Shown is a 0.5 micrometer thick confocal image of a midplane section of a 3T3 fibroblast pulsed for 5 min with BrdU and immuno-labeled with anti-BrdU antibodies. Replication is confined to discrete sites of replication. Arrows denote closely spaced sites.
Cells were fixed with 70% ethanol at -20C, DNA denatured with 4N HCl for 30 min at room temperature, and processed for immunofluorescence using anti-BrdU antibody and an appropriate fluorochrome labeled secondary antibody. Cells were examined in a MRC-1024 confocal microscope mounted on a Nikon Optphot 2 microscope with 60x 1.4 NA objective lens. See: Fig 1A in, Ma, H et al. 1998 Spatial and temporal dynamics of DNA replication sites in mammalian cells. J Cell Biol 143:1415-1425.
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