Pyruvate Dehydrogenase (PDH) Activity

PDH activity was measured in VN homogenates by using the Pyruvate Dehydrogenase (PDH) Enzyme Activity Microplate Assay Kit (Abcam) according to manufacturer’s instructions. Briefly, mitochondria isolated by double centrifugation (50 μg) were loaded into the wells of a 96-well plate that was specific for PDH activity assay. PDH enzyme was immunocaptured by the monoclonal antibodies coated on the wall of each well. The enzymatic activity of PDH was determined based on the production of NADH coupled to a reporter dye whose formation is monitored by measuring the increase in absorbance at 450 nm on a Varioskan LUX plate reader (Thermo Fisher Sci.).