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Phosphorus (Pi) Colorimetric Assay Kit (Phospho Molybdate Method)
SKU: E-BC-K245-M-96
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Phosphorus (Pi) Colorimetric Assay Kit (Phospho Molybdate Method)
| SKU # | E-BC-K245-M |
| Detection Instrument | Microplate reader (620 nm-690 nm, optimum wavelength: 660 nm) |
| Detection method | Colorimetric method |
Product Details
Properties
| Synonyms | Pi |
| Sample Type | Serum, plasma, tissue |
| Sensitivity | 0.004 mmol/L |
| Detection Range | 0.004-2.0 mmol/L |
| Detection Method | Colorimetric method |
| Assay type | Quantitative |
| Assay time | 40 min |
| Precision | Average inter-assay CV: 3% | Average intra-assay CV: 2.100% |
| Other instruments required | Micropipettor, Incubator, Vortex mixer, Centrifuge |
| Other reagents required | Normal saline (0.9% NaCl) |
| 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 (0.004-2.0 mmol/L).
The recommended dilution factor for different samples is as follows (for reference only):
| Sample type | Dilution factor |
| Human serum | 1 |
| Mouse serum | 1 |
| Rat plasma | 1 |
| Human urine | 2-3 |
| 10% Rat liver tissue homogenate | 1 |
| 10% Mouse lung tissue homogenate | 1 |
| 10% Mouse brain tissue homogenate | 1 |
| 10% Rat muscle tissue homogenate | 1 |
Note: The diluent is normal saline (0.9% NaCl).
Detection Principle
Inorganic phosphorus react with molybdic acid to produce phosphomolybdic acid. Phosphomolybdic acid can be reduced to molybdenum blue under the action of reducing agent. And the molybdenum blue have a maximum absorption peak at 660 nm. The phosphorus content can be calculated indirectly be measuring the OD value at 660 nm.
Kit Components & Storage
| Item | Component | Size 1(48 T) | Size 2(96 T) | Storage |
| Reagent 1 | Chromogenic Agent A | 10 mL × 1 vial | 20 mL×1 vial | 2-8℃, 12 months |
| Reagent 2 | Chromogenic Agent B | Powder × 1 vial | Powder × 2 vials | 2-8°C, 12 months, shading light |
| Reagent 3 | Chromogenic Agent C | Powder × 1 vial | Powder × 2 vials | 2-8°C, 12 months, shading light |
| Reagent 4 | Protein Precipitator | 20 mL × 1 vial | 40 mL × 1 vial | 2-8℃, 12 months |
| Reagent 5 | 10 mmol/L Phosphorus Standard | 1 mL × 1 vial | 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 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 (mmol/L) | 0.25 | 0.78 | 1.40 |
| %CV | 2.5 | 2.3 | 1.5 |
Inter-assay Precision
Three human serum samples were assayed 17 times in duplicate by three operators to determine precision between assays.
| Parameters | Sample 1 | Sample 2 | Sample 3 |
| Mean (mmol/L) | 0.25 | 0.78 | 1.40 |
| %CV | 3.3 | 2.6 | 3.1 |
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 101%
| Parameters | Standard 1 | Standard 2 | Standard 3 |
| Expected Conc. (mmol/L) | 0.15 | 0.65 | 1.3 |
| Observed Conc. (mmol/L) | 0.2 | 0.7 | 1.3 |
| Recovery rate (%) | 101 | 103 | 99 |
Sensitivity
The analytical sensitivity of the assay is 0.004 mmol/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.
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 | 0.1 | 0.2 | 0.5 | 0.8 | 1 | 1.5 | 2 |
| OD value | 0.053 | 0.144 | 0.231 | 0.517 | 0.87 | 1.008 | 1.473 | 1.91 |
| 0.049 | 0.142 | 0.227 | 0.508 | 0.824 | 1.016 | 1.451 | 1.929 | |
| Average OD | 0.051 | 0.143 | 0.229 | 0.512 | 0.847 | 1.012 | 1.462 | 1.919 |
| Absoluted OD | 0 | 0.092 | 0.178 | 0.461 | 0.796 | 0.961 | 1.411 | 1.868 |