Fig 1. Characterization of laser induced DNA damage.

(A) At the selected irradiance of 3.0x10¹¹W/cm² phase contrast changes are observed at a laser damage site on prometaphase chromosomes in nocodazole synchronized U2OS cells (4 and 6s). At 16s post laser (see arrows) phase dark material is apparent. Scale bar = 1μm. (B) γH2AX forms around the chromosome region damaged by the laser. A metaphase cell was fixed ~5s after the laser and immnunostained for γH2AX (arrows and inset). This cell was fixed prior to dark material accumulation. Paling is observed in the inset. (C) Positive TUNEL is seen at laser damaged regions (arrows) in prometaphase cells incubated with nocodazole. Two different chromosomes were damaged at different time points within the same cell. The first damage was induced 30 minutes prior to fixation and the second damage was induced 20 minutes after the first damage and fixed after 10 minutes. The TUNEL signal at the second damage site was brighter than the signal at the first damage. A graph of TUNEL signal on the right is the average of six cells per category, n = 6. The average and standard error mean of the late cut (fixed 10 minutes post laser) is 195 ± 61 and 23 ± 8 for the early cut (fixed 30 minutes post laser). p = 0.0363 Scale bar = 1цm.