In vitro assays to evaluate CAR-T cell cytotoxicity

Miao Cao; Robert D Carlson; Ross E Staudt; Adam E Snook

doi:10.1016/bs.mcb.2023.05.009

In vitro assays to evaluate CAR-T cell cytotoxicity

Methods Cell Biol. 2024:183:303-315. doi: 10.1016/bs.mcb.2023.05.009. Epub 2023 Oct 24.

Authors

Miao Cao¹, Robert D Carlson¹, Ross E Staudt¹, Adam E Snook²

Affiliations

¹ Department of Pharmacology, Physiology, & Cancer Biology, Thomas Jefferson University, Philadelphia, PA, United States.
² Department of Pharmacology, Physiology, & Cancer Biology, Thomas Jefferson University, Philadelphia, PA, United States; Department of Microbiology & Immunology, Thomas Jefferson University, Philadelphia, PA, United States; Sidney Kimmel Cancer Center, Thomas Jefferson University, Philadelphia, PA, United States. Electronic address: adam.snook@jefferson.edu.

PMID: 38548415
DOI: 10.1016/bs.mcb.2023.05.009

Abstract

This chapter introduces four commonly used in vitro chimeric antigen receptor (CAR)-T cell cytotoxicity assays (lactate dehydrogenase release assay, ⁵¹Cr release assay, IncuCyte live cell killing assay, and xCELLigence real-time analysis) and provides a detailed protocol for xCELLigence real-time analysis. Focusing on in vitro assays, this chapter starts with explaining the mechanisms and discussing the utilization of each assay to quantify T-cell-induced cytotoxicity. Due to the high-throughput quantification and straightforward workflow of xCELLigence real-time analysis, a protocol entailing reagents and equipment, a 3-day step-by-step procedure, and instructions for data analysis are provided.

Keywords: CAR-T cell; Cytotoxicity; Impedance; Real-time cell analysis; T cell; xCELLigence.

MeSH terms

Apoptosis*
Cell Line, Tumor
T-Lymphocytes*