Fig. 3. Morphological analysis and DNA staining of A20 B-lymphoma cells treated with FasL. Untreated cells (A) or cells treated with 100 ng/ml FasL (B,C) for 16 hours were analyzed by electron microscopy. Necrotic FasL-treated cells (C) were characterized by disruption of the nuclei and abundance of vacuolar structures, whereas apoptotic cells observed in response to FasL were characterized by nuclear fragmentation, strong condensation of chromatin and membrane blebbing (B). The black bar shown for untreated cells (A) is equivalent to 5 µm. All images are shown at the same magnification. Alternatively, cells were left untreated (NT) or incubated with 100 ng/ml FasL for 16 hours, stained with PI and analyzed by confocal microscopy (D). As a comparison, non-treated cells are shown. Non-treated cells permeabilized with methanol (NT, Met-OH) and stained with PI (control, permeabilized) are shown as controls in Fig. 8A at the same magnification. Necrotic nuclei were PI positive, but retained a normal structural appearance (N); apoptotic nuclei were characterized by strong condensation of chromatin (A). Phase contrast and fluorescence images from the same optical section are shown. (E) Alternatively, hypodipliod and normodiploid cells were separated by FACS using PI fluorescence intensity as a parameter and analyzed subsequently by confocal microscopy (D). Images are shown at similar magnification. The bar in white is equivalent to 13 µm.