Figure 6.

Analyses of DM–DNA replication and apoptosis in COLO 320DM cells. (A and B) A culture of rapidly growing COLO 320DM cells was pulse labeled with 10 μM BrdU for 1 h. The nuclei were isolated, fixed with PFA, hybridized with the biotinylated DM painting probe, and then detected using FITC-conjugated streptavidin as in Fig. 4. Sites at which BrdU was incorporated were detected with an anti-BrdU mouse monoclonal antibody followed by rhodamine-conjugated anti–mouse immunoglobulin. The double-labeled nuclei were examined using a confocal laser scanning microscope. The images of BrdU, FISH, and the merged images (red, BrdU; green, FISH) are shown for two representative fields. Nuclear buds that selectively entrap DMs are indicated by arrows, and the cells that were not in S phase during the pulse label (BrdU−) are indicated by arrowheads. (C) Analysis of apoptosis in COLO 320DM cells was done using the TUNEL method as described in Materials and Methods. This representative photograph is from one experiment in which COLO 320DM cells were treated with 100 μM HU for 3 d. The arrow points to a bud in a cell that is not undergoing apoptosis, whereas the arrowhead points to an apoptotic cell in the same field.