Kanellis A K, Solomos T, Mattoo A K
Department of Horticulture, University of Maryland, College Park 20742.
Anal Biochem. 1989 May 15;179(1):194-7. doi: 10.1016/0003-2697(89)90224-8.
A method for visualizing acid phosphatase isoenzymes by activity staining on nitrocellulose filters after electroblotting of proteins fractionated on nondenaturing polyacrylamide gels is described. Reproducible results were obtained when 25 mM Tris-192 mM glycine was used as the transfer buffer instead of 0.7% acetic acid, 50 mM sodium acetate, pH 4, or 0.14 M acetic acid--0.35 M beta-alanine, pH 4.3. Dot-blot analysis of banana fruit extracts on nitrocellulose filters revealed that a minimum of 5 x 10(-3) units (nmol p-nitrophenyl phosphate hydrolyzed g-1.h-1) of acid phosphatase activity can be detected. This method can be suitable for screening a large number of biological samples for monitoring acid phosphatase activity.
描述了一种在非变性聚丙烯酰胺凝胶上分离蛋白质后进行电印迹，然后在硝酸纤维素滤膜上通过活性染色来可视化酸性磷酸酶同工酶的方法。当使用25 mM Tris - 192 mM甘氨酸作为转移缓冲液，而不是0.7%乙酸、50 mM乙酸钠（pH 4）或0.14 M乙酸 - 0.35 Mβ-丙氨酸（pH 4.3）时，可获得可重复的结果。对香蕉果实提取物在硝酸纤维素滤膜上进行斑点印迹分析表明，酸性磷酸酶活性的最低检测量为5×10(-3)单位（每克每小时水解对硝基苯磷酸的纳摩尔数）。该方法适用于筛选大量生物样品以监测酸性磷酸酶活性。
