The indicated rat tissue was processed using the Qproteome Cell Compartment Kit. The respective fractions (C: cytosol, M: membrane, N: nucleus, and S: cytoskeleton) were separated by SDS-PAGE and transferred by western blotting to nitrocellulose membrane. The blot was probed with antibodies against the indicated cell-compartment-specific marker proteins.
Western blots of fractionated NIH 3T3 cells. Protein (20 µg) from the cytosolic (C), membrane (M), and nuclear (N) fractions was separated by SDS-PAGE. After Western blotting, proteins specific to each fraction were detected using [A] annexin, [B] TIM23, and [C] lamin antibodies, and an HRP-conjugated secondary antibody with chemiluminescent detection.
Micrographs (upper panels) of HeLa cells [A] under normal growth conditions and [B] after 4 hour incubation with staurosporine, an inducer of apoptosis. Lower panels show western blots prepared after cell lysates were processed using the Qproteome Cell Compartment Kit. The blots were probed with anti-cytochrome c antibody. It is known that during apoptosis cytochrome c is translocated from the intermembrane space of mitochondria to the cytosol, a process that is reflected in the detection of cytochrome c in the cytosolic fraction of apoptotic cells.