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#DIGT-250 QuantiChrom™ Glutathione Assay Kit
Product name : QuantiChrom™ Glutathione Assay Kit
Catalog number : DIGT-250
Supplier name : Bioassays
Data sheet: Ask more or other datasheet now !
About this Product :QuantiChrom™ Glutathione Assay Kit antibody storage GENTAUR recommends for long therm storage to freeze at -24 C. For short time storage up to 30 days we suggest fridge storage at 1 to 10 C. Prevent multiple freeze taw cycles of QuantiChrom™ Glutathione Assay Kit.
More Details about
Glutathione is a tripeptide of glycine, glutamic acid and cysteine. In the
red blood cell, the reduced form of glutathione is vital in maintaining
hemoglobin in a reduced state and hence protecting the cells from
oxidative damage. Glutathione is involved in detoxification of hydrogen
peroxide through glutathione oxidase. Low levels of glutathione are found
in deficiencies of key enzymes involved in glutathione metabolism, such
as glucose-6-phosphate dehydrogenase, glutathione synthase and
Simple, direct and automation-ready procedures for measuring reduced
glutathione are becoming popular in Research and Drug Discovery.
BioAssay Systems' QuantiChromTM Glutathione Assay Kit is designed to
accurately measure reduced glutathione in biological samples. The
improved 5,5'-dithiobis(2-nitrobenzoic acid (DTNB) method combines
deproteination and detection (Reagent A) into one reagent. DTNB reacts
with reduced glutathione to form a yellow product. The optical density,
measured at 412 nm, is directly proportional to glutathione concentration
in the sample. The optimized formulation has a long shelf life and is
completely free of interference due to sample turbidity.
Sensitive and accurate. Linear detection range 0.4 - 100 μM in 96-well plate.
Simple and convenient. The procedure involves mixing the DTNB
Reagent with sample, removing protein precipitates for proteinaceous
samples, adding a second Reagent and reading the optical density.
Low interference. Amino acids and common buffers do not interfere.
Direct Assays: reduced glutathione in whole blood, plasma, serum,
urine, tissue and cell extracts.
Drug Discovery/Pharmacology: effects of drugs on glutathione
KIT CONTENTS (250 tests in 96-well plates)
Reagent A: 30 mL Reagent B: 30 mL
Calibrator: 10 mL (equivalent to 100 μM glutathione)
Storage conditions. The kit is shipped at room temperature. Store all
components at 4°C. Shelf life: 12 months after receipt.
Precautions: reagents are for research use only. Normal precautions for
laboratory reagents should be exercised while using the reagents.
Please refer to Material Safety Data Sheet for detailed information.
Important: equilibrate Reagents to room temperature. Shake Reagent A
Samples: whole blood samples should be diluted 20-fold with water prior
to the assay (n = 20). Cell lysate can be prepared as follows: collect 2 x
106 cells by centrifugation at 1,000g for 10 min at 4°C. Wash cells in cold
PBS. Lyse cells by homogenization or sonication in 1-2 mL of cold buffer
containing 50 mM MES or phosphate (pH 6-7) and 1 mM EDTA.
Centrifuge at 10,000g for 15 min at 4°C. Use supernatant for assay.
Note: b-mercaptoethanol, dithiothreitol and cysteine are known to
interfere in this assay. Avoid using these compounds during sample
preparation. Amino acids do not interfere.
Procedure using 96-well plate:
1. Blank and Calibrator. Transfer 100 μL water and 100 μL Calibrator into
wells of a clear-bottom 96-well plate. Pipette 200 μL water into the
Blank and Calibrator wells.
2. Samples. Whole blood samples should be diluted 20-fold with water
prior to the assay (n = 20). Deproteination is required for blood, serum,
plasma and other proteinaceous samples. Reagent A contains
components for both color reaction and deproteination.
Mix 120 μL sample with 120 μL Reagent A in 1.5-mL centrifuge
tubes. Vortex to mix well. If turbidity occurs, pellet 5 min at 14,000
rpm in a table centrifuge. If the mixture remains clear, no
centrifugation is necessary.
3. Transfer 200 μL sample/Reagent A mixture into wells of the 96-
well plate. Add 100 μL Reagent B. Tap plate lightly to mix.
4. Incubate 25 min at room temperature. Read OD412nm.
Procedure using Cuvet:
Mix 400 μL sample with 400 μL Reagent A, centrifuge sample tubes
if precipitation occurs. Transfer 600 μL supernatant and mix with
400 μL Reagent B. Incubate 25 min at room temperature. Measure
OD412nm against water. Transfer 400 μL Calibrator and 800 μL Water
into a clean cuvet and measure OD412nm against water.
Subtract blank OD (water) from the Calibrator and Sample OD
values. The glutathione concentration of Sample is calculated as
ODSAMPLE, ODSTD and ODBLANK are optical density values of the
sample, Calibrator and sample “Blank” (water or buffer in which the
sample was dissolved). n is the dilution factor (20 for blood
Conversions: 1 mg/dL glutathione equals 32.5 μM, 0.001% or 10 ppm.
MATERIALS REQUIRED, BUT NOT PROVIDED
Pipeting devices, centrifuge tube and table centrifuge.
Procedure using 96-well plate:
Clear bottom 96-well plates (e.g. Corning Costar) and plate reader.
Procedure using cuvette:
Spectrophotometer and cuvets for measuring OD at 412 nm.
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