An Optimized Tissue Dissociation Protocol for Single-Cell RNA Sequencing Analysis of Fresh and Cultured Human Skin Biopsies

Front Cell Dev Biol. 2022 Apr 28;10:872688. doi: 10.3389/fcell.2022.872688. eCollection 2022.

Abstract

We present an optimized dissociation protocol for preparing high-quality skin cell suspensions for in-depth single-cell RNA-sequencing (scRNA-seq) analysis of fresh and cultured human skin. Our protocol enabled the isolation of a consistently high number of highly viable skin cells from small freshly dissociated punch skin biopsies, which we use for scRNA-seq studies. We recapitulated not only the main cell populations of existing single-cell skin atlases, but also identified rare cell populations, such as mast cells. Furthermore, we effectively isolated highly viable single cells from ex vivo cultured skin biopsy fragments and generated a global single-cell map of the explanted human skin. The quality metrics of the generated scRNA-seq datasets were comparable between freshly dissociated and cultured skin. Overall, by enabling efficient cell isolation and comprehensive cell mapping, our skin dissociation-scRNA-seq workflow can greatly facilitate scRNA-seq discoveries across diverse human skin pathologies and ex vivo skin explant experimentations.

Keywords: ex vivo explants; protocol; scRNAseq; skin biopsy; skin tissue.