Table1_An Optimized Tissue Dissociation Protocol for Single-Cell RNA Sequencing Analysis of Fresh and Cultured Human Skin Biopsies.DOCX

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.本报告提出一种优化后的皮肤细胞悬液制备脱联协议，旨在为新鲜及培养的人体皮肤进行深度单细胞RNA测序（scRNA-seq）分析提供高质量样本。本协议能够从少量新鲜解离的皮肤活检组织中持续分离出高存活率的皮肤细胞，这些细胞被用于scRNA-seq研究。我们不仅重现了现有单细胞皮肤图谱中的主要细胞群体，还识别出如肥大细胞等罕见细胞群体。此外，我们有效地从体外培养的皮肤活检片段中分离出高存活率单细胞，并构建了移除人体皮肤的全球单细胞图谱。生成的scRNA-seq数据集的质量指标在新鲜解离与培养的皮肤之间具有可比性。总体而言，通过实现高效的细胞分离和全面的细胞图谱构建，我们的皮肤脱联-scRNA-seq工作流程能够极大地促进针对多种人体皮肤病理状态和体外皮肤移除实验的单细胞RNA测序发现。