EF1α Promoter Ensures Accurate Luciferase-Based Assessment of CAR-T Cell Cytotoxicity by Avoiding CMV-Driven Signal Bias
A recent study reveals that the EF1α promoter is more suitable than the CMV promoter for driving luciferase tag expression in in vitro cytotoxicity assays of CAR-T cells. The research shows that CAR-T cells can enhance luciferase expression from CMV-driven constructs via the IFN-γ/NF-κB signaling pathway, resulting in artificially elevated viability signals that do not accurately reflect target cell death. In contrast, EF1α-driven luciferase expression demonstrates strong concordance with flow cytometry-based measurements, providing a more reliable and precise evaluation of CAR-T cell cytotoxicity. These findings highlight the importance of promoter selection in assay design to ensure accurate and standardized assessment of immune cell therapeutics.
