform. VU’s panel targets major/minor/rare kidney cell types, immune processes, and biochemical pathways related to cell signaling and lipid biosynthesis.

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10X Multiome (snRNA-seq + snATAC-seq)

Goal: 10X Genomics multiome protocol generates a molecular atlas of the human kidney with comprehensive cell types and minimal processing artifacts. 10X Multiome snATAC-seq + Gene Expression assay allows measurements of chromatin accessibility and transcription from the same cell. This allows direct insights into putative transcription factors or SNPs associated with open chromatin and impact on gene expression.

10X Multiome workflow for paired snRNA-seq and snATAC-seq: biopsy nuclei are processed on Chromium into chromatin accessibility and gene expression libraries, with quality thresholds for biopsy length, nuclei count and library yield.

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Metadata standards

Get metadata template

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