Transduction of Murine T Cells with Retroviral Vector

The transduction using the retroviral vector pMP71 and culture of primary murine T cells has been previously described [47 (link)]. In brief, 1.2 × 10⁶ virus producing 293Vec-Eco cells were seeded into a 6-well plate 24 h prior to splenocyte isolation. After 48 and 72 h, the virus-containing supernatant was used to transduce murine T cells. Murine T cells were expanded from murine splenocytes by activation with anti-CD3 and anti-CD28 antibodies (clones 145-2C11 and 37.51, Thermo Fisher Scientific, Waltham, MA, USA) and human IL-2 (10 IU/ml, Novartis, Basel, Switzerland) for 24 h. Subsequently murine T cells were stimulated Dynabeads™ Mouse T-Activator CD3/CD28 (Thermo Fisher Scientific) and human IL-15 (50 ng/ml, Peprotech). Prior to some experiments Dynabeads™ were removed.