Purified CD22-CAR;PRODH2 and CD22-CAR;PRODH2(Stop) T cells (without cancer stimulation) were collected and washed with PBS, resuspended cell to 1e7 / mL in PBS and Cell-ID Cisplatin (Fluidigm) was added to a final concentration of 5 μM. Cells were incubated at room temperature for 5 min, then washed with Maxpar Cell Staining Buffer (Fluidigm). 1.5e6 CAR T cells per replicate were used for staining, each group has three replicates. Cells were stained with surface marker antibody cocktail first, then fixed and permeabilized. Second round staining was performed using cytoplasmic / secreted antibody cocktail. Finally, cells were incubated in intercalation solution (Fluidigm) in a final concentration of 125 nM, then incubated overnight at 4 °C. Before running on a CyTOF machine, cell concentrations were adjusted to 5–7e5/ mL with water. All data were collected on a CyTOF Helios instrument (Fluidigm). All surface and cytoplasmic / secreted antibodies were purchased from Fluidigm or Yale CyTOF core.
A list of CyTOF antibodies used in this study can be found inKRT .
A list of CyTOF antibodies used in this study can be found in