Figure 2.

Scaffold promotes CAR-T cell ex vivo proliferation and activity. (A) Representative bright-field microscopy images and (B) SEM images of CAR-T cells after 3 days of culture within the scaffold. Scale bar, 50 μm. (C) The expansion curve of CAR-T cells that were untreated (mock), activated with aCD3/CD28 plate or scaffold. (D and E) CFSE-based cell proliferation analysis of CAR-T cells cultured under different conditions. (D) Representative flow cytometry histograms and (E) their relative MFI. n = 5, mean ± s.d. (F and G) The concentrations of human IL-2 and IFN-γ after co-culturing of CAR-T cells with aCD3/CD28 plate or scaffold. Cell number, 5 × 10⁵ CAR-T cells; microsphere number, 50 microspheres. n = 5, mean ± s.d. (H) The killing activity of CAR-T cells after activation on indicated days. n = 4, mean ± s.d. (I) Representative flow cytometric plots (left) and relative quantification (right) of CD8⁺ and CD4⁺ CAR-T cells among CD3⁺ CAR-T cells after 10 days of activation. n = 5, mean ± s.d. (J) Representative flow cytometric plots (left) and the average ratio (right) of PD-1⁺LAG-3⁺ CAR-T cells among CD3⁺ CAR-T cells after 10 days of activation. n = 5, mean ± s.d. Statistical analysis was performed using one-way ANOVA. *P < 0.05; **P < 0.01; ***P < 0.001.