ELECTROPHORETIC TRANSFER OF PROTEINS FROM POLYACRYLAMIDE GELS TO NITROCELLULOSE SHEETS - PROCEDURE AND SOME APPLICATIONS
ELECTROPHORETIC TRANSFER OF PROTEINS FROM POLYACRYLAMIDE GELS TO NITROCELLULOSE SHEETS - PROCEDURE AND SOME APPLICATIONS
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DOI：
10.1073/pnas.76.9.4350
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发表时间：
1979-01-01
影响因子：
11.1
通讯作者：
GORDON, J
中科院分区：
文献类型：
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作者：
TOWBIN, H;STAEHELIN, T;GORDON, J
设计了一种将蛋白质从聚丙烯酰胺凝胶电泳转移到硝酸纤维素片上的方法。该方法导致核糖体蛋白从含尿素的凝胶中定量转移[来自大肠杆菌和鸡肝]。对于十二烷基硫酸钠凝胶，获得了原始带型，分辨率没有损失，但转移不是定量的。该方法允许通过放射自显影检测蛋白质，并且比常规程序更简单。固定化蛋白可通过免疫学方法检测。硝酸纤维素上所有额外的结合能力都被过量的蛋白质阻断;然后结合特异性抗体，第二抗体针对第一抗体。第二种抗体是放射性标记的或与荧光素或过氧化物酶缀合的。然后分别通过紫外光下的放射自显影或过氧化物酶反应产物检测特异性蛋白质。在后一种情况下，可以清楚地检测到低至100 pg的蛋白质。该程序应适用于分析具有特定反应或配体的各种蛋白质。
A method was devised for the electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets. The method results in quantitative transfer of ribosomal proteins [from Escherichia coli and chicken liver] from gels containing urea. For sodium dodecyl sulfate gels the original band pattern was obtained with no loss of resolution, but the transfer was not quantitative. The method allows detection of proteins by autoradiography and is simpler than conventional procedures. Immobilized proteins were detectable by immunological procedures. All additional binding capacity on the nitrocellulose was blocked with excess protein; then a specific antibody was bound and a 2nd antibody directed against the 1st antibody. The 2nd antibody was either radioactively labeled or conjugated to fluorescein or to peroxidase. The specific protein was then detected by either autoradiography under UV light or by the peroxidase reaction product, respectively. In the latter case, as little as 100 pg of protein was clearly detectable. The procedure should apply to analysis of a wide variety of proteins with specific reactions or ligands.