Imaging Chimeric Antigen Receptor (CAR) Activation

Methods Mol Biol. 2020;2111:153-160. doi: 10.1007/978-1-0716-0266-9_13.

Abstract

The chimeric antigen receptor (CAR) has been extensively exploited in cancer immunotherapy. In spite of the success of CAR T cells in clinical applications, the molecular mechanism underlying CAR-T cell activation remains unclear. Key questions remain: how are CARs activated by tumor antigens? How do activated CARs transduce signaling to downstream pathways? Here we introduce a microscopy-based method for studying CAR signaling. We use an antigen-coated supported lipid bilayer to activate CARs and combine it with TIRF microscopy to visualize the initial activation process of CAR T cells. This enables monitoring CAR signaling at high spatial and temporal resolutions.

Keywords: CD19; Chimeric antigen receptor; Microclusters; T cell signaling; TIRF.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antigens, Neoplasm / immunology*
  • HEK293 Cells
  • Humans
  • Jurkat Cells
  • Lipid Bilayers
  • Lymphocyte Activation
  • Microscopy, Fluorescence
  • Molecular Imaging / methods*
  • Neoplasms / immunology*
  • Receptors, Chimeric Antigen / metabolism*
  • Signal Transduction
  • T-Lymphocytes / immunology*

Substances

  • Antigens, Neoplasm
  • Lipid Bilayers
  • Receptors, Chimeric Antigen