Figure 5.

Staining of NB4 and HeLa-S3 cells with anti- cathepsin G antibodies or LysoTracker. (A) NB4 cells were incubated for 1 h at 37°C in media containing 50 μM LysoTracker peptide and then spun onto slides by using a cytocentrifuge. After fixation in neutral-buffered formalin followed by washing, the cells were incubated first with anti-cathepsin G antibodies and then with FITC-labeled secondary antibodies. Cells were prepared for viewing under the microscope by adding mounting media containing DAPI and coverslips. Photographs show DAPI plus cathepsin G staining, DAPI plus LysoTracker, DAPI alone, cathepsin G alone, or LysoTracker alone, as indicated. (B) HeLa-S3 cells were prepared as in A but were not incubated with LysoTracker. (C) NB4 cells were UV-irradiated as described in Materials and Methods; 1 h after irradiation, 50 μM LysoTracker was added to the cell media. After an additional hour of incubation, cells were spun onto a glass slide. After fixing and staining with anti-cathepsin G antibodies, cells were mounted by using DAPI-containing medium. Cell shape and chromatin condensation was used to distinguish apoptotic from nonapoptotic cells.