Kinetics of IR-induced γH2AX suggests more efficient DNA DSB repair in apoptosis-resistant ALLs. (A) (i) Following treatment with 2 Gy of IR and induction of a comparable number of DNA DSBs measured by γH2AX/MDC1 foci at 30 minutes, by 4 hours the resistant ALL 102 shows more efficient DNA repair and resolution of these foci, whereas sensitive ALL 114 retains foci at the same time point. Primary antibody binding was detected using Alexa Fluor 488 and Alexa Fluor 594–labeled secondary antibodies. Images were captured with a Nikon Eclipse E600 microscope (100×/1.40 NA oil objective) with a Hamamatsu digital camera C4742-95 and Volocity 4.2 software. (ii) Percentage of cells with more than 5 γH2AX/MDC1 foci 4 hours after IR is significantly higher in sensitive ALL 114 compared with resistant ALLs 102 and 106 (P = .04 and .02, respectively). The number of foci per cell was counted in 150 to 200 cells in randomly selected fields. The data are expressed as a percentage of cells with greater than 5 foci and the error bars represent the standard deviation from 3 independent counts. (B) Addition of PI3-kinase inhibitor LY294002 significantly delays disappearance of IR-induced foci at 4 hours after IR with 2 Gy in resistant tumor 102 (bottom panel), but shows no effect on resolution of foci in apoptosis-sensitive ALL 114 at the same time point (top panel).