Abcam, ab109902, Pyruvate dehydrogenase (PDH) Enzyme Activity Microplate Assay Kit

Size: 96Test
Pyruvate dehydrogenase (PDH) Enzyme Activity Microplate Assay Kit (ab109902) can be used to determine the activity of PDH in a human, bovine, mouse, or rat sample. Readout on any colorimetric (450 nm) plate reader. - Cited in > 100 publications
Key facts
Detection method:Colorimetric,
Sample types:Tissue Lysate, Cell culture extracts, Purified mitochondria,
Reacts with:Mouse, Rat, Cow, Human,
Assay type:Enzyme activity,
Assay time:3h 30m,
Assay Platform:Microplate reader

Product details:
The Pyruvate Dehydrogenase (PDH) Enzyme Activity Microplate Assay Kit works by immunocapturing the PDH enzyme in microplate wells and measuring its activity through the reduction of NAD+ to NADH, which is linked to a color change detected at 450 nm.
How the assay works
In Pyruvate dehydrogenase (PDH) Enzyme Activity Microplate Assay Kit protocol, the PDH enzyme is immunocaptured within the wells of the microplate and activity is determined by following the reduction of NAD+ to NADH, coupled to the reduction of a reporter dye to yield a colored reaction product with an increase in absorbance at 450 nm at room temperature. Included for performance of the activity assay are buffer, detergent, reagent mix, and a 96-well microplate with monoclonal antibody pre-bound to the wells of the plate, allowing for a stream-lined assay.
This assay is optimized for use with whole tissue extract when the amount of total material available for assay is 20-100 µg or more. If using cell extract of cultured cells 1 mg of material is required due to the very low levels of enzyme and reduced levels of mitochondria in the extract.
The PDH complex is relatively fragile and sensitive to detergent. Please follow the sample preparation steps provided in the protocol booklet. Other preparation methods may decrease PDH activity.
Pyruvate dehydrogenase (PDH) Enzyme Activity Microplate Assay Kit protocol summary:
- Add samples, standards and positive control to wells
- Incubate for 3 hours at room temperature
- Add 200 µL of Assay Solution to each well
- Measure Optical Density (OD450 nm) in a kinetic mode at RT for 30 minutes
How other researchers are using
Pyruvate dehydrogenase (PDH) Enzyme Activity Microplate Assay Kit has been used in a variety of sample type including:
Mouse cardiomyocytes
Mouse muscle tissue samples
Human iPSC-derived cardiomyocytes
References:
1-Mao Y et al. 2023
2-Brenmoehl J et al. 2023
3-Srivastava S et al. 2023
Related and recommended products
See an alternative kit to quantify Pyruvate dehydrogenase (PDH) Enzyme Activity:
- Pyruvate dehydrogenase (PDH) Enzyme Activity Dipstick Assay Kit
ab109882
Store 20X Buffer, Detergent, and Microplate at 4°C.
Store 5X Stabilizer, 20X Reagent Mix, 100X Reagent Dye and 100X Coupler at -80°C.

Properties and Storage Information:
Shipped at conditions-Dry Ice, Appropriate short-term storage conditions-Multi, Appropriate long-term storage conditions-Multi, Storage information-Please refer to protocols

Supplementary Information:
This supplementary information is collated from multiple sources and compiled automatically.
Pyruvate dehydrogenase (PDH) is an enzyme complex also known as the pyruvate dehydrogenase complex (PDC) or PDH complex. It is a multi-enzyme structure with a mass on the order of megadaltons and plays an important role in cellular energy metabolism. Situated in the mitochondrial matrix pyruvate dehydrogenase converts pyruvate into acetyl-CoA through oxidative decarboxylation. This conversion releases one molecule of CO2 and reduces NAD+ to NADH. The complex includes three core enzymes: E1 (pyruvate dehydrogenase) E2 (dihydrolipoamide acetyltransferase) and E3 (dihydrolipoamide dehydrogenase).
Biological function summary
The actions of pyruvate dehydrogenase serve as a bridge between glycolysis and the tricarboxylic acid (TCA) cycle. Pyruvate derived from glucose is transformed into acetyl-CoA before entering the TCA cycle for further energy extraction. The PDH complex ensures efficient energy production by tightly regulating the flow of carbon into the TCA cycle. Regulation occurs through phosphorylation by specific PDH kinases which inactivate E1. This mechanism integrates signals from energy status and substrates availability modulating the carbohydrate metabolism.
Pathways
Pyruvate dehydrogenase is a central player in cellular respiration and energy metabolism. It connects glycolytic pathways with the TCA cycle facilitating energy conversion in eukaryotic cells. Key related proteins involve pyruvate kinase (which generates pyruvate) and citrate synthase (which uses acetyl-CoA) ensuring synchronized activity between upstream and downstream metabolic processes. The proper function of PDH activity is necessary for maintaining the metabolic flow with the PDH complex serving a gating role in the energy pathways.
Pyruvate dehydrogenase deficiency results in metabolic challenges as the inability to convert pyruvate efficiently causes an increase in lactate levels. This condition results in lactic acidosis and severe neurological dysfunction. Furthermore alterations in PDH activity are observed in various forms of cancer as cancer cells often rely on aerobic glycolysis (Warburg effect) rather than complete oxidation of glucose. In this context the PDH protein interacts with oncogenic pathways highlighting its role in tumor metabolism and potential therapeutic targeting.