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Glutamic Acid Colorimetric Assay Kit
SKU: E-BC-K118-S-100
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Glutamic Acid Colorimetric Assay Kit
| SKU # | E-BC-K118-S |
| Detection Instrument | Spectrophotometer (340 nm) |
| Detection Method | Colorimetric method |
Product Details
Properties
| Sample Type | Serum, plasma, animal tissue, cells, cell culture supernatant |
| Sensitivity | 4.00 μmol/L |
| Detection Range | 4.00-450 μmol/L |
| Detection Method | Colorimetric method |
| Assay type | Quantitative |
| Assay time | 60 min |
| Precision | Average inter-assay CV: 2.900% | Average intra-assay CV: 2.200% |
| Other instruments required | Micropipettor, Centrifuge, Incubator, Water bath, Vortex mixer |
| Other reagents required | Normal saline (0.9% NaCl), PBS (0.01 M, pH 7.4) |
| Storage | Reagent 1, Reagent 2, Reagent 8, Reagent 9: 2-8℃; Reagent 3, Reagent 4, Reagent 5, Reagent 6, Reagent 7: -20℃ |
| 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 (4.00-450 μmol/L).
The recommended dilution factor for different samples is as follows (for reference only):
| Sample type | Dilution factor |
| Human serum | 1 |
| Mouse serum | 1 |
| 10% Rat liver tissue homogenate | 1 |
| 10% Rat kidney tissue homogenate | 1 |
| 10% Rat brain tissue homogenate | 1 |
Note: The diluent is normal saline (0.9% NaCl) or PBS (0.01 M, pH 7.4).
Detection Principle
Glutamic acid can react with NAD+ under the catalysis of glutamate dehydrogenase to produce α-ketoglutaric acid, NADH and NH4+. NADH has the maximum absorption at 340 nm. And glutamic acid content can be calculated by measuring the change of NADH.
Kit Components & Storage
| Item | Component | Size 1 (50 Assays) |
Size 2 (100 Assays) |
Storage |
| Reagent 1 | Protein Precipitator | 35 mL ×1 vial | 35 mL × 2 vials | -20℃, 12 months |
| Reagent 2 | Buffer Solution | 60 mL × 1 vial | 60 mL × 2 vials | -20℃, 12 months, shading light |
| Reagent 3 | Chromogenic Agent | Powder ×2 vials | Powder ×2 vials | -20℃, 12 months |
| Reagent 4 | Chromogenic Agent Diluent | 8 mL ×1 vial | 16 mL ×1 vial | -20℃, 12 months |
| Reagent 5 | Accelerator | Powder ×2 vials | Powder ×2 vials | -20℃, 12 months |
| Reagent 6 | Enzyme Reagent | Powder ×2 vials | Powder ×2 vials | -20℃, 12 months |
| Reagent 7 | Enzyme Diluent | 1.5 mL ×1 vial | 1.5 mL ×2 vials | -20℃, 12 months |
| Reagent 8 | Standard | Powder ×2 vials | Powder ×2 vials | -20℃, 12 months |
| Reagent 9 | Standard Diluent | 35 mL ×1 vial | 35 mL ×1 vial | -20℃, 12 months |
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) | 25.40 | 106.80 | 286.50 |
| %CV | 2.5 | 2.1 | 2.0 |
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) | 25.40 | 106.80 | 286.50 |
| %CV | 3.0 | 3.1 | 2.6 |
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 104%.
| Standard 1 | Standard 2 | Standard 3 | |
| Expected Conc. (μmol/L) | 84.5 | 252 | 347 |
| Observed Conc. (μmol/L) | 88.7 | 257.0 | 364.4 |
| Recovery rate (%) | 105 | 102 | 105 |
Sensitivity
The analytical sensitivity of the assay is 4.00 μmol/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 (μmol/L) | 0 | 25 | 50 | 100 | 200 | 400 | 450 |
| Average (A2-A1) | 0.016 | 0.057 | 0.090 | 0.163 | 0.295 | 0.521 | 0.576 |
| Absoluted OD | 0 | 0.041 | 0.074 | 0.147 | 0.279 | 0.505 | 0.560 |