Fig. 2. In vitro characterization of the DART system.
a,b, SEAP levels 24 h after electrical stimulation in culture supernatants of HEK293 cells co-transfected with the DART constructs (pJH1003, pJH1004 and pJH1005). Stimulation for 15 s at the indicated voltages (a), or with 5 V DC for the indicated time periods (b). c, SEAP production kinetics over 72 h by engineered cells exposed to 5 V DC for the indicated periods of time. The induction factors were calculated between stimulated and non-stimulated group at 25 s. d, SEAP produced within 6 h by DART-engineered cells after exposure to 5 V DC for 20 s. The induction factors were calculated between the indicated groups. e, Reversibility of the DART switch. Engineered cells were alternately cultured for 24 h cycles in medium without electrostimulation (OFF) or treated with 5 V DC for 20 s (ON) and SEAP production was measured in the culture supernatants. Every 24 h, the culture medium was exchanged and the cell density was re-adjusted. f, SEAP produced by different mammalian cell lines transiently transfected with the DART constructs and stimulated with 5 V DC for 20 s. Non-stimulated cultures were used as controls. All data are mean ± s.d.; n = 4. P values were calculated between stimulated and non-induced control. Statistical designations with different colors refer to the corresponding data points with the same color (c).