使用高分辨率荧光显微镜进行单细胞 RNA 测序的细胞收集。

Collection of cells for single-cell RNA sequencing using high-resolution fluorescence microscopy.

机构信息

Department of Biomolecular Health Sciences, Faculty of Veterinary Medicine, Utrecht University, Utrecht, the Netherlands.

VYCAP B.V., Enschede, the Netherlands.

出版信息

STAR Protoc. 2021 Aug 5;2(3):100718. doi: 10.1016/j.xpro.2021.100718. eCollection 2021 Sep 17.

DOI:10.1016/j.xpro.2021.100718

PMID:34401784

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC8353340/

Abstract

FACS sorting followed by single-cell RNA-sequencing (SORT-Seq) is a popular procedure to select cells of interest for single-cell transcriptomics. However, FACS is not suitable for measurement of subcellular distribution of fluorescence or for small samples (<1,000 cells). The VYCAP puncher system overcomes these limitations. Here, we describe a workflow to capture, image, and collect fluorescent human retina pigment epithelium cells for SORT-Seq using this system. The workflow can be used for any cell type with a diameter of ∼5-50 μm. For complete details on the use and execution of this protocol, please refer to Segeren et al. (2020).

摘要

荧光激活细胞分选（FACS）后进行单细胞 RNA 测序（SORT-Seq）是一种用于选择单细胞转录组学中感兴趣细胞的常用方法。然而，FACS 不适用于测量荧光的亚细胞分布或小样本（<1000 个细胞）。VYCAP 打孔器系统克服了这些限制。在这里，我们描述了一种使用该系统捕获、成像和收集荧光人视网膜色素上皮细胞进行 SORT-Seq 的工作流程。该工作流程可用于任何直径约为 5-50μm 的细胞类型。有关此协议的使用和执行的完整详细信息，请参阅 Segeren 等人（2020 年）。