The molecular mechanisms of DNA repair following chronic medium-dose-rate (MDR) γ-ray-induced damage remain largely unknown.
We used a cell function imager to quantitatively measure the fluorescence intensity of γ-H2A.X foci in MDR (0.015 Gy/h and 0.06 Gy/h) or high-dose-rate (HDR) (54 Gy/h) γ-ray irradiated… (More)
Figure 1. Detection of c-H2A.X intensity induced by c-ray irradiation. A. Number of c-H2A.X foci (foci/cell) in wild-type (C.B.17/DNA-PKcs+/ +Jcl) MEFs at 1 h after HDR c-ray irradiation (0, 0.54, 1.08, 1.67, 2.16, and 3.24 Gy) was determined by manual image counting. Numbers of c-H2A.X foci were counted in 100 cells at each point. These experiments were obtained by triplicated experiments. Average number of c-H2A.X foci per cell in wild-type MEFs increased in a dose-dependent manner. B. Image of c-H2A.X foci stained by anti-c-H2A.X antibody and Alexa 647-conjugated antimouse-Ig G. C. Image of nucleus stained by 49,6-diamino-2-phenylindole (DAPI). This image was automatically analyzed by a cell function imager (IN Cell Analyzer 1000). Circles traced in blue by the cell function imager indicate nuclei, and circles traced in green indicate c-H2A.X foci. D. Relation between total fluorescence intensity of c-H2A.X foci in nuclei and nuclear area, in non-irradiated cells. Nuclei are surrounded by a green line, as shown in Fig. 1B. Each point in this figure represents one detected cell. This result indicates that the intensity of c-H2A.X foci per nuclear area (I/A) is constant in non-irradiated cells. E. Relation between total intensity of c-H2A.X foci in nuclei and nuclear area, in cells irradiated by HDR at 3.24 G.y. F.