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Aspartate Aminotransferase (AST/GOT) Activity Fluorometric Assay Kit
SKU: E-BC-F043-96
Aspartate Aminotransferase (AST/GOT) Activity Fluorometric Assay Kit
| SKU # | E-BC-F043 |
| Detection Instrument | Fluorescence microplate reader (Ex/Em=535 nm/587 nm) |
| Detection Method | Fluorescence method |
Product Details
Properties
| Synonyms | AST;GOT |
| Sample Type | Cell, animal tissue, serum (plasma), other liquid samples |
| Sensitivity | 0.03 U/L |
| Detection Range | 0.03-1.5 U/L |
| Detection Method | Fluorescence method |
| Assay Time | 80 min |
| Precision | Inter-assay CV: 7.3% | intra-assay CV: 2.2% |
| Storage | This product can be stored at -20°C for 12 months with shading light. |
| Valid period | 12 months |
Dilution Of Sample
The recommended dilution factor for different samples is as follows (for reference only):
| Sample type | Dilution factor |
|---|---|
| 10% Rat heart tissue homogenate | 300-600 |
| 10% Rat brain tissue homogenate | 300-500 |
| 10% Rat liver tissue homogenate | 200-500 |
| Human serum | 5-15 |
| 10% Rat kidney tissue homogenate | 200-500 |
| Rat serum | 10-20 |
| 10% Rat spleen tissue homogenate | 50-100 |
| 293T cell(1×10^6) | 10-20 |
| 10% Mouse lung tissue homogenate | 100-200 |
| 293T cell supernatant | 5-15 |
The diluent is buffer solution. For the dilution of other sample types, please do pretest to confirm the dilution factor.
Detection Principle
AST can catalyze the substrate to produce pyruvate. Pyruvate is oxidized to produce hydrogen peroxide, which causes the fluorescent probe to produce fluoresce. The activity of AST can be calculated by measuring the increase of fluorescence value at the excitation wavelength of 535 nm and the emission wavelength of 587 nm.
Kit Components & Storage
| Item | Component | Size 1(48 T) | Size 2(96 T) | Storage |
| Reagent 1 | Buffer Solution | 60 mL × 1 vial | 60 mL × 2 vials | -20℃, 12 months |
| Reagent 2 | Substrate | 1.5 mL × 1 vial | 1.5 mL × 2 vials | -20℃, 12 months shading light |
| Reagent 3 | Enzyme Reagent | Powder × 1 vial | Powder × 2 vials | -20℃, 12 months shading light |
| Reagent 4 | Aqueous Alkali | 0.3 mL × 1 vial | 0.6 mL × 1 via | -20℃, 12 months shading light |
| Reagent 5 | 100 mmol/L Pyruvate Standard | 1 mL × 1 vial | 1 mL × 1 vial | -20℃, 12 months shading light |
| Black 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 (U/L) | 0.10 | 0.80 | 1.20 |
| %CV | 2.5 | 2.2 | 1.9 |
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 (U/L) | 0.10 | 0.80 | 1.20 |
| %CV | 6.7 | 7.5 | 7.7 |
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. (μmol/L) | 15 | 36.5 | 55 |
| Observed Conc. (μmol/L) | 14.9 | 35.8 | 56.7 |
| Recovery rate (%) | 99 | 98 | 103 |
Sensitivity
The analytical sensitivity of the assay is 0.03 U/L. This was determined by adding two standard deviations to the mean F. obtained when the zero standard was assayed 20 times, and calculating the corresponding concentration.
Standard Curve:
As the fluorescence 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 | 50 | 60 | 80 |
| Average fluorescence value | 1019 | 2598 | 4185 | 5490 | 7018 | 8482 | 9917 | 12725 |
| Absoluted fluorescence value | 0 | 1579 | 3166 | 4471 | 5999 | 7463 | 8898 | 11706 |