Figure 5. Example of co-culture plate layout.

Columns 1–12 are typically used to test T-cells transfected with different T-cell receptors (TCRs). In this context, rows A–H are used to test reactivity of TCR-transfected T-cells under different conditions, e.g., with media only (negative control), in the presence of PMA/ionomycin (positive control), in the presence of one or more different tumor cell lines, and/or in the presence of immunomodulatory substances such as anti-PD-L1 antibodies [23]. It is advisable to keep the row with the PMA/ionomycin condition separated from the rows in which reactivity to tumor cells is tested in order to avoid accidental spillover of PMA/ionomycin-containing media. In case all rows on the plate are used, use rows A and B for PMA/ionomycin and media controls, respectively, as shown in figure. In case six or less rows are used, use row A for media control, rows B–F for testing conditions, and row H for PMA/ionomycin control. Figure created with BioRender.com.