MitoSciences (ab109902)

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

Overview

  • Product namePyruvate dehydrogenase (PDH) Enzyme Activity Microplate Assay Kit
    See all Pyruvate dehydrogenase kits
  • Detection methodColorimetric
  • Tests
    1 x 96 well plate
  • Sample type
    Cell culture extracts, Tissue
  • Assay typeEnzyme activity
  • Assay durationMultiple steps standard assay
  • Species reactivity
    Reacts with: Mouse, Rat, Cow, Human
  • Product overview

    This rapid activity microplate kit is used to determine the activity of PDH in a human, bovine, mouse, or rat sample. 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.

  • Notes

    Store 20X Buffer, Detergent, and Microplate at 4°C.

    Store 5X Stabilizer and 100X Reagent Dye at -20°C.

    Store 20X Reagent Mix and 100X Coupler at -80°C.

  • Tested applicationsFunctional Studiesmore details
  • PlatformMicroplate

Properties

  • Alternative names
    • PDC
    • PDH
    • PDHC

Applications

Our Abpromise guarantee covers the use of ab109902 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
Functional Studies Use at an assay dependent dilution.

Pyruvate dehydrogenase (PDH) Enzyme Activity Microplate Assay Kit images

  • Mitochondria, tissue extracts and whole cultured cell extracts all show linear relationships between signal and sample load at limiting concentrations. The rates shown were determined as change in OD over time, and these are best represented as change in milliOD per minute.
  • Example of microplate reader recorded data from bovine heart mitochondria (100 µg/well) (top trace) and 2-fold dilutions (stepwise lower traces) using Pyruvate dehydrogenase (PDH) Enzyme Activity Microplate Assay Kit (ab109902).
  • Schematic diagram showing the reaction involved in Pyruvate dehydrogenase (PDH) Enzyme Activity Microplate Assay Kit (ab109902).
  • Principle of Pyruvate dehydrogenase (PDH) Enzyme Activity Microplate Assay Kit (ab109902)

Protocols

References for Pyruvate dehydrogenase (PDH) Enzyme Activity Microplate Assay Kit (ab109902)

This product has been referenced in:
  • Prabhu A  et al. Cysteine catabolism: a novel metabolic pathway contributing to glioblastoma growth. Cancer Res 74:787-96 (2014). Functional Studies ; Human . Read more (PubMed: 24351290) »
  • Yamane K  et al. Diisopropylamine dichloroacetate, a novel pyruvate dehydrogenase kinase 4 inhibitor, as a potential therapeutic agent for metabolic disorders and multiorgan failure in severe influenza. PLoS One 9:e98032 (2014). Functional Studies . Read more (PubMed: 24865588) »

See all 15 Publications for this product

Product Wall

Thank you for contacting us. I have heard back from the lab: Yes, liver tissues were tested with ab109902; for example extract of mouse liver homogenate. For extract of liver homogenate, 100 – 500 ug/200 uL should be used, please see attached ex...

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Thank you for your reply.

The laboratory have advised that probably the best in this situation would be to maintain the DCA in all the cell washes after treatment and during the assay itself without NaF, so that full activation of PDH can be o...

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I contacted the laboratories who have confirmed that for phosphatase inhibition we recommend 10mM NaF for use with this kit. A 500mM stock solution can be prepared in water and stored at -20C.

ab65621 is an appropriate protease inhibitor...

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Please see below for further clarification regarding the following questions:

Is it OK to extract mitos from frozen tissues?

A: yes, you can extract mitos from frozen tissues but be aware that freezen/ thaw cycle would weaken the cell...

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Yes, you can use frozen tissues as sample for all three kits. As a reminder, please follow each individual protocol included with each kit to prepare samples for that particular assay.

Thank you for contacting us.



Nome of the reagents in this kit contain thymine.





I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information. Read More

Please find the answers to your questions from my colleagues at Miosciences, who are designing this kits:
1. The dephosphorylation modifications can be done after the immunocapture of the PDH enzyme to the plate. Many related concerns and question...

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Product ab110338 contains whole mitochondria in iso-osmotic buffer. There is no need to measure protein concentration of this product; the product is supplied at 5.5 mg/mL. Simply adjust the protein concentration with PBS to 5.3 mg/mL and add dete...

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There is no need to homogenize mitochondrial preparation in PBS.

I'd recommend measuring protein concentration of mitochondrial PBS suspension on an aliquot diluted in a detergent (for example 100x dilution of mitochondria suspension in 0.25%...

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It is recommended to prepare tissue homogenate in PBS at 23.7 mg/mL. One can use as a guideline the first part of ab110168 protocol. Just substitute all buffers with PBS. In general what you need to do is just mince/grind/homogenize tissue in PBS witho...

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Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"