To use western
blotting for detection of cytochrome c release, a clean
fractionation should be performed in order to differentiate
mitochondrial from cytoplasmic fractions of cells.
Consequently, detection of cytochrome c in cytoplasmic
fraction is an indication of apoptosis. But, a risk of contamination
of cytoplasmic fraction with mitochondrial
fraction is always possible. Therefore, a mitochondrial
marker, called cytochrome c oxidase, should also be
employed for the detection of such contamination (Figure
7). In the case of the presence of this marker in
the cytoplasmic fraction, the experiment should not be
considered valid.