Total Price: $0.00
Contact Information
Vitamin C (VC) Colorimetric Assay Kit
SKU: E-BC-K034-M-500
Vitamin C (VC) Colorimetric Assay Kit
| SKU # | E-BC-K034-M |
| Detection Instrument | Microplate reader (530-540 nm, optimum wavelength: 536 nm) |
| Detection Method | Colorimetric method |
Product Details
Properties
| Synonyms | VC, Ascorbic acid |
| Sample Type | Serum, plasma, tissue |
| Sensitivity | 0.31 μg/mL |
| Detection Range | 0.31-20 μg/mL |
| Detection Method | Colorimetric method |
| Assay type | Quantitative |
| Assay time | 50 min |
| Precision | Average inter-assay CV: 6.100% | Average intra-assay CV: 2.200% |
| Other instruments required | Micropipettor, Centrifuge, Incubator, Vortex mixer |
| Other reagents required | Normal saline (0.9% NaCl), PBS (0.01 M, pH 7.4), Absolute ethanol |
| 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 (0.31-20 μg/mL).
The recommended dilution factor for different samples is as follows (for reference only):
| Sample type | Dilution factor |
| Human serum | 1 |
| Mouse serum | 1 |
| Chicken serum | 1 |
| Horse serum | 1 |
| 10% Mouse kidney tissue homogenate | 1 |
| 10% Mouse liver tissue homogenate | 1 |
| 10% Mouse lung tissue homogenate | 1 |
| 10% Mouse spleen tissue homogenate | 1 |
| 10% Rat liver tissue homogenate | 1 |
| 10% Rat heart tissue homogenate | 1 |
| 10% Plant tissue homogenate | 1 |
Note: The diluent is normal saline (0.9% NaCl) or PBS (0.01 M, pH 7.4).
Detection Principle
The most obvious chemical activity of VC is that reduce Fe3+ to Fe2+, then promote iron absorption in the intestine, promote the storage and utilization of iron. Fe3+ react immediately with reducing ascorbic acid to form Fe2+. then Fe2+ react with phenanthroline and the color developing reaction occurs. The content of vitamin C in sample can be determined. Measure the OD value and calculate the VC content indirectly.
Kit Components & Storage
| Item | Component | Size 1(48 T) | Size 2(96 T) | Storage |
| Reagent 1 | Extracting Solution | 2 mL × 1 vial | 5 mL × 1 vial | 2-8℃, 12 months shading light |
| Reagent 2 | Buffer Solution | 8 mL × 1 vial | 15 mL × 1 vial | 2-8℃, 12 months |
| Reagent 3 | Chromogenic Agent | 3 mL × 1 vial | 6 mL × 1 vial | 2-8℃, 12 months shading light |
| Reagent 4 | Ferrum Reagent | 0.5 mL × 1 vial | 0.5 mL × 1 vial | 2-8℃, 12 months shading light |
| Reagent 5 | Stop Solution | 6 mL × 1 vial | 12 mL × 1 vial | 2-8℃, 12 months |
| Reagent 6 | VC Standard | 6 mg × 2 vials | 6 mg × 2 vials | 2-8℃, 12 months shading light |
| 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 (μg/mL) | 1.50 | 8.40 | 12.50 |
| %CV | 2.7 | 2.1 | 1.8 |
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 (μg/mL) | 1.50 | 8.40 | 12.50 |
| %CV | 5.7 | 6.4 | 6.1 |
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 108%.
| Standard 1 | Standard 2 | Standard 3 | |
| Expected Conc. (μg/mL) | 6.5 | 11 | 13.5 |
| Observed Conc. (μg/mL) | 7.2 | 11.6 | 14.7 |
| Recovery rate (%) | 110 | 105 | 109 |
Sensitivity
The analytical sensitivity of the assay is 0.31 μg/mL VC. 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 (μg/mL) | 0 | 5 | 7.5 | 10 | 12.5 | 15 | 17.5 |
| Average OD | 0.054 | 0.152 | 0.193 | 0.231 | 0.278 | 0.312 | 0.358 |
| Absoluted OD | 0 | 0.098 | 0.139 | 0.177 | 0.224 | 0.258 | 0.304 |