Name: fast and sensitive colloidal coomassie g-250 staining for proteins in polyacrylamide gels
Uploaded: 2009-08-03T00:00:00
Description: Watch this Scientific Journal Video about Fast and Sensitive Colloidal Coomassie G-250 Staining for Proteins in Polyacrylamide Gels at JoVE.com

我们感谢为准备和染色1 - D凝胶Nicola博士Wiethölter。 这项工作是由来自德意志研究联合会（GRK 1089/project 5 ND和SM）的资助，由来自于尔根Manchot的研究奖学金基金会于Nd支持。

第1部分：二维（2 - D）凝胶电泳，用杯装<ol><li> IPG -带补液等电聚焦（IEF） <ul><li>在125μL补液盐补充水分Immobiline DryStrip凝胶，pH值6-11（7厘米）[7 M尿素，硫脲2米，4％CHAPS，50毫米hydroxyethyldisulfide和2％IPG缓冲液的pH值6-11]采用Immobiline DryStrip Reswelling托盘至少10小时。 </li><li>在IEF样品缓冲液溶解沉淀蛋白质样品[7 M尿素，硫脲2米，2％CHAPS，2％的ASB - 14，50 mM的hydroxyethyldisulfide和2％IPG缓冲液的pH值6-11]对?...

<ol>
	<li>Fazekas de St Groth, S., Webster, S. R. G., Datyner, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Two+new+staining+procedures+for+quantitative+estimation+of+proteins+on+electrophoretic+strips.">Two new staining procedures for quantitative estimation of proteins on electrophoretic strips.</a> Biochim. Biophys. Acta. 71, 377-391 (1963).</li><li>Neuhoff, V., Arold, N., Taube, D., Ehrhardt, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Improved+staining+of+proteins+in+polyacrylamide+gels+including+isoelectric+focusing+gels+with+clear+background+at+nanogram+sensitivity+using+Coomassie+Brilliant+Blue+G-250+and+R-250.">Improved staining of proteins in polyacrylamide gels including isoelectric focusing gels with clear background at nanogram sensitivity using Coomassie Brilliant Blue G-250 and R-250.</a> Electrophoresis. 9, 255-262 (1988).</li><li>Candiano, G., Bruschi, M., Musante, L., Santucci, L., Ghiggeri, G. M., Carnemolla, B., Orecchia, P., Zardi, L., Rigetti, P. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Blue+Silver:+A+very+sensitive+colloidal+Coomassie+G-250+staining+for+proteome+analysis.">Blue Silver: A very sensitive colloidal Coomassie G-250 staining for proteome analysis.</a> Electrophoresis. 25, 1327-1333 (2004).</li><li>Kang, D., Gho, S. G., Suh, M., Kang, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Highly+Sensitive+and+Fast+Protein+Detection+with+Coomassie+Brilliant+Blue+in+Sodium+Dodecyl+Sulfate-Polyacrylamide+Gel+Electrophoresis.">Highly Sensitive and Fast Protein Detection with Coomassie Brilliant Blue in Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis.</a> Bull. Korean Chem. Soc. 11, 1511-1512 (2002).</li><li>Shevchenko, A., Wilm, M., Vorm, O., Mann, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mass+spectrometric+sequencing+of+proteins+silver-stained+polyacrylamide+gels.">Mass spectrometric sequencing of proteins silver-stained polyacrylamide gels.</a> Anal. Chem. 68, 850-858 (1996).</li></ol>

创新或只是另一种考马斯亮协议？ 目前存在与考马斯亮蓝染色程序的多个协议。他们大多导致轻微或重大修改Neuhoff和同事2最常用的协议之一。康有为的协议的基础上Neuhoff的公式。但它真的蛋白质组学研究的一个替代的考马斯染色的方法吗？我们将图片的两个主要问题，检测限和可用性，证据表明，它绝对是一个有益的染色协议相比其他考马斯亮协议，甚至银色和荧...

<table border="1" ><tr><td> 产品名称 </td><td> 公司 </td><td> 目录号 </td><td> 评论 </td></tr><tr><td> 对于IEF和SDS - PAGE电泳： </td><td></td><td></td><td></td></tr><tr><td> Immobiline DryStrip凝胶</td><td> GE医疗集团</td><td> 17-6001-94 </td><td>甚至可以使用2年后到期日期。 </td></tr><tr><td> Immobiline DryStrip Reswelling托盘</td><td> GE医疗集团</td><td> 80-6371-84 </td><td>不要用有机溶剂清洗。专为7-18厘米IPG条。 </td></tr><tr><td> Immobiline DryStrip套件</td><td> GE医疗集团</td><td> 18-1004-30 </td><td>包括一个托盘，电极支架，阳极和阴极电极，对准和样品杯酒吧和样品杯。 </td></tr><tr><td>每股盈利3501 XL电源</td><td> GE医疗集团</td><td> 18-1130-05 </td><td>供应电压高达3500 V。 </td></tr><tr><td> Multiphor第二电泳单位</td><td> GE医疗集团</td><td> 18-1018-06 </td><td>移动电极使国际盛事基金在所有长度的IPG胶条（7-24厘米IPG胶条） </td></tr><tr><td> PerfectBlue凝胶体系双人小号</td><td> Peqlab </td><td> 45 - 1010 - C </td><td> SDS - PAGE电泳在10X10厘米的微型胶格式。凝胶室包括一个冷却系统。 </td></tr><tr><td> 胶体考马斯亮蓝G - 250染色： </td><td></td><td></td><td></td></tr><tr><td>染色与盖菜</td><td> VWR </td><td> 216-3412 </td><td>适合用于小型凝胶。摇床可堆叠。 </td></tr><tr><td>硫酸铝- 18 -水合物</td><td>默克公司</td><td> 1.01102.5000 </td><td>我们最好与默克公司的经验。刚上市的5公斤包装。 </td></tr><tr><td>磷酸</td><td> Prolabo </td><td> 20 624.295 </td><td>玻璃瓶出售。 </td></tr></table>

fast and sensitive colloidal coomassie g-250 staining for proteins in polyacrylamide gels

考马斯亮蓝（CBB）是常用的SDS - PAGE分离蛋白的可视化，提供一个简单的染色程序和高定量的染料。此外，它是完全兼容与质谱蛋白质鉴定。但尽管有这些优势，CBB被认为是比银或荧光染色法和较不敏感，因此很少用于分析凝胶为基础的蛋白质组学方法检测蛋白质。 已作出了一些改进原来的考马斯协议 1增加CBB灵敏度。两个重大修改，以提高转换成胶体粒子的染料分子的低丰度蛋白质的检测：在1988年，Neuhoff和他的同事们应用到的CBB G - 250基于染色溶液2 20％甲醇和硫酸铵的浓度较高，和2004年Candiano等。建立了蓝银在硫酸铵和甲醇 3在场的CBB G - 250使用磷酸。然而，所有这些修改，只允许约10毫微克蛋白质的检测。一个广泛琐细的协议，胶体考马斯染色康有为等人在2002年发表了他们关于修改Neuhoff胶体CBB染色协议的络合物质。取而代之的硫酸铵，他们用硫酸铝和甲醇毒性较低的乙醇 4取代。的新型铝为基础的染色，在康有为的研究中表现出卓越的灵敏度，可检测低为1毫微克/乐队（磷酸二）与灵敏度变化不大，对蛋白质而定。 在这里，我们展示了康有为的快速和敏感的胶体考马斯亮蓝染色分析目的蛋白的协议的应用程序。我们将说明使用二维凝胶经常在我们的工作小组进行快速和容易的协议。

Watch this Scientific Journal Video about Fast and Sensitive Colloidal Coomassie G-250 Staining for Proteins in Polyacrylamide Gels at JoVE.com

Fast and Sensitive Colloidal Coomassie G-250 Staining for Proteins in Polyacrylamide Gels

该视频应普及胶体考马斯亮蓝G - 250染色协议，根据康<em&gt;等。</em平均4凝胶吴蛋白质检测。染色完成后2小时内，没有任何努力。我们经常使用凝胶为基础的蛋白质组学分析的目的在康有为的协议。

快速，灵敏的胶体考马斯亮G - 250染色蛋白聚丙烯酰胺凝胶

Coomassie Brilliant Blue (CBB) is a dye commonly used for the visualization of proteins separated by SDS-PAGE, offering a simple staining procedure and ...

Research

JoVE Journal

Biology

Staining Proteins in Gels

在凝胶染色蛋白质

Following separation by electrophoretic methods, proteins in a gel can be detected by several staining methods. This unit describes protocols for ...

科研

生物学

Method for Whole Mount Antibody Staining in Chick

整装在鸡的抗体染色方法

The chick embryo is a valuable tool in the study of early embryonic development. Its transparency, accessibility and ease of manipulation, make it an ...

Staining of Proteins in Gels with Coomassie G-250 without Organic Solvent and Acetic Acid

无有机溶剂和醋酸，蛋白质与考马斯亮蓝G - 250凝胶染色

In classical protein staining protocols using Coomassie Brilliant Blue (CBB), solutions with high contents of toxic and flammable organic solvents ...

Denaturing Urea Polyacrylamide Gel Electrophoresis (Urea PAGE)

Denaturing Urea Polyacrylamide Gel Electrophoresis Urea PAGE

变性尿素聚丙烯酰胺凝胶电泳（尿素页）

Urea PAGE or denaturing urea polyacrylamide gel electrophoresis employs 6-8 M urea, which denatures secondary DNA or RNA structures and is used for their ...

Measuring Fast Calcium Fluxes in Cardiomyocytes

测量心肌细胞的快速钙助焊剂

Cardiomyocytes have multiple Ca2+ fluxes of varying duration that work together to optimize function 1,2. Changes in Ca2+ activity in response to ...

A Strategy for Sensitive, Large Scale Quantitative Metabolomics

敏感，大规模定量代谢组学的策略

Metabolite profiling has been a valuable asset in the study of metabolism in health and disease. However, current platforms have different limiting ...

Development of a Colloidal Gold-based Immunochromatographic Test Strip for Detection of Cetacean Myoglobin

对肌红蛋白鲸类检测基于金免疫层析胶体试纸条的研制

This protocol describes the development of a colloidal gold immunochromatographic test strip based on the sandwich format that can be used to ...

A Fast Air-dry Dropping Chromosome Preparation Method Suitable for FISH in Plants

快速风干滴染色体制备方法适用于FISH在植物

Preparation of chromosome spreads is a prerequisite for the successful performance of fluorescence in situ hybridization (FISH). Preparation of high ...

Detection of Modified Forms of Cytosine Using Sensitive Immunohistochemistry

胞嘧啶修饰形式使用敏感免疫组化检测

Methylation of cytosine bases (5-methylcytosine, 5mC) occurring in vertebrate genomes is usually associated with transcriptional silencing. ...

Quantitative Proteomics Workflow using Multiple Reaction Monitoring Based Detection of Proteins from Human Brain Tissue

使用基于多反应监测的人类脑组织蛋白质的多反应监测工作流程定量蛋白质组学工作流程

The proteomic analysis of the human brain tissue over the last decade has greatly enhanced our understanding of the brain. However, brain related ...