Various techniques are currently used to assess T cell function in order to create more effective therapies, including bulk and single-cell RNA-Seq to measure gene expression; flow cytometry, live-cell imaging, and chromium release to assess target cell killing; and ntracellular cytokine staining, ELISA, and ELISPOT to measure cytokine expression. However, these techniques only allow us to draw correlative rather than causative conclusions because it is impossible to collect all of the required data from the same individual cell. For example, it is not possible to determine whether a specific cytokine-secreting cell can kill tumor targets or link this behavior directly to gene expression or genotype. This is particularly relevant in TCR-mediated therapies since the TCR sequence and associated affinity for antigen is directly linked to the degree of cytokine secretion and kinetics of target killing. Standard assays also make it challenging to discriminate between killing behaviors, such as multiple T cells killing multiple targets and single T cells capable of serial killing. They also make it difficult to assess key killing characteristics such as length of time in conjugation with tumor cell.
In this application note we demonstrate how the Berkeley Lights Opto™ Cell Therapy Development workflow can be used to:
- Correlate cytokine secretion to target cell killing behavior in CAR-mediated antigen recognition
- Discrimnate CAR T cell subsets based on kinetics of target cell killing
- Link cytokine secretion and target cell killing behavior to TCR sequence in TCR-mediated antigen recognition