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. 2020 Jun 5;17(10):1406–1414. doi: 10.7150/ijms.45603

Figure 3.

Figure 3

In vitro functional analysis of EGFRvIII CAR-T cells. (A) Representative flow cytometry analysis of transduction efficiency of EGFRvIII CAR. Surface expression of CAR on T cells was detected by anti-mouse F(ab)2-Alexa 647. NT T and MOCK were used as control. NT T: Non-transduced T cells MOCK: T cells expressing CARs that encode the truncated CD3ζ. (B) Immunoblot analysis of CAR expression. Lysates were separated by SDS-PAGE under reducing condition. Mouse anti-human CD3ζ antibody was used to detect the endogenous and chimeric CD3ζ expression. 1: NT T cells, 2: MOCK, 3: EGFRvIII CAR transduced T cells. (C) Immunoblot analysis of EGFRvIII expression in target cells. 1:EGFRvIII-U87 cells, 2:SMMC7721, 3:U87 cells. (D)When co-incubated with SMMC7721 cells, EGFRvIII CAR-T cells cytotoxicity was measured by Non-Radioactive Cytotoxicity kit. CAR-T cells which were co-cultured with EGFRvIII-U87 or U87 cells were used as positive or negative control, respectively (n=3). (E) Cytokine production of CAR-T cells was quantified by ELISA. EGFRvIII-U87 and U87 cells were used as positive and negative target cell control (n=3). ***P <0.001,*P <0.05.