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GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green) (ab138881)

  • Datasheet
  • SDS
  • Protocol Booklet
Reviews (5)Q&A (10)References (109)

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Functional Studies - GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green) (ab138881)
  • Functional Studies - GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green) (ab138881)
  • Functional Studies - GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green) (ab138881)
  • Functional Studies - GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green) (ab138881)
  • Functional Studies - GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green) (ab138881)
  • Functional Studies - GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green) (ab138881)
  • Functional Studies - GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green) (ab138881)
  • Sample Calibration Curve - Total GSH
  • Sample Calibration Curve - Reduced GSH

Key features and details

  • Assay type: Quantitative
  • Detection method: Fluorescent
  • Platform: Microplate reader
  • Assay time: 30 min
  • Sample type: Cell Lysate, Plasma, Serum, Tissue Extracts, Urine
  • Sensitivity: 10 nM

You may also be interested in

Assay
Product image
GSH/GSSG Ratio Detection Assay Kit II (Fluorometric - Green) (ab205811)

View more associated products

Overview

  • Product name

    GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green)
    See all Glutathione kits
  • Detection method

    Fluorescent
  • Sample type

    Urine, Serum, Plasma, Tissue Extracts, Cell Lysate
  • Assay type

    Quantitative
  • Sensitivity

    10 nM
  • Assay time

    0h 30m
  • Product overview

    GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green) (ab138881) provides an ultrasensitive assay to quantitate glutathione in mammalian samples.


    Note: We recommend using the alternate kit, GSH/GSSG Ratio Detection Assay Kit II (Fluorometric - Green) (ab205811), instead of this kit. ab205811 is a reformulated version of this kit with similar performance. It uses a water-soluble probe, which is more stable, than the DMSO-soluble dye used in this kit. 


     


    The GSH/GSSG assay protocol uses a proprietary non-fluorescent dye that becomes strongly fluorescent upon reacting with GSH. With a one-step fluorimetric method, the assay can detect as little as 1 picomole of GSH or GSSG in a 100 µL assay volume.


    The assay can be performed in a convenient 96-well or 384-well microtiter-plate format and readily adapted to automation without a separation step. Its signal can be easily read by a fluorescence microplate reader at Ex/Em = 490/520 nm.


    GSH/GSSG assay protocol summary:
    - add samples (deproteinized) and standards to wells
    - for GSH assay add Thiol Green in assay buffer, or for total glutathione (GSH + GSSG) assay also add GSSG probe
    - incubate for 10 - 60 min at room temp
    - analyze with microplate reader
    GSSG levels can be calculated by subtracting GSH from total glutathione levels.

  • Notes

    NOTE: For measuring GSH Standard only, there is enough reagent provided to perform 200 tests.

    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).

    Glutathione reductase (GR) recycles GSSG to GSH with the simultaneous oxidation of β-nicotinamide adenine dinucleotide phosphate (β-NADPH2).

    In healthy cells, >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.

  • Platform

    Microplate reader

Properties

  • Storage instructions

    Store at -20°C. Please refer to protocols.
  • Components 100 tests
    Assay Buffer 1 x 25ml
    DMSO 1 x 400µl
    GSH Standard 1 x 62µg
    GSSG Probe 1 vial
    GSSG Standard 1 x 124µg
    Thiol Green Indicator 1 vial
  • 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)

Images

  • Functional Studies - GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green) (ab138881)
    Functional Studies - GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green) (ab138881)Biesemann, Nadine et al., Scientific reports?vol. 8,1 9408., Fig 4, doi:10.1038/s41598-018-27614-8

    Measurement of reduced (GSH) and oxidized (GSSG) forms of glutathione in myotubes treated with 150 µM TBHP for 1 h (n = 6).

  • Functional Studies - GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green) (ab138881)
    Functional Studies - GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green) (ab138881)

    GSH in reduced state measured in cell lysates showing quantity (umol) per 1 mln cells.

    Samples with the concentration of 1e7-1e8 cells/mL were used. Samples were diluted 10-1000 fold.

  • Functional Studies - GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green) (ab138881)
    Functional Studies - GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green) (ab138881)

    Total GSH measured in cell lysates showing quantity (umol) per 1 mln cells.

    Samples with the concentration of 1e7-1e8 cells/mL were used. Samples were diluted 10-1000 fold.

  • Functional Studies - GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green) (ab138881)
    Functional Studies - GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green) (ab138881)

    GSH in reduced state measured in tissue lysates showing quantity (umol) per miligram of extracted protein of tested sample.

    Protein concentration for samples varied from 6 mg/mL to 16 mg/mL. Samples were diluted 10-100 fold.

  • Functional Studies - GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green) (ab138881)
    Functional Studies - GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green) (ab138881)

    Total GSH measured in tissue lysates showing quantity (umol) per miligram of extracted protein of tested sample.

    Protein concentration for samples varied from 6 mg/mL to 16 mg/mL. Samples were diluted 10-100 fold.

  • Functional Studies - GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green) (ab138881)
    Functional Studies - GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green) (ab138881)

    GSH in reduced state measured in biological fluids showing concentration (uM) in tested samples. Human samples were diluted 10 fold. Rat sample was diluted 10-1000 fold.

  • Functional Studies - GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green) (ab138881)
    Functional Studies - GSH/GSSG Ratio Detection Assay Kit (Fluorometric - Green) (ab138881)

    Total GSH measured in biological fluids showing concentration (uM) in tested samples. Samples were diluted 10-100 fold. 

  • Sample Calibration Curve - Total GSH
    Sample Calibration Curve - Total GSH
    Total GSH dose responses were measured with ab138881 in a black 96-well plate using a fluorescence microplate reader. 50 µl of GSSG standards (0.01 to 5 µM), GSH-containing samples or blank control were added into each well, and then 50 µl of Total GSH Reaction Mixture was added. Fluorescence intensity was measured at Ex/Em = 490/520 nm after 30 minutes incubation.
  • Sample Calibration Curve - Reduced GSH
    Sample Calibration Curve - Reduced GSH
    Reduced GSH dose responses were measured in a black 96-well plate with ab138881 using a fluorescence microplate reader. 50 µl of GSH standards (0.01 to 5 µM) or blank control was added into each well, and then 50 µl of GSH Assay Mixture was added. The fluorescence intensity was measured at Ex/Em = 490/520 nm after 30 minutes incubation.

Protocols

  • Protocol Booklet

Click here to view the general protocols

Datasheets and documents

  • SDS download

  • Datasheet download

    Download

References (109)

Publishing research using ab138881? Please let us know so that we can cite the reference in this datasheet.

ab138881 has been referenced in 109 publications.

  • Lin Y  et al. Astragaloside IV Ameliorates Streptozotocin Induced Pancreatic ß-Cell Apoptosis and Dysfunction Through SIRT1/P53 and Akt/GSK3ß/Nrf2 Signaling Pathways. Diabetes Metab Syndr Obes 15:131-140 (2022). PubMed: 35046684
  • Llewellyn SV  et al. Deducing the cellular mechanisms associated with the potential genotoxic impact of gold and silver engineered nanoparticles upon different lung epithelial cell lines in vitro. Nanotoxicology 16:52-72 (2022). PubMed: 35085458
  • Wu J  et al. TNF antagonist sensitizes synovial fibroblasts to ferroptotic cell death in collagen-induced arthritis mouse models. Nat Commun 13:676 (2022). PubMed: 35115492
  • Kolnik S  et al. Vitamin E Decreases Cytotoxicity and Mitigates Inflammatory and Oxidative Stress Responses in a Ferret Organotypic Brain Slice Model of Neonatal Hypoxia-Ischemia. Dev Neurosci 44:233-245 (2022). PubMed: 35134797
  • Wang J  et al. Mitochondrial dysfunction and oxidative stress contribute to cognitive and motor impairment in FOXP1 syndrome. Proc Natl Acad Sci U S A 119:N/A (2022). PubMed: 35165191
View all Publications for this product

Customer reviews and Q&As

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1-10 of 15 Abreviews or Q&A

Detection of GSH levels in microglia BV-2 cells

Good Good 4/5 (Ease of Use)
Abreviews
Abreviews
abreview image
I used GSH/GSSG ratio detection assay kit to measure GSH levels in the BV-2 cells treated with LPS and astaxanthin. Standard curve was perfect to examine total GSH levels and GSH/GSSG ratio. We used 1 million cells for each samples that were appropriate to detect GSH levels. The handbook was great to perfome this assay and calculate our results.
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.

Ms. Ji Hye Han

Verified customer

Submitted Apr 06 2018

Detection of GSH/GSSG ratio in plants

Good Good 4/5 (Ease of Use)
Abreviews
Abreviews
abreview image
I used this kit to measure the GSH/GSSG ratio in plants after treatment with high light. For this, 6-week- old Arabidopsis thaliana (Col) plants were treated with high light (1000 µmol m-2 sec-1) for 0, 2 and 15 minutes and immediately frozen in liquid nitrogen. Three plants were used for each time point. Each sample was then grinded in liquid nitrogen and 20mg of frozen powder was taken for the assay. I also purchased the mammalian lysis buffer from abcam. 400µl of the lysis buffer was immediately added to the sample after weighing. The rest of the assay and calculations were done according to the protocol provided.
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.

Feroza Kaneez

Verified customer

Submitted Jan 24 2018

Detection of GSH levels in wild ype and cystathionine gamma lyase (CSE) silenced cells

Good Good 4/5 (Ease of Use)
Abreviews
Abreviews
abreview image
We measured GSH levels in OVCAR4 cells tranfected with scrambled siRNA or CSE siRNA. We used Cell Lytic M lysis buffer for lysis followed by deproteinisation. The assay was straightforward and the standards gave perfect plots for quantifying amounts of GSH in our samples. We used I million cells for each sample.
Scope of improvement: Providing an assay compatible lysis buffer in the kit will be appreciable.
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.

Abcam user community

Verified customer

Submitted Dec 12 2017

Detection of GSH levels in equine fibroblasts

Average Average 3/5 (Ease of Use)
Abreviews
Abreviews
abreview image
I wanted to measure GSH levels in equine fibroblasts and decided to try the abcam kit. Upon arrival the kit required aliquoting and reconstituting but once this was done it was easy to set up the assay. Some reagents aren't included in the kit (cell lysis buffer and deproteinisation kit) but these are easily ordered from abcam as per the protocol.
This assay worked on equine cells and gave similar values as per other cell types provided in the handbook. The kit did however give similar readings for different sample dilutions (before calculations) and therefore raised some concern regarding sample preparation (I used the mammalian cell lysis buffer and deproteinisation kit from abcam).
The calculations are quite straight forward but not explained particularly well in the handbook, this would be clarified using a working example perhaps.
Figure legend:
Total GSH and reduced GSH levels (uM per 1million cells) in equine fibroblast lysates. Measurements taken at 1/100 dilution of samples. Mean +/- SEM, samples were run in duplicate. Samples were cultured in different glucose concentrations (- :5.5mM, + :25mM).
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.

Abcam user community

Verified customer

Submitted Jun 09 2016

Question

Can we use Triton X-100 as lysis buffer?

Read More

Abcam community

Verified customer

Asked on May 23 2014

Answer

Our in house experiments showed increase in background with Triton X-100 which is why we recommend auto-fluorescent free Triton-X 100 or simply avoiding this lysis buffer.

We recommend following sample preparation protocol:
GSH is labile and easily oxidized, so all samples and reagents should be stored ice cold and used as rapidly as possible. For Cell Lysate preparation: Wash cells with PBS. Resuspend 2-4 x 106 cells in 500 μL of ice-cold 5% Metaphosphoric acid (MPA) working solution. Homogenize cell suspension. Centrifuge homogenate at 4°C for 10 minutes Collect the upper clear aqueous layer and keep at 0-4°C for the assay (within 1 hour). Store on ice if used immediately or freeze at -80ºC for future use. Please neutralize the MPA to pH 4˜6 and then analyse the sample with the kit. Samples may need to dilute 10˜40 folds depending on GSH amount.

Read More

Padamjeet Singh

Abcam Scientific Support

Answered on May 23 2014

Question

I would like some information about ab138881 (GSH/GSSG quantification).:

which lysis solution should be used ?
Is a deproteination step required ?
Which cell concentration should be used?

Read More

Abcam community

Verified customer

Asked on May 21 2014

Answer

For lysis, you can use 0.5% NP40 made up in PBS pH6.0 to lyse your cells. Please do not use the assay buffer to lyse your cells as it contains no detergent.

The deproteinization step is optional, however, we do find that it reduces interference and background in most samples.

With regards to the number of cells to use, this will depend largely on your cells and how much GSH/GSSG is present in your samples. Samples should ideally fall within the middle of standard curve so you may need to try a series of dilutions to determine the optimal conditions to use. As a starting point, the range of cell concentrations that we would recommend would be 10,000 to 100,000 cells/ well/100 uL. Try starting with ˜10,000 to 20,000 cells with adherent cells, and 50,000 to 100,000 cells with suspension cells.

Read More

Elisa Thomas

Abcam Scientific Support

Answered on May 21 2014

Question

Can the kit be used on whole blood?

Read More

Abcam community

Verified customer

Asked on Mar 22 2013

Answer

No direct experiences on total blood but should be OK theoretically. The following may help:

1. add equal vol of ice-cold 10% Metaphosphoric acid (MPA) working solution. mix well, let sit at RT for 5 min.
2. Centrifuge at >2,000xg at 4°C for 5 min.
3. Collect the upper clear aqueous layer and keep at 0-4°C for the assay (within 1 hour).
4. Store on ice if used immediately or freeze at -80ºC for future use.
5. Neutralize the MPA to pH 4~6 and then analyze the sample with the kit. Samples may need to dilute 10~40 folds depending on GSH amount

Read More

Abcam Scientific Support

Answered on Mar 22 2013

Question

Inquiry: Can I lyse the samples with the provided assay buffer or do I have to use another lysis buffer?

Read More

Abcam community

Verified customer

Asked on Feb 05 2013

Answer

We recommend to use 0.5% NP40 as cell lysis buffer. The Assay buffer does not contain any detergent and therefore it is not suitable to be used for lysing the cells.

Read More

Abcam Scientific Support

Answered on Feb 05 2013

Question

All of the standards and samples are giving me the same results. Do you know what could be causing this?

Read More

Abcam community

Verified customer

Asked on Jan 31 2013

Answer

The GSH is very easy to be oxidized. If GSH standard signal is lower than GSSG (or GAM is higher than total GAM), the GSH may be oxidized.

Read More

Abcam Scientific Support

Answered on Jan 31 2013

Question

Can I use a lysis buffer that contains DTT or Beta Mercaptoethanol?

Read More

Abcam community

Verified customer

Asked on Nov 22 2012

Answer

DTT or 2-mercaptoethanol cannot be used for this case. In our kit, GSSG probe is an enzyme-based mixture with other additives that can convert GSSG to GSH, which can be recognized by the probe and generate green fluorescence after reaction. DTT or 2-mercaptoethanol will interfere with the probe, and they should be exclude from the reaction system.

Read More

Abcam Scientific Support

Answered on Nov 22 2012

1-10 of 15 Abreviews or Q&A

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