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. 2020 Sep 13;12(9):2612. doi: 10.3390/cancers12092612

Figure 1.

Figure 1

iKP CAR construction and expression. (A) Schematic diagram of the bicistronic vector expressing iKP/iKPt CAR and CD19 CAR. (B–F) iKP-19-CAR-T/iKPt-19-CAR-T cells or CD19-CAR-T cells were cultured for 14 days in X-VIVO media supplemented with 100U/mL IL-2. Representative iKP/iKPt CAR and CD19 CAR expression in iKP-19-CAR-T/iKPt-19-CAR-T cells were detected on day 4 by flow cytometry using PE-anti-human KIR antibody and Alexa Flour 647-anti-mouse F(ab’)2 antibody (scFv of CD19 CAR is from mouse) (n = 4 different donors) (B). Detection of CD19 CAR-positive rate in iKP-19-CAR-T/iKPt-19-CAR-T and CD19-CAR-T on day 4, day 9 and day 14 by flow cytometry (n = 4 different donors) (C). Viability (D) or total cell numbers (E) of iKP-19-CAR-T/iKPt-19-CAR-T cells and CD19-CAR-T cells were also measured on day 4, day 9 and day 14 using Beckman Coulter counter (n = 4 different donors). Proportion of CD8+ and CD4+ T cell subsets in iKP-19-CAR-T/iKPt-19-CAR-T cells or CD19-CAR-T cells on day 4, day 9 and day 14 was measured using APC-anti-human CD8 antibody and PerCP-anti-human CD4 antibody (n = 4 different donors) (F). Three experiments were performed using PBMCs from each donor. Error bars represent ± SD.