Isolation of human cutaneous immune cells for single-cell RNA sequencing

Ashley A Hailer; David Wu; Abdullah El Kurdi; Michelle Yuan; Raymond J Cho; Jeffrey B Cheng

doi:10.1016/j.xpro.2023.102239

Isolation of human cutaneous immune cells for single-cell RNA sequencing

STAR Protoc. 2023 Apr 28;4(2):102239. doi: 10.1016/j.xpro.2023.102239. Online ahead of print.

Authors

Ashley A Hailer¹, David Wu², Abdullah El Kurdi³, Michelle Yuan¹, Raymond J Cho², Jeffrey B Cheng⁴

Affiliations

¹ Department of Dermatology, University of California, San Francisco, San Francisco, CA 94107, USA; Dermatology, Veterans Affairs Medical Center, San Francisco, CA 94121, USA.
² Department of Dermatology, University of California, San Francisco, San Francisco, CA 94107, USA.
³ Department of Biochemistry and Molecular Genetics, American University of Beirut, Beirut, Lebanon.
⁴ Department of Dermatology, University of California, San Francisco, San Francisco, CA 94107, USA; Dermatology, Veterans Affairs Medical Center, San Francisco, CA 94121, USA. Electronic address: jeffrey.cheng@ucsf.edu.

Abstract

Single-cell RNA sequencing (scRNA-seq) allows for high-resolution analysis of transcriptionally dysregulated cell subpopulations in inflammatory diseases. However, it can be challenging to properly isolate viable immune cells from human skin for scRNA-seq due to its barrier properties. Here, we present a protocol to isolate high-viability human cutaneous immune cells. We describe steps for obtaining and enzymatically dissociating a skin biopsy specimen and isolating immune cells using flow cytometry. We then provide an overview of downstream computational techniques to analyze sequencing data. For complete details on the use and execution of this protocol, please refer to Cook et al. (2022)¹ and Liu et al. (2022).².

Keywords: Cell Isolation; Flow Cytometry/Mass Cytometry; Immunology; Molecular Biology; RNAseq; Single Cell.

Published by Elsevier Inc.