Fig. 5. Apoptosis-associated cytochrome c-release from mitochondria into the cytosol of human-derived HL-60 cells is inhibited by T. gondii. HL-60 cells were infected at a parasite to host ratio of 30:1 and were then treated with 5 µg/ml actinomycin D for 8 hours. Equal numbers of cells per treatment were fractionated into digitonin-soluble and -insoluble extracts, containing cytosolic and mitochondrial proteins, respectively. Lysates were separated by SDS-PAGE and were then transferred to nitrocellulose membranes. Immobilized proteins were probed with a cytochrome c-specific (A) or a cytochrome c oxidase (COX) subunit IV-specific (B) monoclonal antibody and an appropriate secondary antibody using enhanced chemiluminescence detection. COX served as a marker for mitochondrial contamination of the digitonin-soluble extracts.
