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。
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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。
BCA（Bicinchoninic acid）法是⽬前应⽤⽐较⼴泛的蛋⽩质浓度测定⽅法。基于双缩脲反应，即在碱性环境下蛋⽩质将 Cu2+还原成 Cu+，产⽣⼀种紫蓝⾊复合物，在 562nm 处有⾼的吸光值，该反应产物的量与蛋⽩质浓度成正⽐。BCA 蛋⽩浓度测定法实现了蛋⽩质浓度测定的简便、灵敏、快速和稳定性。试剂盒中提供的蛋⽩标准品为⽤户制作标准曲线提供了便利。该 BCA 蛋⽩定量试剂盒可⽤于⽐⾊⽫法检测，也可⽤于微孔板法检测。前者虽需较⼤量（100 μL）的蛋⽩样品，但由于其在检测中使⽤蛋⽩样品与 BCA ⼯作液的⽐率为 1:20（v/v），从⽽降低⼲扰物质带来的影响。后者操作简单⽅便，仅需少量（10-25 μL）的蛋⽩样品。不过，由于其在检测中使⽤蛋⽩样品与 BCA ⼯作液的⽐率为 1:10（v/v），某种程度上限制⼲扰物质的承受浓度以及降低最低检测⽔平。
产品特点
1、灵敏度高，检测范围：20-2000 μg/mL。
2、适用于酶标仪、分光光度计。
3、准确性高，变异系数远小于考马斯亮蓝染色法。
4、不受大部分样品中的化学物质的影响，BCA 法测定蛋白浓度的最大优点是蛋白浓度的测定可以耐受高浓度的去垢剂，可以兼容样品中高达 1% SDS、1% Triton X- 100、1% Tween 20, 60, 80。但受螯合剂和略高浓度的还原剂的影响，需确保 EDTA 低于 10 mM，无 EGTA，DTT 低于1 mM；β-巯基乙醇低于0.01%。详情见附表 1。
5、本试剂盒含有一系列浓度的蛋白质标准溶液（BSA 溶液），即取即用，无需稀释，方便快捷。
图1. 本试剂盒蛋白标准曲线的实测效果图。图中数据仅供参考，实际的检测效果可能会略有不同。
使用方法
1、BCA工作液配制
依据样品数量，将试剂A和试剂B按体积比50:1配制适量BCA工作液， 并充分混匀。 两者混合时会有沉淀形成，彻底混匀后沉淀消失。注：配制BCA工作液前请将试剂A摇晃混匀。新配制的 BCA 工作液室温密封条件下可稳定保存 24 h。
2、微孔板测定法
(1) 各取 10 μL 的标准品和未知样品至相应标记的微孔板中。
(2) 滴加 100 μL BCA 工作液至每孔混匀并充分震荡30秒。
(3) 停止震荡。对于大多数情况，37 ℃放置30分钟； 注：也可以室温放置 2 小时，或 60 ℃放置 30 分钟。 BCA 法测定蛋白浓度时，颜色会随着时间的延长不断加深，并且显色反应会因温度升高而加快。如果浓度较低，适合在较高温度孵育，或适当延长孵育时间。
(4) 在读数仪器中测量 562 nm时的吸光值。
(5) 绘制标准曲线，计算样品中的蛋白浓度。如果蛋白浓度不在标准曲线范围内, 请稀释样品后测定。
注意事项：
1、BCA 蛋⽩测定法测定时颜⾊会随着时间的延⻓不断加深，显⾊反应的速度和温度有关，需注意保持定时和定温，以确保精确定量。
2、⻓期保存，若发现 BCA 试剂 A 或者试剂 B 有沉淀发⽣。请 37ºC 温育并伴随搅拌促使其充分溶解。如发现细菌污染，则应丢弃。
3、实验操作规范，提⾼上样量的精确度。
4、每次测定都需做相应的标准曲线，因为显⾊反应与温度和时间的变化有关，精准蛋⽩定量宜每次都做标准曲线。
5、如果蛋白样品珍贵或量少，可以取 5μL 样品和标准品至相应标记的微孔板中，滴加 95μL BCA工作液，不影响浓度的测定。
6、本公司所有产品仅限用于研发，必须专业人员操作同时佩戴口罩/手套/实验服并遵守生物实验室安全操作规程！
干扰素物质附表1
The BCA (Bicinchoninic acid) method is currently a widely used method for protein concentration determination. Based on the diazotization reaction, that is, in an alkaline environment, proteins will reduce Cu2+ to Cu+ and produce a purple-blue complex, with a high absorbance value at 562nm. The amount of the reaction product is proportional to the protein concentration. The BCA protein concentration determination method achieves the simplicity, sensitivity, rapidity and stability of protein concentration determination. The protein standard substances provided in the kit facilitate users to create a standard curve. This BCA protein concentration determination kit can be used for colorimetric method detection or microplate method detection. The former requires a relatively large amount (100μL) of protein samples, but since the ratio of protein samples to BCA working solution used in the detection is 1:20 (v/v), it reduces the influence of interfering substances. The latter is simple and convenient to operate, only requiring a small amount (10-25μL) of protein samples. However, due to the ratio of protein samples to BCA working solution used in the detection being 1:10 (v/v), it to some extent limits the tolerance concentration of interfering substances and reduces the minimum detection level.
Product features
1. High sensitivity, detection range: 20-2000 μg/mL.
2. Suitable for microplate readers and spectrophotometers.
3. High accuracy, the coefficient of variation is much lower than that of the Coomassie Brilliant Blue staining method.
4. Not affected by most chemical substances in the samples. The greatest advantage of the BCA method for protein concentration determination is that it can tolerate high concentrations of detergents, and can be compatible with up to 1% SDS, 1% Triton X-100, 1% Tween 20, 60, 80 in the samples. However, it is affected by chelating agents and slightly higher concentrations of reducing agents. It is necessary to ensure that EDTA is below 10 mM, without EGTA, DTT is below 1 mM; β-mercaptoethanol is below 0.01%. Details can be found in Table 1.
5. This kit contains a series of protein standard solutions (BSA solutions) at a range of concentrations, which are ready to use without dilution, providing convenience and efficiency.
Figure 1. The actual measurement result of the protein standard curve of this kit. The data shown in the figure are for reference only; the actual detection outcome may vary slightly.
Usage Method
1. Preparation of BCA working solution:
Based on the number of samples, prepare an appropriate amount of BCA working solution by mixing reagent A and reagent B in a volume ratio of 50:1, and mix thoroughly. When the two are mixed, a precipitate will form, but it will disappear after thorough mixing. Note: Shake reagent A well before preparing the BCA working solution. The newly prepared BCA working solution can be stably stored at room temperature for 24 hours under sealed conditions.
2. Microplate Assay Method
(1) Take 10 μL of the standard sample and the unknown sample and add them to the corresponding labeled microplates.
(2) Add 100 μL of BCA working solution to each well and mix well and shake for 30 seconds.
(3) Stop the shaking. For most cases, place at 37 ℃ for 30 minutes; Note: It can also be placed at room temperature for 2 hours or at 60 ℃ for 30 minutes. When using the BCA method to determine protein concentration, the color will deepen over time, and the color development reaction will accelerate due to the increase in temperature. If the concentration is low, it is suitable to incubate at a higher temperature or extend the incubation time appropriately.
(4) Measure the absorbance at 562 nm in the reading instrument.
(5) Draw a standard curve and calculate the protein concentration in the sample. If the protein concentration is not within the range of the standard curve, please dilute the sample and measure it again.
Notes:
1. During the BCA protein determination process, the color will gradually deepen over time. The speed of the color development reaction is related to the temperature. It is necessary to ensure that the reaction is carried out at a fixed time and temperature to achieve precise quantification.
2. For long-term storage, if you find that reagent A or reagent B of BCA has precipitation. Please incubate at 37ºC with stirring to facilitate its complete dissolution. If bacterial contamination is detected, it should be discarded.
3. Follow the experimental operation procedures strictly to improve the accuracy of the sample loading volume.
4. For each determination, a corresponding standard curve should be made because the color development reaction is related to changes in temperature and time. Precise protein quantification should be done with a standard curve every time.
5. If the protein sample is precious or in small quantity, 5 μL of the sample and the standard can be taken to the corresponding labeled microplate wells, and 95 μL of BCA working solution can be added. This will not affect the determination of concentration.
6. All products of our company are only for research use. Professional personnel must operate and wear masks, gloves, and laboratory coats, and follow the safety operation procedures of the biological laboratory.
Interferon Substance List 1
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B1507333 | Components | 500T | 5×500T | Storage Temperature | Quantity Per Test |
B1507333A | BCA试剂 A | 60 mL | 5×60 mL | Room Temperature | 100 μL |
B1507333B | BCA试剂 B | 2 mL | 5×2 mL | Room Temperature | 2 μL |
B1507333C | BSA标准品 (2 mg/mL) | 1 mL | 5×1 mL | 2-8°C | 10 μL |
B1507333D | BSA标准品 (1.5 mg/mL) | 1 mL | 5×1 mL | 2-8°C | 10 μL |
B1507333E | BSA标准品 (1.0 mg/mL) | 1 mL | 5×1 mL | 2-8°C | 10 μL |
B1507333F | BSA标准品 (0.75 mg/mL) | 1 mL | 5×1 mL | 2-8°C | 10 μL |
B1507333G | BSA标准品 (0.5 mg/mL) | 1 mL | 5×1 mL | 2-8°C | 10 μL |
B1507333H | BSA标准品 (0.25 mg/mL) | 1 mL | 5×1 mL | 2-8°C | 10 μL |
B1507333I | BSA标准品 (0.125 mg/mL) | 1 mL | 5×1 mL | 2-8°C | 10 μL |
B1507333J | BSA标准品 (0 mg/mL) | 1 mL | 5×1 mL | 2-8°C | 10 μL |
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 |
|---|---|---|
 ZJ26F0434245 |
分析证书 | B1507333 |
| ZJ26F0332976 |
分析证书 | B1507333 |
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