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Nitric Oxide (NO) Colorimetric Assay Kit– MSE Supplies LLC

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Nitric Oxide (NO) Colorimetric Assay Kit

SKU: E-BC-K035-M-96

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Nitric Oxide (NO) Colorimetric Assay Kit

SKU # E-BC-K035-M
Detection Instrument Microplate reader(540-550 nm)
Detection Method Colorimetric method


Product Details

Properties

Synonyms NO
Sample Type Serum, plasma, tissue, saliva
Sensitivity 0.16 μmol/L
Detection Range 0.16-100 μmol/L
Detection Method Colorimetric method
Assay type Quantitative
Assay time 55 min
Precision Average inter-assay CV: 3.700% | Average intra-assay CV: 2.400%
Other instruments required Micropipettor, Vortex mixer, Centrifuge
Other reagents required Normal saline (0.9% NaCl), PBS (0.01 M, pH 7.4)
Storage Reagent 6: -20℃, others: 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 (0.16-100 μmol/L).

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

Sample type Dilution factor
Human serum 1
Human plasma 1
10% Mouse liver tissue homogenization 1
Rat serum 1
Mouse serum 1
10% Epipremnum aureum tissue homogenization 1


Note: The diluent is double distilled water, normal saline (0.9% NaCl) or PBS (0.01 M, pH 7.4).

 

Detection Principle

NO is easily oxidized to form NO2- in vivo or in aqueous solution, and a reddish azo compound is formed with the color developing agent, and the concentration of the azo compound is linearly related to the concentration of NO. The concentration of NO can be calculated indirectly by measuring the OD value at 550 nm.


Kit Components & Storage

Item Component Size 1(48 T) Size 2(96 T) Storage
Reagent 1 Sulphate Solution 12 mL × 1 vial 24 mL × 1 vial 2-8℃, 12 months
Reagent 2 Alkali Reagent 12 mL × 1 vial 12 mL × 1 vial 2-8℃, 12 months
Reagent 3 Chromogenic
Agent A
1.9 mL × 1 vial 1.9 mL × 2 vials 2-8℃, 12 months
shading light
Reagent 4 Chromogenic
Agent B
Powder × 1 vial Powder × 1 vial 2-8℃, 12 months
shading light
Reagent 5 Acid Solution 1.3 mL × 1 vial 1.3 mL × 2 vials 2-8℃, 12 months
Reagent 6 Sodium Nitrite
Standard
Powder × 1 vial Powder × 2 vials 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 human serum 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 (μmol/L) 1.30 36.50 74.20
%CV 2.8 2.1 2.3


Inter-assay Precision

Three human serum samples were assayed 20 times in duplicate by three operators to determine precision between assays.

Parameters Sample 1 Sample 2 Sample 3
Mean (μmol/L) 1.30 36.50 74.20
%CV 3.5 3.8 3.8


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 102%.


Standard 1 Standard 2 Standard 3
Expected Conc. (μmol/L) 15 34 78
Observed Conc. (μmol/L) 15.2 35.4 78.8
Recovery rate (%) 101 104 101


Sensitivity

The analytical sensitivity of the assay is 0.16 μmol/L NO. 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 (μmol/L) 0 10 20 30 40 60 80 100
Average OD 0.036 0.065 0.097 0.117 0.162 0.223 0.287 0.351
Absoluted OD 0.000 0.028 0.061 0.081 0.125 0.187 0.251 0.314