gamma-H2AX and 53BP1 foci induction by X-rays. (A) Immunofluorescence microscopy images were taken at 0.5h following X-irradiation of normal human fibroblasts. Each image is 20 μm wide. In the merged images 53BP1 is red, gamma-H2AX green and the nuclear margins are shown in blue. Co-localising foci appear yellow or orange. (B) Colocalisation analysis of gamma-H2AX and 53BP1 foci. Pearson’s correlation coefficients were calculated as described in . A value of one represents total co-localisation. The significance of correlation coefficients was determined for individual cells using Costes’ spatial statistics approach . Each point represents one cell. Filled red circles: non-significant, open green circles: significant correlation. Blue triangles, connected by blue line: mean correlation coefficient; error bars are standard errors from the analysis of 10–20 cells for each dose. (C) Gamma-H2AX versus 53BP1 foci count per cell, manually scored in the same 1,000 double-immunostained cells following exposure to a range of X-ray doses. Each data point corresponds to one cell. Shading of data point symbols reflects the number of coinciding points. The blue line indicates a 1:1 ratio.