Diamandis E P, Christopoulos T K, Bean C C
Department of Clinical Biochemistry, Toronto Western Hospital, Ontario, Canada.
J Immunol Methods. 1992 Mar 4;147(2):251-9. doi: 10.1016/s0022-1759(12)80015-9.
We describe a new method of staining and quantification of proteins blotted or spotted on nitrocellulose. Blotted or spotted proteins are first reacted with specific antibodies followed by reaction with biotinylated secondary antibodies. The immunocomplex is then reacted with a streptavidin-based macromolecular complex labeled with the fluorescent europium chelate of 4,7-bis(chlorosulfophenyl) 1,10-phenanthroline-2,9-dicarboxylic acid (BCPDA). The fluorescent spots or bands can then be assessed by visual inspection under UV illumination, by instant photography or quantified by scanning with a time-resolved fluorometer. The method does not involve enzyme detection, is simple, sensitive and gives sharp bands which remain fluorescent for long periods of time (months to years).
我们描述了一种对硝酸纤维素膜上印迹或点样的蛋白质进行染色和定量的新方法。印迹或点样的蛋白质首先与特异性抗体反应，随后与生物素化二抗反应。然后免疫复合物与基于链霉亲和素的大分子复合物反应，该复合物用4,7-双（氯磺苯基）1,10-菲咯啉-2,9-二羧酸（BCPDA）的荧光铕螯合物标记。然后可通过在紫外光照射下目视检查、即时摄影来评估荧光斑点或条带，或用时间分辨荧光计扫描进行定量。该方法不涉及酶检测，操作简单、灵敏，能产生清晰的条带，且条带能长时间保持荧光（数月至数年）。
