Figure 2. Nucleofection of RNPs leads to highly efficient target gene KO in nonactivated human T cells. (A) Systematic optimization of nucleofection parameters for RNP transfection of nonactivated human CD4+ T cells. Analysis of transfection efficiency (ATTO550 expression and MFI), cell viability, and CXCR4 KO frequency 48 h after transfection. Data are from one experiment. (B) Examples of transfection efficiency (ATTO550-labeled tracrRNA) with different nucleofection pulses 48 h after transfection (gated on live cells). (C) Comparison of KO efficiency and cell viability by flow cytometry 72 h after RNP transfection using selected nucleofection pulses and buffers for targeting CXCR4 in nonactivated human CD4+ T cells. Data are presented as mean ± SD (n = 2) and representative of two independent experiments. (D) CXCR4 expression by flow cytometry 72 h after RNP transfection using the EH100/P2 condition. (E) KO efficiency as measured by flow cytometry using optimized RNP transfection in nonactivated human CD4+ T cells targeting CXCR4, CCR7, CD127, or IFN-γ compared with target expression in cells transfected with NTC. Data are presented as mean ± SD (n = 2) and representative of two independent experiments. **, P < 0.01; ***, P < 0.001; ****, P < 0.0001 by one-way ANOVA.
Figure 2.

Nucleofection of RNPs leads to highly efficient target gene KO in nonactivated human T cells. (A) Systematic optimization of nucleofection parameters for RNP transfection of nonactivated human CD4+ T cells. Analysis of transfection efficiency (ATTO550 expression and MFI), cell viability, and CXCR4 KO frequency 48 h after transfection. Data are from one experiment. (B) Examples of transfection efficiency (ATTO550-labeled tracrRNA) with different nucleofection pulses 48 h after transfection (gated on live cells). (C) Comparison of KO efficiency and cell viability by flow cytometry 72 h after RNP transfection using selected nucleofection pulses and buffers for targeting CXCR4 in nonactivated human CD4+ T cells. Data are presented as mean ± SD (n = 2) and representative of two independent experiments. (D) CXCR4 expression by flow cytometry 72 h after RNP transfection using the EH100/P2 condition. (E) KO efficiency as measured by flow cytometry using optimized RNP transfection in nonactivated human CD4+ T cells targeting CXCR4, CCR7, CD127, or IFN-γ compared with target expression in cells transfected with NTC. Data are presented as mean ± SD (n = 2) and representative of two independent experiments. **, P < 0.01; ***, P < 0.001; ****, P < 0.0001 by one-way ANOVA.

This Feature Is Available To Subscribers Only

or Create an Account

Close Modal
Close Modal