Pohl Stephanie T, Prada Maria Llamazares, Espinet Elisa, Jurkowska Renata
Division of Biomedicine, School of Biosciences, Cardiff University, Cardiff, UK.
Division of Cancer Epigenomics, German Cancer Research Center (DKFZ) and Translational Lung Research Center, Heidelberg, Germany.
Methods Mol Biol. 2023;2584:371-387. doi: 10.1007/978-1-0716-2756-3_19.
Single-cell and single-nucleus RNA sequencing have revolutionized biomedical research, allowing analysis of complex tissues, identification of novel cell types, and mapping of development as well as disease states. Successful application of this technology critically relies on the dissociation of solid organs and tissues into high-quality single-cell (or nuclei) suspensions.In this chapter, we examine several key aspects of the tissue handling workflow that need to be considered when establishing an efficient tissue processing protocol for single-cell RNA sequencing (scRNA-seq). These include tissue collection, transport, and storage, as well as the choice of the dissociation conditions. We emphasize the importance of the tissue quality check and discuss the advantages (and potential limitations) of tissue cryopreservation. We provide practical tips and considerations on each of the steps of the processing workflow, and comment on how to maximize cell viability and integrity, which are critical for obtaining high-quality single-cell transcriptomic data.
单细胞和单细胞核RNA测序彻底改变了生物医学研究，能够对复杂组织进行分析、鉴定新型细胞类型以及绘制发育和疾病状态图谱。这项技术的成功应用严重依赖于将实体器官和组织解离成高质量的单细胞（或细胞核）悬液。在本章中，我们将探讨在为单细胞RNA测序（scRNA-seq）建立高效的组织处理方案时需要考虑的组织处理工作流程的几个关键方面。这些方面包括组织采集、运输和储存，以及解离条件的选择。我们强调组织质量检查的重要性，并讨论组织冷冻保存的优点（和潜在局限性）。我们针对处理工作流程的每个步骤提供实用技巧和注意事项，并就如何最大化细胞活力和完整性发表评论，这对于获得高质量的单细胞转录组数据至关重要。
