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This product is available through the Promega Helix onsite stocking program in a –20°C Helix Freezer. The program offers numerous convenient solutions to meet your lab's needs. Helix Freezers are available in two sizes: 5.7 cubic ft. or 9.7 cubic ft.
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The GSH-Glo™ Assay is a luminescent-based assay for the detection and quantification of glutathione (GSH) in cells or in various biological samples. A change in GSH levels is important in the assessment of toxicological responses and is an indicator of oxidative stress, potentially leading to apoptosis or cell death. The assay is based on the conversion of a luciferin derivative into luciferin in the presence of GSH. The reaction is catalyzed by a glutathione S-transferase (GST) enzyme supplied in the kit. The luciferin formed is detected in a coupled reaction using Ultra-Glo™ Recombinant Luciferase that generates a glow type luminescence that is proportional to the amount of glutathione present in cells. The assay provides a simple, fast and sensitive alternative to colorimetric and fluorescent methods and can be adapted easily to high-throughput applications.
The assay is performed in two steps. In the 1st step, cells are lysed in the presence of the luciferin-NT substrate and glutathione S-transferase. Glutathione in the cells drives the formation of luciferin. In the 2nd step, Luciferin Detection Reagent is added to produce light that is directly proportional to the amount of GSH in the reaction.
HeLa cells (5000 cells/well in 96 well format) were exposed to l-Buthionine-sulfoximine (BSO). BSO inhibits GSH synthesis thus reducing cellular GSH level.
20hr BSO: Cells treated with BSO and assayed for GSH after 20 hours.
BSO + Recovery: Cells treated with BSO for 20 hours then washed 2X with PBS and covered with fresh media without BSO. Cells assayed for GSH after 40 hours.
40hr BSO: Cells treated with BSO, washed at 20 hours and fresh media plus BSO added. Cells assayed for GSH after 40 hours.
The simple two-reagent-addition assay minimizes the number of assay steps compared to conventional GSH assays and is adapted easily to higher throughput applications. No deproteination step is required, and results are achieved in as quickly as 30 minutes.
The luminescent method avoids inherent background fluorescence associated with other methods thereby providing excellent signal-to-background ratios. A half-life greater than 5 hours results in a stable signal as well.
GSH levels in lysates from adherent cells in 24 well plates were measured with the GSH-Glo Glutathione Assay. GSH concentrations were determined by interpolation from a GSH standard curve generated using the bioluminescent system. Exposure to 5mM acetaminophen for three hours substantially reduced GSH only after treatment for two days with a P450 inducer, 30 μM pregnenolone 16a-carbonitrile (PCN).
GSH-Glo™ Glutathione Assay Technical Bulletin
(1 MB pdf)
Reconstitution Buffer with esterase
GSH-Glo™ Reaction Buffer
Luciferin Detection Reagent
PDF 0 B
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U.S. Pat. Nos. 6,602,677, 7,241,584, 8,030,017 and 8,822,170 and other patents and patents pending.
Certain applications of this product may require licenses from others.
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