Fig. 3.

CM-H₂DCFDA uptake and its ROS- mediated oxidation. (A) Measurement of uptake and oxidation of CM-H₂DCFDA. (A) Cells were pretreated with PEG-catalase followed by washing and treatment with H₂O₂. The changes in uptake and free radical-mediated oxidation of CM-H₂DCFDA were acquired for 15 min as described under Materials and methods. (B) Radiation-induced changes in intracellular ROS generation. The kinetics of radiation-induced changes in generation of ROS was measured at different time intervals after 0.5 Gy radiation by flow cytometry. After irradiation, cells were washed with PBS and incubated with CM-H₂DCFDA for 1 h at 37°C in the dark. Fluorescence (Fl) of CM-DCF was acquired at 488-nm excitation wavelength and 535-nm emission wavelength by flow cytometry and data were analyzed using FlowJo 6.4.1. Results are expressed as the mean fluorescence±SD of three independent experiments and statistical analysis was performed using Student’s t test (*p<0.05, **p<0.01, ***p<0.001).