American Society of Hematology

Figure 2.

$Figure 2. Stable transgene expression in human primary PBLs by the SB transposon cis construct. Freshly isolated PBLs (5 × 106) were nucleofected at concentrations of 10 or 5 μg trans vector pT2/DsRed or cis vector pT2/DsRed//-SB10 using the T-cell Nucleofector kit (Amaxa, Gaithersburg, MD). On day 1 or 2 after nucleofection, a fraction of cells was analyzed for DsRed expression by flow cytometry, and the remaining cells were stimulated with anti-CD3/28 beads for 3 to 5 days. After removal of beads, the cells were cultured in human T-cell medium with IL-2 (50 IU/mL) and IL-7 (10 ng/mL). DsRed transgene expression was analyzed at the indicated time points by flow cytometry. The cells were restimulated once every 10 to 14 days. The data from 10 individual PBLs are shown.$

Stable transgene expression in human primary PBLs by the SB transposon cis construct. Freshly isolated PBLs (5 × 10⁶) were nucleofected at concentrations of 10 or 5 μg trans vector pT2/DsRed or cis vector pT2/DsRed//-SB10 using the T-cell Nucleofector kit (Amaxa, Gaithersburg, MD). On day 1 or 2 after nucleofection, a fraction of cells was analyzed for DsRed expression by flow cytometry, and the remaining cells were stimulated with anti-CD3/28 beads for 3 to 5 days. After removal of beads, the cells were cultured in human T-cell medium with IL-2 (50 IU/mL) and IL-7 (10 ng/mL). DsRed transgene expression was analyzed at the indicated time points by flow cytometry. The cells were restimulated once every 10 to 14 days. The data from 10 individual PBLs are shown.

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