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

Note: The diluent is normal saline (0.9% NaCl). For the dilution of other sample types, please do pretest to confirm the dilution factor.

 

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.