Overview

  • Product name
    Thioredoxin Reductase Assay Kit (Colorimetric)
    See all Thioredoxin reductase kits
  • Sample type
    Urine, Cell culture extracts, Other biological fluids, Tissue Extracts
  • Assay type
    Enzyme activity
  • Assay time
    0h 40m
  • Product overview

    Thioredoxin Reductase Assay Kit (ab83463) is a specific assay for detecting Thioredoxin Reductase (TrxR) activity in various samples. TrxR catalyzes the reduction of DTNB (5, 5'-dithiobis (2-nitrobenzoic) acid) to TNB2- (5-thio-2-nitrobenzoic acid) in presence of NADPH, which generates a strong yellow color (ODmax = 412 nm). Other enzymes present in crude biological samples such as glutathione reductase and glutathione peroxidase can also reduce DTNB. In order to measure TrxR-only activity, a TrxR specific inhibitor is used in a separate reaction to determine TrxR specific activity. The difference between total DTNB reduction in the sample and DTNM reduction in the sample in presence of TrxR inhibitor is the value of specific TrxR activity in the sample.
    Visit our FAQs page for tips and troubleshooting.

  • Notes

    Thioredoxin reductase (TrxR, EC 1.8.1.9) is a ubiquitous mammalian enzyme that catalyzes the NADPH-dependent reduction of the redox protein thioredoxin, as well as of other endogenous and exogenous compounds such as selenite, lipid hydroperoxides and hydrogen peroxide. TrxR is the only enzyme known to catalyze the reduction of thioredoxin in order to form reduced disulfide bounds in cells, and it is likely that also plays a role in protection against oxidant injury, cell growth and transformation.

  • Tested applications
    Suitable for: Functional Studiesmore details

Properties

  • Storage instructions
    Store at -20°C. Please refer to protocols.
  • Components Identifier 100 tests
    DTNB Red 1 vial
    NADPH Blue 1 vial
    TNB Standard Brown 1 vial
    TrxR Assay Buffer WM 1 x 25ml
    TrxR Inhibitor Clear 1 vial
    TrxR Positive Control Green 1 vial
  • Research areas
  • Relevance
    Thioredoxin reductase (TrxR) (EC 1.8.1.9) is a ubiquitous enzyme which is involved in many cellular processes such as cell growth, p53 activity, and protection against oxidation stress, etc. The mammalian TrxR reduces thioredoxins as well as non-disulfide substrates such as selenite, lipoic acids, lipid hydroperoxides, and hydrogen peroxide.
  • Cellular localization
    TXNRD1: Cytoplasmic. TXNRD2: Cytoplasmic. Nuclear. Microsome. Endoplasmic reticulum. TXNRD3: Mitochondrial.
  • Alternative names
    • EC 1.8.1.9
    • gene associated with retinoic and interferon-induced mortality 12 protein
    • Gene associated with retinoid IFN induced mortality 12 protein
    • GRIM 12
    • KM 102 derived reductase like factor
    • KM-102-derived reductase-like factor
    • MGC9145
    • oxidoreductase
    • Selenoprotein Z
    • SELZ
    • TGR
    • Thioredoxin and glutathione reductase
    • Thioredoxin glutathione reductase
    • Thioredoxin reductase 1
    • thioredoxin reductase 1, cytoplasmic
    • Thioredoxin reductase 2
    • thioredoxin reductase 2, mitochondrial
    • Thioredoxin reductase 3
    • Thioredoxin reductase beta
    • Thioredoxin reductase GRIM 12
    • thioredoxin reductase TR1
    • thioredoxin reductase TR2
    • thioredoxin reductase TR3
    • TR
    • TR beta
    • TR1
    • TR2
    • TR3
    • TrxR
    • TRXR1
    • TRXR2
    • TRXR3
    • TXNR
    • TXNRD1
    • TXNRD2
    • TXNRD3
    see all

Applications

Our Abpromise guarantee covers the use of ab83463 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.

Images

  • Thioredoxin reductase measured in mouse tissue lysates showing activity (mU) per mg protein of sample tested

  • Thioredoxin reductase measured in cell lysates showing activity (mU) per 1 mln of cells tested

  • Standard curve (colourimetric) : mean of duplicates (+/-SD) with background readings substracted

  • Thioredoxin reductase Kinetic Data using ab83463.
  • Activity of endogenous Brugia thioredoxin reductase from soluble worm lysates following incubation with 1% DMSO or 0.3 μM, 0.1 μM, or 0.03 μM of auranofin in vitro. Percentages indicate the percent activity of TrxR compared to DMSO controls. Thioredoxin reductase activity was significantly reduced (p < 0.05) to 15%, 33% and 69% of endogenous activity, respectively, compared to the activity in DMSO-treated worms.

    Thioredoxin reductase activity of worm lysates was assayed using female B. malayi treated in vitro with either 0.3 μM, 0.1 μM, or 0.03 μM auranofin or 1% DMSO. After 5 hours of treatment, worm motility was measured using the Worminator, and then worms (24 in each group) were pooled, washed three times in PBS, and lysed by douncing in a glass homogenizer in assay buffer (ab83463) with 1 mM PMSF. The crude lysates were centrifuged at 10,000 rcf for 15 minutes at 4°C to pellet insoluble material. The total protein concentrations of soluble lysates were measured using the Bradford assay. The soluble lysates were incubated for 20 minutes in assay buffer or assay buffer with a proprietary thioredoxin reductase specific inhibitor before adding a specific substrate, DTNB (5, 5′-dithiobis (2-nitrobenzoic) acid), and measuring activity at 20 second intervals for 40 minutes using the SpectraMax Plus Microplate Reader (Molecular Devices, Sunnyvale, CA) at λ = 412 nm. Lysates were tested in duplicate. TrxR activity was calculated based on the linear amount of TNB produced per minute per mg of total protein and adjusted for background activity from enzymes other than TrxR in the lysates.

Protocols

References

This product has been referenced in:
  • Ahmad F  et al. TGF-ß-induced hCG-ß regulates redox homeostasis in glioma cells. Mol Cell Biochem 399:105-12 (2015). Read more (PubMed: 25300619) »
  • Bulman CA  et al. Repurposing auranofin as a lead candidate for treatment of lymphatic filariasis and onchocerciasis. PLoS Negl Trop Dis 9:e0003534 (2015). Functional Studies . Read more (PubMed: 25700363) »

See all 3 Publications for this product

Customer reviews and Q&As

Ab83463 can be used to measure TrxR in plasma. It does not need any special preparation. It can be used directly or diluted depending on the TrxR content.

Several volumes must be tested in a pilot assay to ensure readings are within the linear...

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I wanted to let you know that I have been informed by the lab that there is EDTA in this kit.

Thank you for contacting us.

The standard in this kit is from rat.

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

Thank you for contacting us.

I can confirm, that regrettably we can not provide any data using serum samples with this kit. However, every sample type and application stated for a product on our datasheets is covered by our six month guarantee...

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Thank you for contacting us and your interest in our products.

I can confirm this product is sourced externally and the molar concentration of the TrxR inhibitor is proprietary information. Regrettably we are not allowed to chare this informat...

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I have had some feedback from the lab to share with you which may help to improve the results seen so far.


The assay buffer needs to be allowed to come to room temperature before use. This may have significantly reduced the reactivity o...

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Thank you for providing that information.

Could I ask you confirm a few additional details?

1. How was the kit stored for the month in which it was not used?

2. In particular, how was the NADPH kept, both before reconstitu...

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Thank you for contacting us and reporting the problems experienced with the Thioredoxin (TrxR) Reductase Assay Kit (ab83463). From having spoken to you over the phone, there does not immediately appear to be an obvious solution to what you have been ob...

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Thank you for contacting us. The Thioredoxin (TrxR) Reductase Assay Kit ab83463 contains a TrxR specific inhibitor that is utilized to determine TrxR specific activity. Unfortunately, we have not tested the kit or the inhibitor with insect cells. ...

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Thank you for contacting us.  The Thioredoxin (TrxR) Reductase Assay Kit ab83463 contains a TrxR specific inhibitor that is utilized to determine TrxR specific activity. Unfortunately, we have not tested the kit or the inhibitor with insect cells. ...

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

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