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Malate Dehydrogenase 2 (MDH2) Activity Assay (ab119693) is used to determine mitochondrial malate dehydrogenase activity (MDH2) in a sample. The enzyme is captured within the wells of the microplate and activity is determined by following the production of NADH in the following MDH2 catalyzed reaction: malate + NAD → oxaloacetic acid + NADH (↑ Absorbance at 450 nm). The generation of NADH is coupled to the 1:1 reduction of a reporter dye to yield a colored (yellow) reaction product whose concentration can be monitored by measuring the increase in absorbance at 450 nm. In each well, ab119693 immunocaptures only native MDH2 from the chosen sample; this removes all other enzymes, including MDH1 in cytosol.
This product allows researchers to focus on TCA cycle, studying isotype-specific malate dehydrogenase (MDH2) activity assay without the necessity of isolating mitochondria.
Mitochondrial malate dehydrogenase (MDH2, P40926) is a 35.5 kDa enzyme that catalyzes the conversion of malate into oxaloacetate (using NAD+) and vice versa. (EC 126.96.36.199) Several isozymes of malate dehydrogenase exist, depending on where they are localized in the cell and their specific dependence on NAD+ or NADP+ (only in chloroplasts). There are two main isoforms in eukaryotic cells. One is found in the mitochondrial matrix (MDH2), participating as a key enzyme in the citric acid cycle that catalyzes the oxidation of malate. The other is found in the cytoplasm (MDH1), assisting the malate-aspartate shuttle with exchanging reducing equivalents so that malate can pass through the mitochondrial membrane to be transformed into oxaloacetate for further cellular processes. Because malate dehydrogenase is closely tied to the citric acid cycle, regulation is highly dependent on TCA products. High malate concentrations stimulate MDH activity, and, in a converse manner, high oxaloacetate concentrations inhibit the enzyme. Enzyme activity is enhanced by acetylation.
Storage: All components are shipped cold. Reagent dye, coupler, malate and NAD+ are shipped lyophilized. Before use rehydrate by adding 0.25 mL pure H2O to each tube and vortex each tube thoroughly to dissolve. After hydration unused amounts of these four materials should be stored at -80°C for 6 months. Store all other components at 4°C. This kit is stable for 6 months from receipt.
|Components||1 x 96 tests|
|100X Coupler||1 unit|
|100X NAD+||1 unit|
|100X Reagent Dye||1 unit|
|100X Sodium Malate||1 unit|
|10X Blocking Buffer||1 x 8ml|
|20X Buffer||1 x 20ml|
|Base Buffer||1 x 24ml|
|Extraction Buffer||1 x 15ml|
|MDH2 Microplate||1 unit|
Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"