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甘油三酯（TG）是由三分子长链脂肪酸与甘油缩合形成的脂肪分子，是机体重要的能量储备物质与呼吸底物。
检测原理：甘油三酯先经脂蛋白脂肪酶水解为甘油和游离脂肪酸；甘油在甘油激酶（GK）催化下生成甘油‑1‑磷酸（G‑1‑P）；甘油‑1‑磷酸再经甘油磷酸氧化酶（GPO）氧化产生过氧化氢（H₂O₂）；过氧化氢与 4‑氨基安替比林等底物反应生成红色醌类化合物，该产物在 510 nm 处有特征吸收峰，通过检测此波长下的吸光度，即可定量计算样本中甘油三酯含量。
自备仪器和试剂
操作步骤
建议先选取1-3个差异大的样本（例如不同类型或分组）进行预实验，熟悉操作流程，根据预实验结果确定或调整样本浓度，以防造成样本或试剂不必要的浪费！
1. 样本提取
1.1 组织样本
称取约0.1 g组织样本加入研钵中，加入1 mL提取液，在冰上进行匀浆，12000rpm，4℃或室温离心10 min，取上清液待测。
注：若组织样本为高脂样本或部分为高脂样本，需用无水乙醇进行提取。
1.2 细菌/细胞样本
先收集细菌或细胞到离心管内，离心后弃上清；取约500万细菌或细胞加入1 mL提取液，超声波破碎细菌或细胞（冰浴，功率200 W，超声3 s，间隔10 s，重复30次）；12000rpm 4℃ 或室温离心10 min，取上清，置冰上待测。
注：若增加样本量，可按照细菌/细胞数量（10⁴）：提取液（mL）为500~1000：1的比例进行提取。
1.3 液体样本
澄清的液体样本直接测定，若浑浊则离心后取上清检测。
2. 检测步骤
2.1 酶标仪预热30 min，调节波长到510 nm。
2.2 所有试剂平衡至室温（25℃）。
2.3 在96孔板中依次加入：
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注：
3. 结果计算
3.1 按样本质量计算
TG (μg/g 重量)=(C标准×V2)×(A测定-A空白)÷(A标准-A空白)÷(W×V1÷V)×Mr×D=C标准×(A测定-A空白)÷(A标准-A空白)÷W×Mr×D
3.2 按细胞数量计算
TG (μg/10⁴ cells)=(C标准×V2)×(A测定-A空白)÷(A标准-A空白)÷(500×V1÷V)×Mr×D=C标准×(A测定-A空白)÷(A标准-A空白)×1.278×D
3.3 液体中TG含量计算
TG (μg/mL)=(C标准×V2)×(A测定-A空白)÷(A标准-A空白)÷V1×Mr×D =C标准×(A测定-A空白)÷(A标准-A空白)×Mr×D
TG (mmol/L)=(C标准×V2) ×(A测定-A空白)÷(A标准-A空白)÷V1×D=C标准×(A测定-A空白)÷(A标准-A空白)×D
3.4 按蛋白浓度计算
TG (μg/mg prot)=(C标准×V2)×(A测定-A空白)÷(A标准-A空白)÷(Cpr×V1÷V)×D=C标准×(A测定-A空白)÷(A标准-A空白)÷Cpr×Mr×D
参数说明
C标准：标品浓度，2.45mmol/L；
Mr：甘油三酯分子量，639；
V：提取液体积，1mL；
V1：样本加入体积，0.005 mL；
V2：标准品加入体积，0.005mL；
D：稀释倍数，未稀释即为1；
W：样本取样质量，g；
500：细胞数量，500×10⁴个；
A测定：测定管吸光度值；
A标准：标准管吸光度值；
A空白：空白管吸光度值。
注意事项
1. 试剂二和标准品对光敏感，全程避光保存和使用，避免阳光直射。
2. 冰浴操作的样本和试剂，避免反复冻融，未用完的试剂及时放回 4℃保存。
3. 推荐使用阿拉丁B665595 BCA蛋白定量试剂盒或R1491648 即用型BCA蛋白定量试剂盒。
Triglycerides (TG) are fat molecules formed by the condensation of three long-chain fatty acids with glycerol. They serve as crucial energy reserve substances and respiratory substrates in organisms.
Assay Principle:
Triglycerides are first hydrolyzed by lipoprotein lipase into glycerol and free fatty acids. Glycerol is then phosphorylated by glycerol kinase (GK) to produce glycerol‑1‑phosphate (G‑1‑P). Glycerol‑1‑phosphate is subsequently oxidized by glycerol‑3‑phosphate oxidase (GPO), generating hydrogen peroxide (H₂O₂). The hydrogen peroxide reacts with substrates such as 4‑aminoantipyrine to form a red quinone compound. This product has a characteristic absorption peak at 510 nm. By measuring the absorbance at this wavelength, the triglyceride content in the sample can be quantitatively determined.
Reagents, consumables and Equipments not provided
Procedure
It is recommended to first perform a pilot experiment using 1-3 samples with significant differences (e.g., different types or groups) to familiarize yourself with the procedure. Determine or adjust sample concentrations based on pilot results to avoid unnecessary waste of samples or reagents.
1. Sample Extraction
1.1 Tissue Samples
Weigh approximately 0.1 g of tissue and add it to a mortar with 1 mL of Extraction Solution. Homogenize on ice. Centrifuge at 12,000 rpm for 10 minutes at 4°C or room temperature. Collect the supernatant for assay.
Note: For high-fat tissue samples or partially high-fat samples, extraction with absolute ethanol is required.
1.2 Bacterial/Cell Samples
Collect bacteria or cells into a centrifuge tube, centrifuge, and discard the supernatant. Take approximately 5 million bacteria/cells and add 1 mL of Extraction Solution. Sonicate to disrupt the bacteria/cells (ice bath, 200 W power, 3 seconds sonication with 10-second intervals, repeated 30 times). Centrifuge at 12,000 rpm for 10 minutes at 4°C or room temperature. Collect the supernatant and keep it on ice for assay.
Note: To increase sample size, extraction can be performed at a ratio of 500~1000 (bacteria/cell count, ×10⁴) : 1 (Extraction Solution, mL).
1.3 Liquid Samples
Clear liquid samples can be assayed directly. If turbid, centrifuge and use the supernatant for detection.
2. Assay Procedure
2.1 Preheat the microplate reader for 30 minutes and set the wavelength to 510 nm.
2.2 Thaw all reagents to room temperature (25°C).
2.3 Add the following to a 96-well plate in order:
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Notes:
If A<sub>Assay</sub> > 0.3, dilute the sample (using Extraction Solution) or reduce the sample volume V1 (e.g., to 2 μL, with a corresponding increase in Reagent 1). The dilution factor D or the adjusted V1 must be used in the calculation formula.
For high-TG samples like serum or egg yolk (which requires ethanol extraction), sample volume V1 can be reduced to 2.5 μL and made up to 20 μL total with 17.5 μL distilled water, then proceed according to the table above. The adjusted V1 must be used in the calculation.
If (A<sub>Assay</sub> - A<sub>Blank</sub>) < 0.01, increase the sample volume V1 (e.g., from 5 μL to 10 μL, with a corresponding decrease in Reagent 1). The adjusted V1 must be used in the calculation formula.
3. Calculation of Results
3.1 Calculation Based on Sample Weight
TG (μg/g weight) = (CStd × V2) × (AAssay - ABlank) ÷ (AStd - ABlank) ÷ (W × V1 ÷ V) × Mr × D
= CStd × (AAssay - ABlank) ÷ (AStd - ABlank) ÷ W × Mr × D
3.2 Calculation Based on Cell Number
TG (μg/10⁴ cells) = (CStd × V2) × (AAssay - ABlank) ÷ (AStd - ABlank) ÷ (500 × V1 ÷ V) × Mr × D
= CStd × (AAssay - ABlank) ÷ (AStd - ABlank) × 1.278 × D
3.3 Calculation of TG Content in Liquid
TG (μg/mL) = (CStd × V2) × (AAssay - ABlank) ÷ (AStd - ABlank) ÷ V1 × Mr × D
= CStd × (AAssay - ABlank) ÷ (AStd - ABlank) × Mr × D
TG (mmol/L) = (CStd × V2) × (AAssay - ABlank) ÷ (AStd - ABlank) ÷ V1 × D
= CStd × (AAssay - ABlank) ÷ (AStd - ABlank) × D
3.4 Calculation Based on Protein Concentration
TG (μg/mg prot) = (CStd × V2) × (AAssay - ABlank) ÷ (AStd - ABlank) ÷ (Cpr × V1 ÷ V) × D
= CStd × (AAssay - ABlank) ÷ (AStd - ABlank) ÷ Cpr × Mr × D
Parameter Definitions:
CStd: Standard concentration 2.45mmol/L.
Mr: Molecular weight of triglyceride, 639.
V: Volume of Extraction Solution, 1 mL.
V1: Sample volume added, 0.005 mL.
V2: Standard volume added, 0.005 mL.
D: Dilution factor, 1 if not diluted.
W: Sample weight taken, g.
500: Cell number, 500 × 10⁴ cells.
AAssay: Absorbance of the assay well.
AStd: Absorbance of the standard well.
ABlank: Absorbance of the blank well.
Notes
Reagent 2 and the Standard are light-sensitive. Store and use protected from light at all times, avoiding direct sunlight.
For samples and reagents requiring ice bath operation, avoid repeated freeze-thaw cycles. Return unused reagents to 4°C storage promptly.
Aladdin's BCA Protein Quantification Kit (B665595) or Ready-to-Use BCA Protein Quantification Kit (R1491648) is recommended for protein concentration measurement.
| T1505523 | Component | 100T | Storage |
| T1505523A | 提取液 | 100 mL | 2-8℃. |
| T1505523B | 试剂一 | 15 mL | 2-8℃. |
| T1505523C | 试剂二 | 10 mL | 2-8℃. Store in the dark. |
| T1505523D | 标准品 (2.45mmol/L) | 1 EA | 2-8℃. Store in the dark |
通过匹配包装上的批号来查找并下载产品的 COA，每批产品都进行了严格的验证，您可放心使用！
| 批号(Lot Number) | 证书类型 | 货号 |
|---|---|---|
 ZJ26F0535652 |
分析证书 | T1505523 |
| ZJ26F0332662 |
分析证书 | T1505523 |
危险品化学品经营许可证（带存储）