Immunofluorescence analysis of HeLa cells treated with STS. A. Uninfected cells stained with antibodies against key points in the STS-induced apoptosis pathway, namely Bad activation at 1 hour, cytochrome c release at 2 hours, and activated caspase-3 and DNA damage with DAPI nuclear staining at 3 hours STS treatment. B. A monolayer of HeLa cells was infected with Shigella harboring a low copy number plasmid expressing GFP to allow green fluorescence. Left: Merged image of the monolayer for GFP and the activated form of caspase-3. Only the uninfected cells have a positive signal for caspase-3 activation. Right: Phase contrast view to visualize the bacteria in the cytoplasm of the HeLa cell. Images are representative of three repeated experiments, and cell counts of at least 300 cells for each treatment demonstrated consistent results (data not shown).