BCA蛋白定量试剂盒
BCA Protein Assay Kit
本产品冰袋运输; BSA标准品 -20℃保存,其它组分4℃保存,保质期12个月。
货号规格
goodZJ101L 500次(微孔)
goodZJ101L 500次(微孔)×5
产品内容
组分 |
ZJ101 |
ZJ101L |
试剂A |
100 mL |
100 mL×5 |
试剂B |
3 mL |
3 mL×5 |
BSA标准品(5mg/mL) |
1 mL |
1 mL×5 |
产品特点
g
d准确性高——变异系数远小于考马斯亮蓝染色法;
g
d线性范围宽——灵敏,检测范围:20~2,000 μg/mL;
g
d兼容性好——与金属离子、还原剂、螯合剂及去污剂兼容性较好。
产品简介
goodBCA蛋白定量法是目前广泛使用的蛋白定量方法之一。本产品是基于BCA(Bicin-choninic Acid)法研制而成,实现了对蛋白质进行快速、稳定、灵敏的浓度测定。其原理是在碱性环境下蛋白质分子中的肽链结构能与Cu2+络合生成络合物,同时将Cu2+还原成Cu+,BCA试剂可敏感特异地与Cu+结合,形成稳定的有颜色的复合物,其在562 nm处有高的光吸收值,颜色的深浅与蛋白质浓度成正比,可根据吸收值的大小来测定蛋白质的含量。本试剂盒含有牛血清白蛋白(BSA)溶液作为蛋白质标准品溶液,测定范围为20~2,000 μg/mL。
使用说明
good◈以微孔酶标仪法为例:
gooddd1. 稀释BSA标准品:
gooddd1. 取120 μL蛋白标准品(需完全融解),用与待测蛋白样品相一致的稀释液稀释至300 μL,使终浓度为2 mg/mL。
注意:为方便起见,也可以用0.9%生理盐水或PBS缓冲液稀释标准品。
gooddd2. 配置显色工作液:
gooddd1. a. 计算显色工作液总量:
goodgoodgoodd工作液总量 = (BSA标准品样本个数+待测样本个数)×复孔数×每个样本显色工作液体积
gooddd1. a. 举例:BSA标准品样本个数为9个,待测样本个数3个,复孔数3个。
goodgoodgoodd显色工作液总量 = (9个BSA标准品样本+3个待测样本)×3个复孔×200 μL(每个样本工作液体积) = 7.2 mL
gooddd1. b. 根据计算出的所需显色工作液用量,将试剂A和试剂B按照50:1的体积比,配制显色工作液,充分混匀。
gooddd1. b. 注意:1)由于加样可能存在误差,建议配制BCA工作液时,多配制1~2个孔的量;
gooddd1. b. 注意:2)新配制的BCA工作液室温密封条件下可稳定保存24 h。
gooddd3. 定量检测
gooddd3. (1)将稀释后的标准品按0,1,2,4,6,8,10,15,20 μL分别加到96孔板中,加入用于稀释标准品的溶液补足到20 μL(为避免枪尖损失,可先补足稀释液后加入标准品);
|
|
孔号 |
1 |
2 |
3 |
4 |
5 |
6 |
7 |
8 |
9 |
|
|
稀释后的标准品(μL) |
0 |
1 |
2 |
4 |
6 |
8 |
10 |
15 |
20 |
|
|
稀释液(μL) |
20 |
19 |
18 |
16 |
14 |
12 |
10 |
5 |
0 |
|
|
BSA终浓度(μg/mL) |
0 |
100 |
200 |
400 |
600 |
800 |
1000 |
1500 |
2000 |
gooddd3. (2)将样品适当稀释(可以多作几个梯度,如2倍、4倍、8倍稀释),加20 μL到96孔板的样品孔中;
gooddd3. (3)各孔加入200 μL显色工作液,充分混匀,盖上96孔板盖,37℃孵育30 min,冷却至室温;
gooddd3. (3)注意:也可以室温放置2 h,或60℃放置30 min。BCA法测定蛋白浓度时,吸光度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。如果蛋白浓度较低,可在较高温度孵育,或延长孵育时间。
gooddd3. (4)用酶标仪测定每个样品及BSA标准品的A562,或540~590 nm之间的其它波长的吸光度,注意要减去空白对照(稀释液+工作液)的吸光度。
gooddd3. (5)绘制标准曲线,计算样品中的蛋白浓度。
gooddd3. (3)注意:数据处理时需要去除明显错误的值。待测样品浓度可以从标准曲线中查得, 实际浓度需要乘以样品的稀释倍数。如果是计算机绘制的曲线,可以从计算机给出的线性方程式计算出待测样品的浓度。
注意事项
good1. 本产品可以采用酶标仪(微孔检测法)或者分光光度计(试管检测法)测定蛋白浓度,如使用普通的分光光度计测定,需根据比色皿的最小检测体积,适当加大BCA工作液的用量使其不小于最小检测体积,样品和标准品的用量可相应按比例放大。使用分光光度计测定蛋白浓度时,每个试剂盒可以测定的样品数量可能会显著减少;
good2. 试剂在低温条件或长期保存出现沉淀时,可搅拌或37℃温育使其溶解;
good3. 建议每次测定蛋白样品时,都须绘制标准曲线,以获得准确数据;
good4. BSA标准品的稀释液需与待测样品的稀释液一致(可用1×PBS或0.9%生理盐水进行稀释);
good5. 如待测样品中含较多的干扰物质(具体见附表),可采用其它蛋白定量产品;
good6. 为了您的安全和健康,请穿实验服并戴一次性手套操作;
good7. 本产品仅限科研使用。
干扰物质附表
化合物 |
耐受浓度 |
化合物 |
耐受浓度 |
缓冲液 |
去垢剂和变性剂 |
乙酸盐 |
0.2 M |
Brij35 |
1% |
甘氨酸 |
1 M |
CHAPS |
1% |
HEPES |
0.1 M |
盐酸胍 |
4 M |
MES |
50 mM |
NP-40 |
1% |
MOPS |
50 mM |
辛葡糖 |
1% |
柠檬酸钠 |
<1 mM |
SDS |
1% |
PIPES |
50 mM |
Triton X-100 |
1% |
磷酸钠 |
0.1 M |
糖类 |
乙酸钠 |
0.2 M pH5.5 |
葡萄糖 |
10 mM |
TES |
50 mM |
蔗糖 |
1 M |
Tris |
0.1 M |
螯合剂 |
盐类 |
EDTA |
10 mM |
硫酸铵 |
干扰 |
还原剂 |
NaCl |
1 M |
β-巯基乙醇 |
50 μM |
尿素 |
3 M |
DTT |
1 mM |
极性化合物 |
其它 |
DMSO |
5% |
HCl/NaOH |
0.1 M |
甘油 |
10% |
脂类 |
干扰 |
相关论文
good1. Pang Y, Wu D, Ma Y, et al. Reactive oxygen species trigger NF-κB-mediated NLRP3 inflammasome activation involvement in low-dose CdTe QDs exposure-induced hepatotoxicity[J]. Redox biology, 2021, 47: 102157.(IF 11.799)
good2. Zhou Y, Zhao X, Hu W, et al. Acute and subacute oral toxicity of propylene glycol enantiomers in mice and the underlying nephrotoxic mechanism[J]. Environmental Pollution, 2021, 290: 118050.(IF 8.071)
good3.Hua T, Wang H, Fan X, et al. BRD4 Inhibition Attenuates Inflammatory Pain by Ameliorating NLRP3 Inflammasome-Induced Pyroptosis[J]. Frontiers in immunology, 2022, 13: 837977-837977.(IF 7.561)
good4. Cheng, B., Zhang, H., Hu, J., Peng, Y., Yang, J., Liao, X., ... & Lu, H. (2020). The immunotoxicity and neurobehavioral toxicity of zebrafish induced by famoxadone-cymoxanil. Chemosphere, 247, 125870. (IF 7.086)
good5. WANG, Feng, et al. Adrenomedullin 2 improves bone regeneration in type 1 diabetic rats by restoring imbalanced macrophage polarization and impaired osteogenesis. Stem cell research & therapy, 2021, 12.1: 1-15.(IF 6.832)
good6. Chen Q, Yin H, Pu N, et al. Chemokine C‐C motif ligand 21 synergized with programmed death‐ligand 1 blockade restrains tumor growth[J]. Cancer science, 2021, 112(11): 4457-4469.(IF 6.716)
good7. Cong P, Wu T, Huang X, et al. Identification of Role and Clinical Prognostic Value of Target Genes of m6A RNA Methylation Regulators in Glioma[J]. Frontiers in Cell and Developmental Biology, 2021: 2472.(IF 6.684)
good8. Wang F, Wang W, Kong L, et al. Accelerated bone regeneration by Adrenomedullin 2 through improving the coupling of osteogenesis and angiogenesis via β-catenin signaling[J]. Frontiers in Cell and Developmental Biology, 2021, 9.(IF 6.684)
good9. Xiao L, Sun Y, Liu C, et al. Molecular Behavior of HMGB1 in the Cochlea Following Noise Exposure and in vitro[J]. Frontiers in Cell and Developmental Biology, 2021, 9: 350.(IF 6.684)
good10. WANG, Feng, et al. Accelerated Bone Regeneration by Astragaloside IV through Stimulating the Coupling of Osteogenesis and Angiogenesis. International Journal of Biological Sciences, 2021, 17.7: 1821.(IF 6.580)
good11. Tan X, Gong L, Li X, et al. Promethazine inhibits proliferation and promotes apoptosis in colorectal cancer cells by suppressing the PI3K/AKT pathway[J]. Biomedicine & Pharmacotherapy, 2021, 143: 112174.(IF 6.529)
good12. JIANG, Qian, et al. 4-Phenylbutyric acid accelerates rehabilitation of barrier function in intestinal porcine epithelial cell (IPEC-J2) monolayer model. Animal Nutrition, 2021.(IF 6.383)
good13. LIU, Chang, et al. Value of Pyruvate Carboxylase in Thyroid Fine-Needle Aspiration Wash-Out Fluid for Predicting Papillary Thyroid Cancer Lymph Node Metastasis. Frontiers in oncology, 2021, 11: 1625.(IF 6.244)
good14. Wang C, Wang T, Zhang K, et al. Pan-KRAS inhibitors suppress proliferation through feedback regulation in pancreatic ductal adenocarcinoma[J]. Acta Pharmacologica Sinica, 2022: 1-13.(IF 6.150)
good15. LIU, Heze, et al. Prenatal dexamethasone exposure induces nonalcoholic fatty liver disease in male rat offspring via the miR-122/YY1/ACE2-MAS1 pathway. Biochemical Pharmacology, 2021, 185: 114420.(IF 5.858)
good16. Zhang J, Li C, Zhang L, et al. Andrographolide induces noxa-dependent apoptosis by transactivating atf4 in human lung adenocarcinoma cells[J]. Frontiers in pharmacology, 2021, 12: 924.(IF 5.810)
good17. PIAO, Jinmei, et al. Effects of real-ambient PM2. 5 exposure on lung damage modulated by Nrf2. Frontiers in pharmacology, 2021, 12: 913.(IF 5.810)
good18. Wang F, Hou W, Li X, et al. SETD8 cooperates with MZF1 to participate in hyperglycemia-induced endothelial inflammation via elevation of WNT5A levels in diabetic nephropathy[J]. Cellular & Molecular Biology Letters, 2022, 27(1): 1-18.(IF 5.787)
good19. Jiao X, Yan B, Huang J, et al. Redox Proteomic Analysis Reveals Microwave-Induced Oxidation Modifications of Myofibrillar Proteins from Silver Carp (Hypophthalmichthys molitrix)[J]. Journal of Agricultural and Food Chemistry, 2021, 69(33): 9706-9715.(IF 5.279)
good20. Feng, F., Cheng, P., Xu, S., Li, N., Wang, H., Zhang, Y., & Wang, W. (2020). Tanshinone IIA attenuates silica-induced pulmonary fibrosis via Nrf2-mediated inhibition of EMT and TGF-β1/Smad signaling. Chemico-Biological Interactions, 109024.(IF 5.192)