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1 碧云天网站 微信公众号 碧云天生物技术 /Beyotime Biotechnology 订货热线 : 或 订货 技术咨询 网址 : BCA 蛋白浓度测定试剂盒 ( 增强型 ) 产品编号产品名称包装 P0010S BCA 蛋白浓度测定试剂盒 ( 增强型 ) 200 次 产品简介 : BCA 蛋白浓度测定试剂盒 ( 增强型 ) (Enhanced BCA Protein Assay Kit) 是根据目前世界上最常用的两种蛋白浓度检测方法之一 BCA 法研制而成, 实现了蛋白浓度测定的简单 高稳定性 高灵敏度和高兼容性 和碧云天生产的普通 BCA 蛋白浓度测定试剂盒相比, 灵敏度更高, 检测浓度下限达到 10µg/ml, 最小检测蛋白量达到 0.2µg, 待测样品体积为 1-20µl 和碧云天生产的普通 BCA 蛋白浓度测定试剂盒相比, 显色速度更快, 相同的样品孵育较短时间即可进行吸光度测定 在 µg/ml 浓度范围内有较好的线性关系 本产品从 到 0.5mg/ml 的标准曲线参考图 1 图 1. 本试剂盒蛋白标准曲线的效果图 左图为加入 BCA 工作液后 37ºC 分别孵育 和 120 分钟后的吸光度实测效果图, 右图为 37ºC 孵育 60 分钟时的实际显色效果图 图中数据仅供参考, 实际的检测效果可能会略有不同 BCA 法测定蛋白浓度不受绝大部分样品中的化学物质的影响, 可以兼容样品中高达 5% 的 SDS,5% 的 Triton X-100,5% 的 Tween 但本试剂盒受螯合剂和略高浓度的还原剂的影响, 需确保 EDTA 低于 10mM, 无 EGTA, 二硫苏糖醇 (DTT) 低于 1mM,β- 巯基乙醇 (β-mercaptoethanol) 低于 0.01% 不适用 BCA 法时建议试用碧云天生产的 Bradford 蛋白浓度测定试剂盒 (P0006) BCA 蛋白浓度测定试剂盒 ( 增强型 ) 对样品中各种物质的详细的兼容性和普通的 BCA 蛋白浓度测定试剂盒相同, 请参考碧云天如下网页 : Chart For BCA Kit.pdf 每个试剂盒可以检测 200 个样品 包装清单 : 产品编号 产品名称 包装 P0010S-1 BCA 试剂 A 40ml P0010S-2 BCA 试剂 B 1.2ml P0010S-3 蛋白标准 (BSA) 20mg P0010S-4 蛋白标准配制液 1ml 说明书 1 份 保存条件 : 室温保存 蛋白标准配制成溶液后 -20ºC 冻存 注意事项 : 需酶标仪一台, 测定波长为 nm 之间,562nm 最佳 需 96 孔板 如果没有酶标仪, 也可以使用普通的分光光度计测定, 但测定时, 需根据比色皿的最小检测体积, 适当加大 BCA 工作液的用量使不小于最小检测体积, 样品和标准品的用量可相

2 应按比例放大也可不变 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 如发现样品稀释液或裂解液本身就有较高背景, 请试用碧云天生产的 Bradford 蛋白浓度测定试剂盒 (P0006) 为了加快 BCA 法测定蛋白浓度的速度可以适当用微波炉加热, 但是切勿过热 EDTA 浓度必须小于 10mM, 不兼容 EGTA 不适用 BCA 法时, 请试用碧云天生产的 Bradford 蛋白浓度测定试剂盒 (P0006) 本产品仅限于专业人员的科学研究用, 不得用于临床诊断或治疗, 不得用于食品或药品, 不得存放于普通住宅内 为了您的安全和健康, 请穿实验服并戴一次性手套操作 使用说明 : 1. 蛋白标准品的准备 a. 取 1.2ml 蛋白标准配制液加入到一管蛋白标准 (30mg BSA) 中, 充分溶解后配制成 25mg/ml 的蛋白标准溶液 配制后可立即使用, 也可以 -20ºC 长期保存 b. 取适量 25mg/ml 蛋白标准, 稀释至终浓度为 0.5mg/ml 例如取 20µl 25mg/ml 蛋白标准, 加入 980µl 稀释液即可配制成 0.5mg/ml 蛋白标准 蛋白样品在什么溶液中, 标准品也宜用什么溶液稀释 但是为了简便起见, 也可以用 0.9% NaCl 或 PBS 稀释标准品 稀释后的 0.5mg/ml 蛋白标准可以 -20ºC 长期保存 2. BCA 工作液的配制根据样品数量, 按 50 体积 BCA 试剂 A 加 1 体积 BCA 试剂 B(50:1) 配制适量 BCA 工作液, 充分混匀 例如 5ml BCA 试剂 A 加 100µl BCA 试剂 B, 混匀, 配制成 5.1ml BCA 工作液 BCA 工作液室温 24 小时内稳定 3. 蛋白浓度检测 a. 将标准品按 µl 加到 96 孔板的标准品孔中, 加标准品稀释液补足到 20µl, 相当于标准品浓度分别为 mg/ml b. 加适当体积样品到 96 孔板的样品孔中 如果样品不足 20µl, 加标准品稀释液补足到 20µl 请注意记录样品体积 c. 各孔加入 200µl BCA 工作液,37ºC 放置 分钟 注 : 也可以室温放置 2 小时, 或 60ºC 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 d. 用酶标仪测定 A562, 或 nm 之间的波长的吸光度 e. 根据标准曲线和使用的样品体积计算出样品的蛋白浓度 常见问题 : 1. 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严重干扰 BCA 法测定蛋白浓度的物质, 详细的 BCA 法的兼容性列表请参考碧云天如下网页 : Chart For BCA Kit.pdf 2. 是否每次测定时都需要做标准曲线? 建议每次测定时都做标准曲线 因为 BCA 法测定时颜色会随着时间的延长不断加深, 并且显色反应的速度和温度有关, 所以除非精确控制显色反应的时间和温度, 否则如需精确测定宜每次都做标准曲线 相关产品 : 产品编号 产品名称 包装 P0006 Bradford 蛋白浓度测定试剂盒 1000 次 P0006C Bradford 蛋白浓度测定试剂盒 ( 去垢剂兼容型 ) 800 次 P0007 蛋白标准 (5mg/ml BSA) 1ml P0009 BCA 蛋白浓度测定试剂盒 ( 增强型 ) 5000 次 P0010 BCA 蛋白浓度测定试剂盒 ( 增强型 ) 500 次 P0010S BCA 蛋白浓度测定试剂盒 ( 增强型 ) 200 次 P0011 BCA 蛋白浓度测定试剂盒 5000 次 P0012 BCA 蛋白浓度测定试剂盒 500 次 P0012S BCA 蛋白浓度测定试剂盒 200 次 使用本产品的文献 : 1. Liu MJ, Wang Z, Li HX, Wu RC, Liu YZ, Wu QY. Mitochondrial dysfunction as an early event in the process of apoptosis induced by woodfordin I in human leukemia K562 cells. Toxicol Appl Pharmacol Jan 15;194(2): Wu RC, Chen DF, Liu MJ, Wang Z. Dual effects of cycloheximide on U937 apoptosis induced by its combination with VP-16. Biol Pharm Bull Jul; 27(7): Zheng CH, Gao JQ, Zhang YP, Liang WQ. A protein delivery system: biodegradable alginate-chitosan-poly(lactic-co-glycolic acid) composite microspheres. Biochem Biophys Res Commun Oct 29; 323(4): Wu RC, Wang Z, Liu MJ, Chen DF, Yue XS. Chen and X.-S. Yue.β2- integrins mediate a novel form ofchemoresistance in cycloheximide- induced U937 apoptosis. Cell. Mol. Life Sci. 2004;61(16): Liu MJ, Yue PY, Wang Z, Wong RN. Methyl protodioscin induces G2/M arrest and apoptosis in K562 cells with the hyperpolarization of mitochondria. Cancer Lett. 2005;224: Liu MJ, Wang Z, Ju Y, Wong RN, Wu QY. Diosgenin induces cell cycle arrest and apoptosis in human leukemia K562 cells with the disruption of Ca2+ homeostasis. Cancer Chemother Pharmacol Jan;55(1): Le-feng Zhang, Shuang-qing Peng, Sheng Wang. Influence of lead (Pb2+) on the reactions of in vitro cultured rat aorta to 5-hydroxytryptamine. Toxicology Letters. 2005;159: Wu ZQ, Li M, Chen J, Chi ZQ, Liu JG. Involvement of camp/camp-dependent protein kinase signaling pathway in regulation of Na+,K+-ATPase upon activation of opioid receptors by morphine. Mol Pharmacol Mar;69(3): Yi ZC, Liu YZ, Li HX, Yin Y, Zhuang FY, Fan YB, Wang Z. Tellimagrandin I enhances gap junctional communication and attenuates the tumor phenotype of human cervical carcinoma HeLa cells in vitro. Cancer Lett Oct 8;242(1): / 7 P0010S BCA 蛋白浓度测定试剂盒 ( 增强型 ) / 碧云天 /Beyotime

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