The Jurkat T cell line was obtained commercially (#TIB-152, ATCC, Manassas, VA) for initial nanoemulsion cell labeling characterizations. Jurkat cells were grown in RPMI-1640 media (Gibco, Waltham, MA) plus 10% fetal bovine serum (FBS), 10 mM HEPES buffer (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid), 1 mM sodium pyruvate and 1.5 mg/mL sodium bicarbonate.
Primary human T cells were obtained from blood samples sourced from the San Diego Blood Bank and enriched for T cells by Ficoll (Histopaque 1077, Sigma Aldrich) gradient density centrifugation and magnetic assisted cell sorting (Dynabeads, Thermo Fisher). T-cells were then activated with human T-activator CD3/CD28 Dynabeads and allowed to expand for two days in RPMI-1640 supplemented with 10% FBS and 100 units/mL of recombinant human interleukin 2 (IL-2, Peprotech, Rocky Hill, NJ). For transduction, we employed a vector specific to epidermal growth factor receptor variant III (EGFR-vIII) as described by Johnson et al. (30 ). A detailed CAR virus production and human T cell transduction is available elsewhere (31 (link)). CAR receptor expression was confirmed by flow cytometry. We used T cell populations with >70% CAR+ expression.
Primary human T cells were obtained from blood samples sourced from the San Diego Blood Bank and enriched for T cells by Ficoll (Histopaque 1077, Sigma Aldrich) gradient density centrifugation and magnetic assisted cell sorting (Dynabeads, Thermo Fisher). T-cells were then activated with human T-activator CD3/CD28 Dynabeads and allowed to expand for two days in RPMI-1640 supplemented with 10% FBS and 100 units/mL of recombinant human interleukin 2 (IL-2, Peprotech, Rocky Hill, NJ). For transduction, we employed a vector specific to epidermal growth factor receptor variant III (EGFR-vIII) as described by Johnson et al. (30 ). A detailed CAR virus production and human T cell transduction is available elsewhere (31 (link)). CAR receptor expression was confirmed by flow cytometry. We used T cell populations with >70% CAR+ expression.
