I have been trying to activate CD19 CAR-T cells using plate-bound anti-CAR Abs. I am getting activation both in cell lines and primary CD8 T-cells. I am using pErk as a readout for activation. I see Erk phosphorylation peak at 5min and is sustained for a little longer than 10min. However, in case of primary CD8 T-cells, in my negative control (well with no Ab coating), I am also getting a lower but similar pattern where %pErk peaks at 5min. I do not see this with my cell line. Also, I did not see this when I had the primary cells rested on the plate for more than 30min and stimulated them with CD19+ tumor cells. Does anyone have any experience with this? Is there a way to reduce the Erk activation in the unstimulated group?
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