Figure 2. In vitro gene expression at pH 7.4 and 6.8 transfection.

(a,b) The protein expression of p53 (a) and KillerRed (b) via DNA-complex or bPEI_25K/DNA-complex delivery in H1299 cells in culture medium at pH 7.4 or 6.8, 48 h post transfection. p53 protein in cell lysates was measured by ELISA assay using 450 nm absorbance for quantification. Confocal images showing the p53-mediated apoptosis was observed by TUNEL assay using a fluorescein. The fluorescence intensity and TE of KillerRed-positive cells were analysed by flow cytometry. Confocal images show KillerRed expression or ROS generation mediated by KillerRed photoactiviation was observed in transfected cells, in which the CellRox green dye and DAPI stains identify ROS and nuclei, taken 20 min exposure (593 nm, 100 mW cm⁻²). (c) Cell viability of transgene-expressing cells incubated at pH 7.4 or 6.8 with and without irradiation with a yellow laser, determined by the MTS assay. (d) Percentages of apoptotic cells stained by TUNEL assay incubated after a light exposure of 20 min and different pH values, analysed by flow cytometry. Confocal images show that the apoptotic cells were observed by TUNEL assay. All results show mean of measurements conducted in triplicate±s.d. Scale bar, 50 μm.