Mitochondrial Protein Detection Protocol

Mitochondrial fractions were isolated to detect mitochondrial proteins using a mitochondria isolation kit for cell cultures (Catalog: 89874, Option B method) from Life Technologies, Thermo Fisher Scientific, according to the manufacturer's instructions. The cytosolic fraction from the mitochondrial preparation was further concentrated using a Microcon-10 kDa Centrifugal Filter Unit with Ultracel-10 membrane (Millipore) to detect cytosolic proteins. Protein was estimated by the Bradford assay and 20–30 µg of protein were used for western blotting. The purity of the mitochondrial fractions was established by the absence of cytosolic or nuclear proteins, LDH and Lamin B1, respectively, as described before (Devi et al., 2017 (link)).