计算溶液所需的质量、体积或浓度。
总蛋白含量检测试剂盒（双缩脲, 比色法）
有货
为什么选择此级别
BioReagent,用于蛋白分析 级 ，适用于对基线干扰要求严格的色谱和分析工作流程。
储存与运输
2-8°C储存,室温。低温运输 。请查阅批次 COA 获取详细规格。
质量文档
SDS、COA、产品数据表及规格说明书均可下载。可通过批号查询获取批次 COA。
文献证明
在色谱分析、有机合成和交叉偶联反应领域已被 0 篇同行评审文献引用。
概述
总蛋白（Total Protein，TP）由白蛋白和球蛋白组成，对于生物体液（血清、尿液、脑脊液）中总蛋白质含量的测定，一般要基于如下两个假设：1、所有蛋白质分子由纯多肽组成，含氮量的质量百分比为16%；2、体液中含有数百个蛋白质分子，每个分子对测定反应都具有非常相似的特性，目前常用的检测总蛋白的方法有：双缩脲法、紫外分光光度法、染料结合法、凯氏定氮法、沉淀法等。
本产品多用于人或动物血清、血浆、组织等样本中的总蛋白含量测定，双缩脲反应的原理是在呈蓝色的碱性硫酸铜溶液存在的情况下，肽键与铜离子结合，生成蓝紫色的化合物，此化合物在540nm的吸光度与肽键的数量呈正比例关系，因此可计算出蛋白质的含量。双缩脲法测蛋白浓度兼容性亦很好，不受大部分样本中其他成分的影响，但易受铜离子螯合剂的影响，另外对于血清总蛋白的双缩脲分析，胆红素、脂类、血红蛋白、葡聚糖具有一定干扰作用，双缩脲法在5～160mg/mL浓度范围内有较好的线性关系。该试剂盒仅用于科研领域，不适用于临床诊断或其他用途。
| T1507297 | 组分名称 | 物理外观 | 120 T | 储存 | 单次测试使用量 |
| T1507297A | 双缩脲试剂 | 液体 | 250 mL | 2-8℃. | 2 mL |
| T1507297B | 蛋白标准 | 粉末 | 80 mg | RT | 40 μL |
| T1507297C | 蛋白标准配制液 | 液体 | 5 mL | RT | 按需 |
| T1507297D | 双缩脲空白试剂 | 液体 | 100 mL | 2-8℃. | 2 mL |
自备仪器和试剂
1、待测样品（血清、血浆、组织）、生理盐水（0.9% NaCl）或PBS
2、离心管或小试管、水浴锅或恒温箱、分光光度计、比色杯、匀浆器或研钵
实验流程
1、标准品的配制
取0.5ml蛋白标准配制液或稀释液加入到80mg蛋白标准中，充分溶解后即配制成蛋白标准溶液（160mg/ml），配制后可立即使用，溶解后的蛋白标准溶液应-20℃保存，亦可根据需要进行稀释，如稀释至40mg/ml。特别提示：待测蛋白溶解于什么样的稀释液中，蛋白标准也宜溶解于什么样的稀释液中，例如待测蛋白溶解于蔗糖溶液中，亦取蛋白标准溶解于蔗糖溶液中，常用0.9%NaCl或PBS作为总蛋白标准品的稀释液。
2、样本处理
2.1 血清、血浆样本，直接取40μl检测；
2.2 对于组织样本，按组织质量(g)：生理盐水(ml)=1：9比例，加入9倍体积的生理盐水，冰浴下匀浆后2500g离心10min，取上清液并补生理盐水至10ml待检。
3、TP加样
按照下表设置空白管、标准管、测定管，溶液应按照顺序依次加入，并注意避免产生气泡；如果样品浓度过高，可以减少样品用量或适当稀释后再进行测定，样品的检测最好能设置平行管。
| 加入物 (ml) | 空白管 | 标准管 | 测定管 |
| ddH₂O | 0.04 | － | － |
| 蛋白标准溶液 (如40mg/ml) | － | 0.04 | － |
| 待测样品 | － | － | 0.04 |
| 双缩脲试剂 | 2 | 2 | 2 |
注意：当遇到浑浊或溶血样本时，可设“样本空白管”：取0.04ml待测样品加入2ml双缩脲空白试剂即为“样本空白管”，以双缩脲空白试剂调零，读取“样本空白管”的吸光度，用测定管的吸光度减去“样本空白管”的吸光度后的净吸光度，计算总蛋白浓度。
4、TP测定
混匀，37℃孵育10min，比色杯光径1cm，以空白管调零，用分光光度计测定540nm处各管的吸光度，记为A标准、A测定，如无540nm，510～562nm之间的波长也可。
5、结果计算
血清、血浆样本总蛋白浓度(g/L)=A测定/A标准×C
浑浊或溶血样本总蛋白浓度(g/L)= (A测定-A样本空白)/A标准×C
100g组织样本总蛋白含量(g/100g)=A测定/A标准×C×VT×100/m
参数说明：
A测定：测定管的吸光度
A标准：标准管的吸光度
A样本空白：样本空白管的吸光度
C：蛋白标准液浓度(mg/ml=g/L)
VT：组织样本上清液总体积=10ml
m：组织样本的质量(g)
100：100g组织样本的质量
参考区间：
| 健康成人走动后血清总蛋白浓度 | 64~83g/L |
| 健康成人静卧时血清总蛋白浓度 | 60~78g/L |
注意事项
1、蛋白标准粉末溶解于蛋白标准配制液后，即获得蛋白标准原液，该原液中含有防腐剂，不影响后续检测，该蛋白标准原液-20℃长期保存。
2、待测蛋白和蛋白标准加入双缩脲试剂后，如果发现检测效果不佳，可以室温放置1h或60℃放置15min，颜色会随着时间的延长不断加深。
3、测定标准曲线时发现随着标准品浓度的增加吸光度或颜色没有明显变化，可能的原因是样品中含有铜离子螯合剂。
4、蛋白浓度低于20mg/ml时，颜色反应呈不明显的蓝色；当大于40mg/ml时，即可呈现明显的蓝紫色反应。
5、该试剂盒的线性范围是5~160 mg/ml，样品蛋白浓度应大于1mg/ml，以20mg/ml以上为宜；如需测定低浓度蛋白样品(1mg/ml)，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。
6、该试剂盒反应生成的蓝紫色的化合物的吸光度，与试剂批次、pH值、反应温度有关，因此建议每次测定都做标准对照。
7、氨基酸和二肽不发生双缩脲反应，三肽、寡肽和多肽与铜离子的双缩脲复合物，呈粉红色-紫红色，与蛋白质的双缩脲反应结果不同。
8、如果没分光光度计，也可以使用酶标仪测定，使用酶标仪测定蛋白浓度时，每个试剂盒可以测定的样品数量会显著增加。
9、检测中发现所有孔都呈暗紫色，可能原因是样品含有还原剂，应适当透析或稀释样品。
10、试剂开封后请尽快使用，以防影响后续实验效果。
附参考标准曲线范围
用酶标仪测定蛋白质标准1、5、10、20、40、60、80、100、120、140、160mg/ml在540nm的吸光度(未调零)，空白调零后40mg/ml的吸光度为0.15~0.18，据此做出其标准曲线如下：
测定结果显示，1mg/ml的吸光度比较低，与空白管接近，因此建议测定样品浓度应大于1mg/ml，以20mg/ml以上为宜；如需测定低浓度蛋白样品(1mg/ml)，请使用阿拉丁B665595 BCA蛋白定量试剂盒或R1491648 即用型BCA蛋白定量试剂盒。
Total Protein (TP) consists of albumin and globulin. The determination of total protein content in biological fluids (serum, urine, cerebrospinal fluid) is generally based on two assumptions: 1) All protein molecules are composed of pure polypeptides, with a mass percentage of nitrogen content of 16%; 2) The body fluid contains hundreds of protein molecules, each exhibiting very similar characteristics in the assay reaction. Commonly used methods for total protein detection include the biuret method, UV spectrophotometry, dye-binding methods, the Kjeldahl method, and precipitation methods.
This product is primarily used for determining total protein content in human or animal serum, plasma, tissues, and other samples. The principle of the biuret reaction is that peptide bonds complex with copper ions in the presence of an alkaline copper sulfate solution (which is blue), generating a blue-violet compound. The absorbance of this compound at 540 nm is directly proportional to the number of peptide bonds, allowing for the calculation of protein content. The biuret method for protein concentration determination also shows good compatibility and is largely unaffected by other components in most samples, although it can be susceptible to copper ion chelators. Additionally, for biuret analysis of serum total protein, bilirubin, lipids, hemoglobin, and dextran can cause some interference. The biuret method demonstrates good linearity within the concentration range of 5–160 mg/mL. This kit is intended for research use only and is not suitable for clinical diagnosis or other purposes.
| T1507297 | Component Name | Appearance | 120 T | Storage | Quantity Per Test |
| T1507297A | Biuret Reagent | Liquid | 250 mL | 2-8℃. | 2 mL |
| T1507297B | Protein Standard | Powder | 80 mg | RT | 40 μL |
| T1507297C | Protein Standard Diluent | Liquid | 5 mL | RT | As needed |
| T1507297D | Biuret Blank Reagent | Liquid | 100 mL | 2-8℃. | 2 mL |
Materials Required but Not Provided
1. Test samples (serum, plasma, tissue), physiological saline (0.9% NaCl) or PBS
2. Centrifuge tubes or small test tubes, water bath or incubator, spectrophotometer, cuvettes, homogenizer or mortar and pestle
Experimental Procedure
1. Preparation of Standard
Add 0.5 mL of Protein Standard Diluent (or an appropriate diluent) to the 80 mg Protein Standard vial. Dissolve completely to prepare the Protein Standard Solution (160 mg/mL). This solution can be used immediately after preparation. The dissolved Protein Standard Solution should be stored at -20°C. It can also be diluted as needed, for example to 40 mg/mL.
Special Note: The protein standard should ideally be dissolved/diluted in the same solution as the protein in the test samples. For instance, if the test protein is dissolved in a sucrose solution, the protein standard should also be dissolved in sucrose solution. Commonly, 0.9% NaCl or PBS is used as the diluent for the total protein standard.
2. Sample Preparation
2.1 Serum/Plasma Samples: Use 40 μL directly for detection.
2.2 Tissue Samples: Add 9 volumes of physiological saline per mass of tissue (g) (e.g., 1 g tissue + 9 mL saline). Homogenize in an ice bath, then centrifuge at 2500 × g for 10 minutes. Collect the supernatant and adjust the volume to 10 mL with physiological saline for testing.
3. TP Assay Setup
Set up Blank, Standard, and Test tubes according to the table below. Add reagents in the specified order, avoiding bubble formation. If the sample concentration is too high, reduce the sample volume or dilute appropriately before assay. Running sample duplicates is recommended.
| Reagent (mL) | Blank Tube | Standard Tube | Test Tube |
| ddH₂O | 0.04 | － | － |
| Protein Standard Solution (e.g., 40 mg/mL) | － | 0.04 | － |
| Test Sample | － | － | 0.04 |
| Biuret Reagent | 2 | 2 | 2 |
Note: For turbid or hemolyzed samples, set up a "Sample Blank Tube": Add 0.04 mL of test sample to 2 mL of Biuret Blank Reagent – this is the "Sample Blank Tube". Zero the instrument with the Biuret Blank Reagent, then read the absorbance of the "Sample Blank Tube". Use the net absorbance (A<sub>test</sub> - A<sub>sample blank</sub>) to calculate the total protein concentration.
4. TP Measurement
Mix well and incubate at 37°C for 10 minutes. Using a cuvette with a 1 cm light path and zeroing the instrument with the Blank tube, measure the absorbance of the Standard and Test tubes at 540 nm using a spectrophotometer, recorded as A<sub>standard</sub> and A<sub>test</sub>. If 540 nm is unavailable, wavelengths between 510–562 nm can also be used.
5. Calculation of Results
Serum/Plasma Sample Total Protein Concentration (g/L) = (Atest/ Astandard) × C
Turbid/Hemolyzed Sample Total Protein Concentration (g/L) = [(Atest- Asample blank) / Astandard] × C
Total Protein Content per 100g Tissue (g/100g) = (Atest/ Astandard) × C × VT × 100 / m
Parameter Description:
Atest: Absorbance of the test tube
Astandard: Absorbance of the standard tube
A<sub>sample blank</sub>: Absorbance of the sample blank tube
C: Protein Standard Solution concentration (mg/mL or g/L)
VT: Total volume of tissue supernatant = 10 mL
m: Mass of the tissue sample (g)
100: Mass for reporting content per 100g of tissue
Reference Interval:
| Condition | Serum Total Protein Concentration |
| Healthy ambulatory adults | 64 ~ 83 g/L |
| Healthy recumbent adults | 60 ~ 78 g/L |
Precautions
Dissolving the Protein Standard powder in the Protein Standard Diluent yields the stock Protein Standard Solution. This stock contains preservatives which do not interfere with the assay and can be stored long-term at -20°C.
If detection performance is suboptimal after adding Biuret Reagent to the test protein and standard, letting the reaction proceed at room temperature for 1 hour or at 60°C for 15 minutes may help, as the color continues to develop over time.
If no significant increase in absorbance or color is observed with increasing standard concentration during standard curve generation, the sample might contain copper ion chelators.
At protein concentrations below 20 mg/mL, the color reaction is a faint blue. A distinct blue-violet reaction becomes apparent at concentrations greater than 40 mg/mL.
The linear range of this kit is 5–160 mg/mL. Sample protein concentration should be greater than 1 mg/mL, preferably above 20 mg/mL. For low-concentration protein samples (~1 mg/mL), the Aladdin B665595 BCA Protein Quantification Kit or R1491648 Ready-to-Use BCA Protein Quantification Kit is recommended.
The absorbance of the blue-violet compound generated by this kit depends on the reagent batch, pH, and reaction temperature. Therefore, it is recommended to run a standard control with each assay.
Amino acids and dipeptides do not yield the biuret reaction. The biuret complexes of tripeptides, oligopeptides, and polypeptides with copper ions are pink to purplish-red, differing from the protein biuret reaction result.
If a spectrophotometer is unavailable, a microplate reader can also be used. Using a microplate reader significantly increases the number of samples measurable per kit.
If all tubes appear dark purple, the sample might contain reducing agents. Consider appropriate dialysis or dilution of the sample.
Use reagents promptly after opening to prevent potential effects on subsequent experimental results.
Appendix: Reference Standard Curve Range
The absorbance (un-zeroed) of protein standards at 1, 5, 10, 20, 40, 60, 80, 100, 120, 140, 160 mg/mL was measured at 540 nm using a microplate reader. After zeroing with the blank, the absorbance for 40 mg/mL typically ranged from 0.15 to 0.18. The resulting standard curve is shown below:
The results show that the absorbance for 1 mg/mL is relatively low and close to the blank. Therefore, it is recommended that the sample concentration for assay be greater than 1 mg/mL, preferably above 20 mg/mL. For low-concentration protein samples (~1 mg/mL), please use the Aladdin B665595 BCA Protein Quantification Kit or R1491648 Ready-to-Use BCA Protein Quantification Kit.
规格
别名
双缩脲法蛋白含量检测试剂盒 | 双缩脲法蛋白含量检测试剂盒 | 总蛋白（TP）比色法测试盒（双缩脲法）
英文别名
Biuret Protein Colorimetric Assay Kit | Biuret Protein Colorimetric Assay Kit
规格或纯度
BioReagent, 用于蛋白分析
英文名称
Protein Content Assay Kit (Biuret, Colorimetric Method)
应用
Protein Quantification
储存与运输
储存条件
2-8°C储存,室温
运输条件
低温运输
稳定性与储存
各组分在相应的储存条件下保质期1年。
组分与储存
| T1507297 | 组分名称 | 物理外观 | 120 T | 储存 | 单次测试使用量 |
| T1507297A | 双缩脲试剂 | 液体 | 250 mL | 2-8℃. | 2 mL |
| T1507297B | 蛋白标准 | 粉末 | 80 mg | RT | 40 μL |
| T1507297C | 蛋白标准配制液 | 液体 | 5 mL | RT | 按需 |
| T1507297D | 双缩脲空白试剂 | 液体 | 100 mL | 2-8℃. | 2 mL |
技术文档
高级数据
质检证书（CoA,COO,BSE/TSE 和分析图谱)
C of A & Other Certificates(BSE/TSE, COO):
输入批号以搜索分析图谱:
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 |
|---|---|---|
 G2608553 |
分析证书 | T1507297 |
| ZJ26F0131501 |
分析证书 | T1507297 |
溶液计算器
常见问题
什么是「用于蛋白分析」？
「用于蛋白分析」表示该产品经过加工和检测，适用于分子生物学和蛋白质组学。典型规格包括：低或不可检测的核酸酶/蛋白酶活性、低内毒素、受控的微生物限度，以及高批间重现性。
什么是「BioReagent」？
「BioReagent」表示该产品经过加工和检测，适用于生物和生化应用。规格可能包括内毒素限值、微生物控制、无菌性、低 DNase/RNase/蛋白酶活性，以及在相关情况下的生物活性验证。
本产品应如何储存？
请于2–8 °C条件下保存。需冷藏保存以维持规定的有效期。
本产品如何运输？
本产品采用湿冰冷藏运输。收货后请立即拆包，并按上述储存条件保存。
如何获取批次专用COA？
每个订单都会随附分析证书（COA），您也可以从账户订单历史中下载。如需订购前的COA，请联系您的客户代表，或使用PDP上的"申请COA"按钮——我们将提供代表性批次的COA。
产品仅提供给经过验证的企业、机构和合格的专业人员用于研发。不适用于人类、动物、诊断或治疗。
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