Total Cholesterol(TC) Content Assay Kit
Spectrophotometry
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description Product Description
The Total Cholesterol (TC) Content Assay Kit is widely used in clinical diagnostics and research laboratories to measure cholesterol levels in biological samples such as blood, serum, or plasma. It is essential for assessing cardiovascular health, as elevated cholesterol levels are a risk factor for heart disease and atherosclerosis. The kit is also employed in pharmaceutical studies to evaluate the efficacy of cholesterol-lowering drugs or therapies. In nutritional research, it helps analyze the impact of diets or supplements on cholesterol metabolism. Additionally, it is utilized in veterinary medicine to monitor cholesterol levels in animals. The assay provides a reliable, quick, and accurate method for quantifying total cholesterol, making it a valuable tool in both medical and scientific settings.
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| Test Parameter | Specification |
|---|---|
| Performance test | PASS |
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Total Cholesterol (TC) Content Assay Kit
Product code: 112612
Method: Spectrophotometric method
Product Introduction
Total cholesterol (TC) includes free cholesterol and cholesteryl esters. TC refers to the total cholesterol contained in all lipoproteins in the tissue.
In this assay, cholesteryl esters are hydrolyzed by esterase to produce free cholesterol (FC) and free fatty acids (FFA). The FC is then oxidized by cholesterol oxidase to generate Δ4-cholestenone and H2O2. Finally, peroxidase catalyzes H2O2to oxidize 4-aminoantipyrine and phenol, producing a red quinone compound. This compound has a characteristic absorption peak at 500 nm, and the color intensity is directly proportional to the TC content.
Package Contents
| Pack Size | Code | Item | Quantity | Notes |
|---|---|---|---|---|
| 50T | 112612.1 | Reagent I | User prepared | Isopropanol |
| 50T | 112612.2 | Reagent II | 1 bottle | |
| 50T | 112612.3 | Reagent III | 1 bottle | |
| 50T | 112612.4 | Reagent IV | 1 bottle | |
| 50T | 112612.5 | TC Standard | 1 bottle | |
| 50T | 112612.m | Manual | 1 copy |
Quality Standards and Safety Instructions
| Raw Material and Packaging Name | Quality Standard | Main Toxicity |
|---|---|---|
| Reagent II | - | - |
| Reagent III | - | - |
| Reagent IV | - | - |
| TC Standard | - | - |
Transportation and Storage Conditions
| Item | Condition |
|---|---|
| Transport | Transport with ice packs. |
| Storage | Store at 2-8 ℃, protected from light. |
| Shelf Life | 180 days. |
Instructions for Use
1. Total Cholesterol Extraction
Tissue samples: Extract TC from tissue according to a tissue mass (g) to Reagent I volume (mL) ratio of 1:5-10. It is recommended to weigh about 0.1 g tissue and add 1 mL Reagent I. Homogenize in an ice bath, centrifuge at 8000 g and 4 ℃ for 10 min, then collect the supernatant as the test sample.
Cells or bacteria: Collect 400-500 × 104cells or bacteria into a centrifuge tube and discard the supernatant. Add 1 mL Reagent I, then sonicate for lysis for 1 min at 20% power using a cycle of 2 s sonication and 1 s pause. Use this as the test sample.
Serum (plasma) and other samples: Measure directly.
2. Reagent Preparation
Before use, prepare the TC working solution by pipetting about 0.8 mL of Reagent II and adding it separately to the Reagent III and Reagent IV bottles. Dissolve completely, then transfer all contents back into the Reagent II bottle and mix thoroughly. Incubate the TC working solution in a 37 ℃ water bath for 10 min before use.
Unused working solution should be stored at 4 ℃ for one month.
3. Procedure
Preheat the spectrophotometer for 30 min or longer, set the wavelength to 500 nm, and zero the instrument with distilled water.
Add reagents to EP tubes according to the table below.
Component Standard Tube Assay Tube Blank Tube TC standard (μL) 100 - - Sample solution (μL) - 100 - Reagent I (μL) - - 100 Working solution (μL) 900 900 900 Mix thoroughly, incubate at 37 ℃ for 10 min, then measure the absorbance at 500 nm. Record the values as Astandard, Aassay, and Ablank. The standard tube and blank tube only need to be measured once.
4. Total Cholesterol Content Calculation
4.1 Serum (plasma): TC (μmol/dL) = Cstandard× (Aassay- Ablank) ÷ (Astandard- Ablank) × 100 mL = 50 × (Aassay- Ablank) ÷ (Astandard- Ablank)
4.2 By sample protein concentration: TC (μmol/mg prot) = Cstandard× (Aassay- Ablank) ÷ (Astandard- Ablank) ÷ Cpr = 0.5 × (Aassay- Ablank) ÷ (Astandard- Ablank) ÷ Cpr
4.3 By sample mass: TC (μmol/g fresh weight) = Cstandard× (Aassay- Ablank) ÷ (Astandard- Ablank) ÷ W = 0.5 × (Aassay- Ablank) ÷ (Astandard- Ablank) ÷ W
4.4 In cells and bacteria: TC (μmol/104cells) = Cstandard× (Aassay- Ablank) ÷ (Astandard- Ablank) ÷ bacteria or cells (104cells/L) = 0.5 × (Aassay- Ablank) ÷ (Astandard- Ablank) ÷ bacteria or cells (104cells/L)
Cstandard: 0.5 μmol/mL
100 mL: 1 dL = 100 mL
Cpr: sample protein concentration, mg/mL
W: sample mass, g/mL
Precautions
- Before formal testing, select 2-3 samples with large expected differences for preliminary testing.
- Required instruments and supplies not provided with the kit: visible spectrophotometer, benchtop centrifuge, adjustable-volume pipette, 1 mL glass cuvette, mortar, ice, isopropanol, and distilled water.
- This 50T kit can test 48 samples.
- The minimum detection limit is 1 nmol/mL.
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