Sucrase Activity Assay Kit

$ 272⁹⁵

Sucrase Activity Assay Kit

SKU #	E-BC-K751-M
Detection Instrument	Microplate reader (500-520 nm, optimum wavelength: 505 nm)
Detection method	Colorimetric method

Product Details

Properties

Sample type	Animal tissue
Sensitivity	20 U/mL
Detection range	20-2000 U/mL
Detection method	Colorimetric method
Assay type	Enzyme Activity
Assay time	70 min
Precision	Average inter-assay CV: 6.500% \| Average intra-assay CV: 5.400%
Other instruments required	Micropipette, Vortex mixer, Centrifuge
Other reagents required	PBS (0.01 M, pH 7.4)
Storage	2-8℃
Valid period	12 months

Dilution of Sample

It is recommended to take 2~3 samples with expected large difference to do pre-experiment before formal experiment and dilute the sample according to the result of the pre-experiment and the detection range (20-2000 U/mL).

The recommended dilution factor for different samples is as follows (for reference only):

Sample type	Dilution factor
20% Rat ileum tissue homogenate	1
20% Rat stomach tissue homogenate	1
20% Rat liver tissue homogenate	1

Note: The diluent is PBS (0.01 M, pH 7.4).

Detection Principle

Sucrase catalyzes its substrate (sucrose) to produce glucose, which produces hydrogen peroxide under the action of glucose oxidase. Hydrogen peroxide reacts with chromogenic agent to produce red substance, which has a strong absorption peak at 505 nm. In a certain concentration range, It's absorbance is proportional to glucose concentration. Therefore, the activity of sucrase can be calculated by measuring the OD value at 505 nm.

Kit Components & Storage

Item	Component	Size (96 T)	Storage
Reagent 1	Substrate	Powder × 1 vial	2-8℃, 12 months
Reagent 2	Buffer Solution	10 mL × 1 vial	2-8℃, 12 months
Reagent 3	Phenol Solution	12 mL × 1 vial	2-8℃, 12 months, shading light
Reagent 4	Enzyme Solution	12 mL × 1 vial	2-8℃, 12 months, shading light
Reagent 5	Stop Solution	Powder × 1 vial	2-8℃, 12 months
Reagent 6	50 mmol/L Glucose Standard	1 mL × 1 vial	2-8℃, 12 months
	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 rat liver tissue 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/mL)	32.50	135.00	482.00
%CV	5.7	5.3	5.2

Inter-assay Precision

Three rat liver tissue samples were assayed 20 times in duplicate by three operators to determine precision between assays.

Parameters	Sample 1	Sample 2	Sample 3
Mean (U/mL)	32.50	135.00	482.00
%CV	6.2	6.4	6.9

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 100%.

	Standard 1	Standard 2	Standard 3
Expected Conc. (mmol/L)	1.5	7.8	17.5
Observed Conc. (mmol/L)	1.5	8.0	16.8
Recovery rate (%)	101	103	96

Sensitivity

The analytical sensitivity of the assay is 20 U/mL. 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.

Standard Curve

As the OD value of the standard curve may vary according to the conditions of the actual assay performance (e.g. operator, pipetting technique or temperature effects), so the standard curve and data are provided as below for reference only:

Concentration (mmol/L)	0	1	2	5	10	15	20	25
Average OD	0.083	0.112	0.140	0.233	0.378	0.527	0.683	0.840
Absoluted OD	0.000	0.030	0.058	0.150	0.295	0.444	0.600	0.757