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| Product Name | Glutathione Reductase (GR) Activity Colorimetric Assay Kit |
| Catalog No. | ETR-HMM-0082 |
| Description | Colorimetric assay for glutathione reductase activity based on the reduction of oxidized glutathione (GSSG) to reduced glutathione (GSH) with concomitant oxidation of NADPH to NADP+, monitored as a decrease in absorbance at 340 nm. GR is essential for maintaining cellular GSH levels and redox homeostasis, with implications in cancer drug resistance, aging, and oxidative stress-related diseases. |
| Intended Use | Quantitative measurement of glutathione reductase activity in cell and tissue lysates, erythrocytes, and purified enzyme preparations; oxidative stress research; cancer drug resistance mechanism studies; evaluation of antioxidant defense systems. |
| Principle / Technology | GR + NADPH + H⁺ + GSSG → GR + NADP⁺ + 2 GSH; oxidation of NADPH is monitored as a decrease in A340 (ε340 = 6.22 mM⁻¹cm⁻¹); reaction rate is proportional to GR activity |
| Detection Method | Colorimetric; kinetic mode at 340 nm; absorbance decrease measured every 30 sec for 5-10 min at 25 °C or 37 °C |
| Sample Type | Cell/tissue lysates (10-100 µg protein), erythrocyte lysates, purified GR; 10-50 µL sample per 200 µL assay |
| Sensitivity / LOD | LOD 0.1 mU/mL GR activity (1 mU = 1 nmol NADPH oxidized/min); linear range 0.1-50 mU/mL |
| Specificity | Specific for NADPH-dependent glutathione reductase; no cross-reactivity with thioredoxin reductase due to GSSG substrate specificity; NADH does not substitute for NADPH efficiently |
| Reaction Conditions / Protocol | Add sample to assay buffer containing GSSG (1 mM) and NADPH (0.2 mM); monitor A340 decrease kinetically; calculate activity from initial linear rate using NADPH extinction coefficient |
| Components / Formulation | Assay buffer (potassium phosphate 100 mM pH 7.5 with EDTA 1 mM), GSSG substrate (50 mM), NADPH (20 mM in Tris buffer), GR positive control (purified yeast GR, 1 U/mL), 96-well UV-transparent plate (optional) |
| Storage Conditions | -20 °C (NADPH and GSSG); assay buffer at 4 °C; GR control at -80 °C |
| Shelf Life | 12 months at -20 °C; NADPH solution 1 week at 4 °C, 2 months at -20 °C aliquoted |
| Package Specifications | 100 assays, 200 assays, 500 assays in 96-well format |
| Product Form | Lyophilized and liquid reagents; UV-transparent plate optional |
| Key Features | Direct kinetic measurement at 340 nm; NADPH extinction coefficient provides absolute quantification; includes purified GR positive control; optimized substrate concentrations for standard kinetic analysis; compatible with 96-well UV plate readers |
| Purity | NADPH ≥97% by enzymatic assay; GSSG ≥98%; GR control specific activity >200 U/mg |
| Concentration | As specified per kit; calibrator and substrate concentrations traceable to reference |
| Activity / Unit Definition | One unit defined as the amount of enzyme that catalyzes conversion of 1 µmol substrate per minute under specified conditions |
| Molecular Weight | Varies by target enzyme as documented |
| Source / Origin | Recombinant or purified from natural sources; synthetic chromogenic/fluorogenic substrates |
| pH Range / Optimal pH | pH 6.0–9.0 depending on specific enzyme assay |
| Shipping Conditions | Cold pack -20 °C; NADPH light-sensitive — ship in amber vials |
| Expiration Date / Stability | 12 months at -20 °C; NADPH stock solution 2 months at -20 °C; discard NADPH if A340/A260 ratio decreases below 0.35 (indicating oxidation to NADP+) |
| Regulatory / Compliance | For laboratory and research use only; RUO; ISO 9001 |
| Compatibility | Compatible with standard microplate readers (absorbance, fluorescence, luminescence modes) |
| Recommended Buffer System | Tris or phosphate buffer optimized per enzyme assay |
| Application Notes / Precautions | Include positive control and inhibitor control in each assay; read within linear range; protect light-sensitive substrates |
| Batch-to-Batch Consistency | Enzyme activity and standard curve linearity within ±15% of reference lot |
For research use only, not for clinical use.
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