Evaluating RevCAR T Cell Cytotoxicity

In order to evaluate the cytotoxic potential of RevCAR T cells, co-injection experiments were performed in vivo using six weeks old male NMRI-nude mice (Rj:ATHYM-Foxn1nu/nu) obtained by Janvier Labs. All mice were randomly assigned to experimental groups of five individuals and housed in sterile cages in a pathogen-free facility with a 12 h light/dark cycle. 1*10⁶ LNCaP^PSCA+/Luc+ cells were injected subcutaneously in the presence or absence of RevCAR T cells (1*10⁶) and RevTM (102 µg) in a total volume of 100 µl PBS (Gibco™ Cat#10010015). Subsequently, anesthetized mice were i.p. injected with 200 µl D-Luciferin (15 µg/ml in PBS, PerkinElmer Cat#122799) and analyzed for bioluminescence in the planar X-ray (Bruker In-vivo Xtreme) at different time points. Measured luminescence was evaluated using the Bruker MI and Multispectral Software as described previously.^{43 (link),44 (link)} All in vivo studies have been approved by the local ethics committee for animal experiments (Landesdirektion Dresden, 24–9165.40-4/2013, 24.9168.21–4/2004-1) and were performed at the Helmholtz-Zentrum Dresden-Rossendorf (HZDR) in appliance with the terms of German regulations of animal welfare.