3D bioluminescence imaging assays for measuring in vitro CAR T cell cytotoxicity

For the past decades, many advances were made in the field of immunotherapies, most notably the development of chimeric antigen receptor (CAR) T cells, that allowed development of potent and targeted treatment options against cancers and other diseases. To determine the preclinical cytotoxic efficacy of T cell therapies, such as CAR T cell therapy, bioluminescence-based assays have emerged as a common technique, thanks to their high sensitivity, reproducibility and ease of use. Because the extracellular matrix (ECM) plays a key role in tumor development, numerous 3D cancer models have been generated to reduce the use of animals while simultaneously reproducing the tumor microenvironment during the testing of novel cell therapies. In this chapter, we describe a simple and reliable method to determine CAR T cell cytotoxicity in 3D matrices using bioluminescence detection, eliminating the need to harvest cells from the matrix.