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Choline/Acetylcholine Fluorometric Assay Kit
SKU: E-BC-F055-96
Choline/Acetylcholine Fluorometric Assay Kit
| SKU # | E-BC-F055 |
| Detection Instrument | Fluorescence microplate reader (Ex/Em=535 nm/587 nm) |
| Detection Method | Fluorescence method |
Product Details
Properties
| Sample Type | Serum, plasma, animal tissue |
| Sensitivity | 0.06 μmol/L |
| Detection Range | 0.06-10 μmol/L |
| Detection Method | Fluorescence method |
| Precision | Inter-assay CV: 8.2-10% | intra-assay CV: 3.2-5% |
| Other Reagents Required | Normal saline (0.9% NaCl) |
| 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% Mouse heart tissue homogenate | 10-20 |
| 10% Rat liver tissue homogenate | 10-20 |
| 10% Mouse lung tissue homogenate | 10-20 |
| 10% Mouse kidney tissue homogenate | 15-25 |
| 10% Mouse brain tissue homogenate | 15-25 |
| 10% Mouse liver tissue homogenate | 10-20 |
| Human serum | 3-5 |
| Rat serum | 3-7 |
| Rat plasma | 1 |
The diluent is normal saline (0.9% NaCl). For the dilution of other sample types, please do pretest to confirm the dilution factor.
Detection Principle
Choline and acetylcholine play important roles in many biological processes. Choline is an essential nutrient, often grouped with B complex vitamins, that plays a key role in many biological processes. Choline is a precursor to the synthesis of acetylcholine (AchE), an important neurotransmitter in the peripheral and central nervous systems. Choline is measured by enzyme coupling reaction to generate fluorescent products in this kit, and the content of fluorescent substances is proportional to the presence of choline. Acetylcholinesterase was added to the reaction to hydrolyze acetylcholine into choline and acetic acid, and the acetylcholine content was further calculated by measuring the total amount of choline and acetylcholine.
Kit Components & Storage
| Item | Component | Size (96 T) | Storage |
| Reagent 1 | Buffer Solution | 20 mL ×1 vial | -20℃, 12 months |
| Reagent 2 | Hydrolase | 1.3 mL ×2 vials | -20℃, 12 months shading light |
| Reagent 3 | Complex Enzyme | 0.4 mL ×1 vial | -20℃, 12 months shading light |
| Reagent 4 | 1 mmol/L Standard Solution |
0.1 mL ×1 vial | -20℃, 12 months shading light |
| Reagent 5 | Chromogenic Agent | 0.1 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 (μmol/L) | 1.00 | 3.00 | 7.00 |
| %CV | 5.0 | 3.2 | 3.8 |
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 (μmol/L) | 1.00 | 3.00 | 7.00 |
| %CV | 8.2 | 10.0 | 8.8 |
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 100%.
| Standard 1 | Standard 2 | Standard 3 | |
| Expected Conc. (μmol/L) | 2.5 | 5 | 8.5 |
| Observed Conc. (μmol/L) | 2.5 | 5.0 | 8.5 |
| Recovery rate (%) | 101 | 99 | 100 |
Sensitivity
The analytical sensitivity of the assay is 0.06 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 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 | 2 | 3 | 4 | 6 | 8 | 9 | 10 |
| Fluorescence value | 1021 | 2487 | 3379 | 4326 | 6118 | 7603 | 8179 | 9317 |
| 1083 | 2485 | 3565 | 4265 | 6220 | 7886 | 8611 | 9524 | |
| Average fluorescence value | 1052 | 2486 | 3472 | 4296 | 6169 | 7745 | 8395 | 9421 |
| Absoluted fluorescence value | 0 | 1434 | 2420 | 3244 | 5117 | 6693 | 7343 | 8369 |