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    glutathione-assay-kit-fluorometric-ab65322.pdf

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Glutathione Assay Kit (Fluorometric) (ab65322)

  • Datasheet
  • SDS
  • Protocol Booklet
Reviews (1)Q&A (4)References (19)

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Glutathione Detection Kit (Fluorometric) (ab65322)
  • Functional Studies - Glutathione Detection Kit (Fluorometric) (ab65322)

Key features and details

  • Assay type: Quantitative
  • Detection method: Fluorescent
  • Platform: Microplate reader
  • Assay time: 2 hr
  • Sample type: Cell culture media, Cell Lysate, Other biological fluids, Plasma, Serum, Tissue Extracts, Urine

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Overview

  • Product name

    Glutathione Assay Kit (Fluorometric)
    See all Glutathione kits
  • Detection method

    Fluorescent
  • Sample type

    Urine, Serum, Plasma, Other biological fluids, Tissue Extracts, Cell Lysate, Cell culture media
  • Assay type

    Quantitative
  • Assay time

    2h 00m
  • Species reactivity

    Reacts with: Mammals, Other species
  • Product overview

    Glutathione Assay Kit (Fluorometric) (ab65322) provides a simple in vitro assay for detection of total glutathione changes during cellular response to toxicity, apoptosis and other conditions. The assay uses the dye monochlorobimane (MCB), which forms an adduct with glutathione in a reaction catalyzed by glutathione-S-Transferase (GST). The unbound MCB is almost nonfluorescent, whereas it emits a fluorescent blue light (Ex/Em = 380nm/461nm) when bound to reduced or oxidized glutathione. Thus, the amount of glutathione can be easily detected using a fluorometer or a 96-well fluorometric plate reader.


    This product does not measure total glutathione in the sample. It measures the relative level of glutathione between untreated and treated samples.



    Visit our FAQs page for tips and troubleshooting.

  • Notes

    This product is manufactured by BioVision, an Abcam company and was previously called K251 Glutathione Fluorometric Assay Kit. K251-100 is the same size as the 100 test size of ab65322.

    Glutathione (GSH) is a tripeptide that contains L-cysteine, L-glutamic acid and glycine. It is the smallest intracellular protein thiol molecule in the cells, which prevents cell damage caused by reactive oxygen species such as free radicals and peroxides. Glutathione exists in reduced (GSH) and oxidized (GSSG) states. Reduced glutathione (GSH) is a major tissue antioxidant that provides reducing equivalents for the glutathione peroxidase (GPx) catalyzed reduction of lipid hydroperoxides to their corresponding alcohols and hydrogen peroxide to water. In the GPx catalyzed reaction, the formation of a disulfide bond between two GSH molecules generates oxidized glutathione (GSSG). The enzyme glutathione reductase (GR) recycles GSSG to GSH with the simultaneous oxidation of β-nicotinamide adenine dinucleotide phosphate (β-NADPH2). In healthy cells, more than 90% of the total glutathione pool is in the reduced form (GSH). When cells are exposed to increased levels of oxidative stress, GSSG accumulates and the ratio of GSSG to GSH increases. An increased ratio of GSSG-to-GSH is an indication of oxidative stress. The monitoring of reduced and oxidized GSH in biological samples is essential for evaluating the redox and detoxification status of the cells and tissues against oxidative and free radicals mediated cell injury.

    Abcam has not and does not intend to apply for the REACH Authorisation of customers’ uses of products that contain European Authorisation list (Annex XIV) substances.
    It is the responsibility of our customers to check the necessity of application of REACH Authorisation, and any other relevant authorisations, for their intended uses.

  • Platform

    Microplate reader

Properties

  • Storage instructions

    Store at -20°C. Please refer to protocols.
  • Components Identifier 100 tests
    Cell Lysis Buffer 1 x 25ml
    GSH Standard Yellow 1 vial
    GST Reagent Green 1 x 200µl
    Monochlorobimane Substrate Red 1 x 200µl
  • Research areas

    • Kits/ Lysates/ Other
    • Kits
    • Apoptosis Kits
    • Other Apoptosis Kits
    • Kits/ Lysates/ Other
    • Kits
    • Cell Damage Kits
    • Oxidative stress
    • Kits/ Lysates/ Other
    • Kits
    • Cell Metabolism Kits
    • Coenzyme and cofactor assay kits
    • Glutathione assay kits
    • Kits/ Lysates/ Other
    • Kits
    • Cell Damage Kits
    • Antioxidant
  • Relevance

    Glutathione is a small peptide composed of three amino acids: cysteine, glutamic acid, and glycine and is present in tissues in concentrations as high as one millimolar. Glutathione is the principal intracellular low-molecular-weight thiol that plays a critical role in the cellular defense against oxidative and nitrosative stress in mammalian cells. Diminished glutathione levels have been observed in the early stages of apoptosis.
  • Alternative names

    • GSH
    • GSSG

Associated products

  • Related Products

    • Ethacrynic acid, Glutathione S-transferase (GST) inhibitor (ab141399)
    • Dihydromyristicin, Glutathione S-transferase inducer (ab142290)
    • 10kD Spin Column (ab93349)

Images

  • Glutathione Detection Kit (Fluorometric) (ab65322)
    Glutathione Detection Kit (Fluorometric) (ab65322)Dasgupta A et al., PLoS One, 5, , 2010 Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/
    Glutathione pool measured in THP-1 macrophages: uninfected cells; WT: infected with M.tuberculosis wild type; KO: infected with M.tuberculosis OppD knock-out; COM: infected with M.tuberculosis OppD knock-out complemented with OppDA gene. 106 cells were infected and lysed by treating them with 100µl of ice cold lysis buffer. Cell lysate was diluted and mixed as described in the kit protocol. After 30 min incubation at 37C, fluorescence was measured at Ex=380nm/ Em=460nm. Results represent the means of ± S.D. of three determinations.
    Image obtained from Dasgupta A. et al; PLoS One; 2010 Aug 17; 5(8): e12225.
  • Functional Studies - Glutathione Detection Kit (Fluorometric) (ab65322)
    Functional Studies - Glutathione Detection Kit (Fluorometric) (ab65322)
    Glutathione assays were performed using various amounts of Glutathione as indicated. Results were analyzed according to the kit instructions.

Protocols

  • Protocol Booklet

Click here to view the general protocols

Datasheets and documents

  • SDS download

  • Datasheet download

    Download

References (19)

Publishing research using ab65322? Please let us know so that we can cite the reference in this datasheet.

ab65322 has been referenced in 19 publications.

  • Gay MD  et al. Targeting the Cholecystokinin Receptor: A Novel Approach for Treatment and Prevention of Hepatocellular Cancer. Cancer Prev Res (Phila) 14:17-30 (2021). PubMed: 33115780
  • Huang J  et al. HMGCR inhibition stabilizes the glycolytic enzyme PKM2 to support the growth of renal cell carcinoma. PLoS Biol 19:e3001197 (2021). PubMed: 33905408
  • Wang J  et al. Betalain exerts a protective effect against glaucoma is majorly through the association of inflammatory cytokines. AMB Express 10:125 (2020). PubMed: 32666339
  • Younis NS  et al. Protective Effect of Geraniol on Oxidative, Inflammatory and Apoptotic Alterations in Isoproterenol-Induced Cardiotoxicity: Role of the Keap1/Nrf2/HO-1 and PI3K/Akt/mTOR Pathways. Antioxidants (Basel) 9:N/A (2020). PubMed: 33053761
  • Manfredini F  et al. Rehabilitation Improves Mitochondrial Energetics in Progressive Multiple Sclerosis: The Significant Role of Robot-Assisted Gait Training and of the Personalized Intensity. Diagnostics (Basel) 10:N/A (2020). PubMed: 33080806
View all Publications for this product

Customer reviews and Q&As

Show All Reviews Q&A
Submit a review Submit a question

1-5 of 5 Abreviews or Q&A

Glutathione assay on bacteria

Excellent Good 4/5 (Ease of Use)
Abreviews
Abreviews
abreview image
Pros: The kit works with chemically defined media (CDM) which is a big bonus. Gram-positive and gram-negative bacterial supernatants both worked well. All steps of the kit are straightforward, but be careful of forming bubbles. Customer service while troubleshooting was helpful.

Cons: Intracellular bacterial levels of glutathione may be off - during cell lysis, glutathione turnover may be high. About half of the kit has to be used for controls and that is quite expensive.
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

Alex Klementiev

Verified customer

Submitted Nov 10 2022

Question

Does this kit measure total glutathione (GSH + GSSG) or just sample GSH concentration?

Read More

Abcam community

Verified customer

Asked on Mar 27 2014

Answer

This kit detects total glutathione.

Read More

Caitlin Valued Customer

Abcam Scientific Support

Answered on Mar 27 2014

Question

What plates are recommended for use with this kit?

Read More

Abcam community

Verified customer

Asked on Jan 31 2014

Answer

Black 96-well plates with transparent bottom (flat-bottom wells) can be used for bottom-reading fluorometers or opaque white plates can be used for top-reading fluorometers.

Read More

Jeremy Kasanov

Abcam Scientific Support

Answered on Jan 31 2014

Question

Hi,

I am using Greiner Thincert Culture Inserts for 6-well plates and A549 cells in my exposure experiments. The goal is to expose the cells at an air/liquid interphase for a certain period and to measure cell viability, oxidative stress markers and cell cycle stage after the exposure.

Until now I have tried to detach the cells from the inserts and measure the MTT assay and do flow cytometry analysis. However I have been having problems detaching the cells from the inserts, trypsin/EDTA is ineffective and scraping destroys the cells, so I have not been able to get sensible data from our exposure experiments.

Can you recommend any kits/procedures which could be applied directly in the inserts without prior detachments of the cells?

The minimum parameters I would need to evaluate after the exposure are cell viability and apoptosis/cell cycle stage, but I would like to include a marker for oxidative stress (e.g. glutathione) as well. I should be able to measure all the mentioned parameters simultaneously in one insert without too much quenching and fluorescence interference/overlay.

We have a Victor III microplate reader and an inverted fluorescence microscope for which we will acquire quantification software shortly.

Thank you for your help.

With best regards,

Read More

Abcam community

Verified customer

Asked on Mar 06 2012

Answer

Thank you for your enquiry and interest in our products.

MTT assay can be carried out on 96 well plates using either adherent or floating cells. You do not necessary need to remove (harvest) the cells by trypsinization. You can culture the cells in the wells of the 96 well plates, then apply the MTT solution (20ul) to each well. During incubation (37C, 5% CO2) for 1-5 hours MTT will be metabolized. The formazan (MTT metabolic product) can then be dissolved in DMSO and the optical density can be read at 560 nm using a plate reader - without harvesting the cells. There may be other cell viability/cytotoxicyprotovols but this is the most widely used.

Cell viability kits:

We have different cell viability assay kits in our catalogue i.e. ab112120, ab112121, ab112122, ab112123.

https://www.abcam.com/Cell-Viability-Assay-Kit-Fluorometric-Blue-ab112120.html

https://www.abcam.com/Cell-Viability-Assay-Kit-Fluorometric-Dual-Green-Red-ab112121.html

https://www.abcam.com/Cell-Viability-Assay-Kit-Fluorometric-Green-ab112122.html

https://www.abcam.com/Cell-Viability-Assay-Kit-Fluorometric-Near-InfraRed-ab112123.html

GSH kits:

GSH can be detected/quantified either by fluorimetric or chromogenic (DTNB - Ellman's reagent) assays and currently we have also some assay kits for detecting this tripeptide, you may wish to take a look at the datasheets for further information.

- ab65322: Glutathione Fluorometric Detection Kit

https://www.abcam.com/Glutathione-Fluorometric-Detection-Kit-ab65322.html

- ab112132:Intracellular GSH Assay Kit

https://www.abcam.com/Intracellular-GSH-Assay-Kit-ab112132.html

Normally, intracellular GSH levels can be expressed per cell numbers or per protein concentration.

I hope this helps and if I can assist further, please do not hesitate to contact me.

Read More

Abcam Scientific Support

Answered on Mar 06 2012

Question

Dear technical team, Our customer would like to purchase ab65322 and wants to confirm plasma samples are availble for this kit.   Please check this for our customer.     Best regards,

Read More

Abcam community

Verified customer

Asked on Dec 06 2011

Answer

Thank you for contacting us. The Glutathione Fluorometric Detection Kit (ab65322) can be used with a number of different types of sample: Cells, tissues or liquid samples such as plasma. As long as your sample contains Glutathione which can be detected the kit should be applicable to your customers sample.  A GSH standard is provided with the kit in order to produce the standard curve. I hope this information has been of help. If you require any further information please do not hesitate to contact us again.

Read More

Abcam Scientific Support

Answered on Dec 06 2011

Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
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