Glutathione Peroxidase 4 (GPX4) Activity Assay Kit
Glutathione Peroxidase 4 (GPX4) Activity Assay Kit
| SKU # | E-BC-K883-M |
| Detection Instrument | Microplate reader (340 nm) |
|
Detection method |
Colorimetric method |
Product Details
Properties
| Synonyms | GPX4 |
| Sample type | Animal tissue and cell |
| Sensitivity | 3.22U/L |
| Detection range | 3.22-44.69U/L |
| Detection method | Colorimetric method |
| Assay type | Enzyme Activity |
| Precision | Average inter-assay CV: 0.4-3.4% | Average intra-assay CV: 1.2-.3.1% |
| Other instruments required | Incubator |
| Other reagents required | Double distilled water, Normal saline(0.9% NaCl) |
| Storage | -20℃ |
| Valid period | 12 months |
Dilution of Sample
The recommended dilution factor for different samples is as follows (for reference only):
| Sample type | Dilution factor |
| 10% Mouse liver tissue homogenate | 5-50 |
| 10% Mouse kidney tissue homogenate | 2-6 |
| 10% Mouse lung tissue homogenate | 2-5 |
| 2.05×106 293T cells | 1 |
| 1.9×106 4T1 cells | 1 |
| 2.03×106 Jurkat cells | 1 |
| 2×106 Molt-4 cells | 1 |
Detection Principle
Glutathione peroxidase 4(GPX4) is a kind of glutathione peroxidase containing selenocysteine, which can reduce phospholipid hydroperoxide to phosphatidyl alcohol, thus protecting the cell membrane from oxidative damage. GPX4 catalyzes the substrate, and the product consumes the reducing agent with the addition of enzyme reagents. The reducing agent has a maximum absorbance at 340 nm, and the GP4-specific enzyme activity can be calculated by measuring non-specific enzyme activity and total enzyme activity by adding GPX-4 inhibitor to the system.
Kit Components & Storage
| Item | Component | Size 1(48 T) | Size 2(96 T) | Storage |
| Reagent 1 | Buffer Solution | 25 mL × 1 vial | 50 mL × 1 vial | -20℃, 12 months |
| Reagent 2 | Substrate | Powder × 1 vial | Powder × 2 vials | -20℃, 12 months, shading light |
| Reagent 3 | Enzyme Reagent | 0.25 mL ×1 vial | 0.5 mL × 1 vial | -20℃, 12 months, shading light |
| Reagent 4 | Oxidant | 0.15 mL ×1 vial | 0.3 mL × 1 vial | -20℃, 12 months, shading light |
| Reagent 5 | Inhibitor | 0.25 mL ×1 vial | 0.5 mL × 1 vial | -20℃, 12 months, shading light |
| Reagent 6 | Reducing Reagent | Powder × 1 vial | Powder × 2 vials | -20℃, 12 months, shading light |
| Reagent 7 | Accelerant | 4 mL ×1 vial | 8 mL × 1 vial | -20℃, 12 months, shading light |
| Reagent 8 | Stabilizer | 0.5 mL ×1 vial | 1 mL × 1 vial | -20℃, 12 months, shading light |
| UV Microplate | 96 wells | No requirement | ||
| Plate Sealer | 2 pieces | |||
Note: The reagents must be stored strictly according to the preservation conditions in the above table. The reagents in different kits cannot be mixed with each other. For a small volume of reagents, please centrifuge before use, so as not to obtain sufficient amount of reagents.
Technical Data:
Parameter:
Intra-assay Precision
Three mouse liver samples were assayed in replicates of 20 to determine precision within an assay (CV = Coefficient of Variation).
| Parameters | Sample 1 | Sample 2 | Sample 3 |
| Mean (U/L) | 6.80 | 16.20 | 32.50 |
| %CV | 3.1 | 1.4 | 1.2 |
Inter-assay Precision
Three mouse liver samples were assayed 20 times in duplicate by three operators to determine precision between assays.
| Parameters | Sample 1 | Sample 2 | Sample 3 |
| Mean (U/L) | 6.80 | 16.20 | 32.50 |
| %CV | 3.4 | 2.2 | 0.4 |
Recovery
Take three samples of high concentration, middle concentration and low concentration to test the samples of each concentration for 6 times parallelly to get the average recovery rate of 98%.
| Sample 1 | Sample 2 | Sample 3 | |
| Expected Conc. (U/L) | 8.5 | 19 | 37 |
| Observed Conc. (U/L) | 8.5 | 18.3 | 36.1 |
| Recovery rate(%) | 100 | 96.3 | 97.7 |
Sensitivity
The analytical sensitivity of the assay is 3.22 U/L. This was determined by adding two standard deviations to the mean O.D. obtained when the zero standard was assayed 20 times, and calculating the corresponding concentration.