Dish/vessel: 10 cm dish. Growth medium: DMEM 10 % HI FCS. Seeding: 3x10^6 per dish the day before. DNA : 30 µg. Reagent Amount: 30 µL. Ratio: 1:1. Complexation medium: DMEM. Complexation time: 15 min RT.

Redirected T cells genetically modified with a chimeric antigen receptor (CAR) induced spectacular remissions of so far refractory leukemia/lymphoma in early phase trials attracting interest to use CAR T cells in a variety of other applications including solid cancer and non-malignant diseases. However, extensive pre-clinical explorations demand highly effective and robust procedures for the genetic modification of blood T cells; the same applies for engineering with a recombinant T cell receptor (TCR). We present laboratory procedures in a step-by-step protocol to engineer human and mouse T cells with a CAR by retro- or lentiviral transduction for further pre-clinical testing.