Glutathione, γ-L-glutamyl-L-cysteinyl-glycine or GSH, is the most abundant non-protein thiol in cells. GSH is involved in the regulation of a number of essential biochemical processes within the cell. Primarily recognized as a key intracellular source of reducing power for combating the toxic accumulation of free radical byproducts, GSH is also involved with detoxifying and removal of exo/endogenous toxins and alkylating agents. In its role as a cell signaling agent, GSH is involved in DNA synthesis and cell proliferation regulation. Due to the genetically conserved molecular processes by which cells die, intracellular levels of GSH can favor a cell death pathway via apoptosis (adequate intracellular GSH stores) or via necrosis or autophagy (depleted intracellular GSH stores). Detection of a drop in intracellular GSH concentration in an experimental cell population relative to a negative (healthy cell) control is often indicative of an apoptosis induction event. ICT’s Glutathione Colorimetric Detection Kit is designed to quantitatively measure GSH and GSSG in a variety of samples.
Glutathione Colorimetric Detection Kit
Quantitatively measure glutathione (GSH) and oxidized glutathione (GSSG) present in a variety of samples, including whole blood, serum, plasma, erythrocytes, urine, cell lysates, and tissue samples. Analyze the absorbance colorimetric signal by spectrophotometer.
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1. Prepare samples; treat with SSA to remove thiol-bearing protein.
2. Prepare standard curve and blanks.
3. If measuring oxidized GSH (GSSG), treat blanks, samples, and standards with 2VP.
4. Add 50 µL blanks, standards, and samples to plate.
5. Add 25 µL Colorimetric GSH Detection Substrate.
6. Add 25 µL Reaction Mixture.
8. Incubate 20 minutes at room temperature.
9. Read at 405 nm.
Absorbance of Oxidized Glutathione Standard With and Without Treatment of 2-VP. The standard curve derived from the 2-VP treatment, which covalently blocks Free GSH, is used to calculate Oxidized Glutathione. The standard curve derived from the untreated standard (without 2-VP) is used to calculate Total Glutathione.
• Target: Glutathione and Oxidized Glutathione
• Method of Analysis: Absorbance Plate Reader
• Types of Samples: whole blood, serum, plasma, erythrocytes, urine, cell lysates, and tissue samples
• Storage: 2-8°C
• Shipping: Ships overnight (domestic), International Priority Shipping
• 4 clear half-area polystyrene 96-well microtiter plates, #267
• Oxidized Glutathione (GSSG) Standard, 250 µM, 350 µL, #6624
• Colorimetric GSH Detection Substrate, 1 mL, #6625
• Glutathione Assay Buffer, 225 mL, #6626
• NADPH Concentrate, 1 mL, #6627
• Glutathione Reductase Concentrate, 1 mL, #6622
• Kit Manual