Fig 2: Leukemia cells repair themselves when damaged. (A) Cells were treated with VP16 for 2 h, and the drug was then washed away. Expression of ?-H2AX was detected by comet assay at 0, 2, 4, and 6 h. (B) After treatment of VP16 for 2 h, the expression of ?-H2AX was detected by flow cytometry at 0, 2, 4, and 6 h. The results were expressed as mean fluorescence intensity (MFI); n=3; 0 h, ****P<0.0001 (both cell lines); 2 h, ****P<0.0001 (both cell lines); 4 h, Kasumi-1 ***P<0.001, KG1a **P<0.01. There was no significant difference (ns) at 6 h. VP16, etoposide.

Fig 3: Comparison of ?-H2AX dose estimates.Foci yields were determined for X-irradiated and 2 h incubated blood samples processed with the lyse/fix versus histopaque protocol. At least 50 cells or 200 foci were manually scored per coded sample. Foci yields were converted to dose estimates using calibration data from Horn, Barnard & Rothkamm (2011). Actual doses were 0, 0.2, 0.6, 1.1, 1.2, 2.1 and 4.3 Gy. The line indicates the ideal 1:1 relationship.

Fig 4: Comparison of manual ?-H2AX foci counts.Different volunteer blood samples were processed by the lyse/fix or histopaque method following 0.5 Gy X-irradiation and 30 min incubation at 37°C.

Fig 5: Absence of Mapk7 in Atm-/- mice restores the DNA damage response to ionizing irradiationMice were treated with 6 Gy and two hours later, thymocytes were isolated and lysed in SDS-buffer or fixed for cell cycle analysis. A. Phosphorylation of H2AX (pH2AX) in thymocytes isolated from irradiated (IR) mice. Representative Western blot with pH2AX and tubulin as loading control (left); pH2AX:tubulin ratio normalized to control Mapk7hemat-/- (right; n = 3 for each genotype). n.s., not significant. B. Cell cycle distribution of live thymocytes isolated from the same mice evaluated in A., as analyzed by PI staining. C. Cell death of the same thymocyte samples evaluated in B. (sub-G1 fraction). D. Representative flow cytometry profiles of DNA content in thymocytes based on PI staining. The quantitation of these results is represented in B. for live thymocytes and in C. for dead cells.