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ÇóÖú ÓÃ×Ô¼ºÖƱ¸µÄRuNHS°´ÕÕÎÄÏ׵ķ½·¨Ó뿹Ìå½áºÏ£¬ÓÃÆÏ¾ÛÌÇÄý½ºÖùÏ´ÍѺóÀíÂÛÉÏÏȳöÀ´µÄÊÇ¿¹Ì壬µ«ÊÇÔÚ400-600nm´¦²»³ö·å£¬ËùÒÔ²»ÖªµÀ¿¹ÌåºÍRuÁ¬ÉÏÁËûÓС£¹òÇó×ö¹ý¸ÃÀàʵÑéµÄС»ï°éÃÇ£¬ÄܸæÖªÏêϸµÄ²Ù×÷·½·¨¡£¡¢ÎҲο¼µÄÎÄÏ×£ºRu tris(2,2'-bipyridine-4,4'-dicarboxylic acid) N-hydroxysuccinimide ester (4). 0.46 g of DCC and 0.238 g of N-hydroxysuccinimide were dissolved in 3 ml of DMF with stirring and cooled in an ice bath. A solution of 0.38 g of Ru tris(2,2'-bipyridine-4,4'-dicarboxylic acid) (2) was added, and the mixture was stirred for a few hours. The formed precipitate was removed by filtration through a syringe filter, and the filtrate containing the active Ru-complex was used for labeling the substrates. The NHS-ester (3) was prepared in analogy to (4). The proteins HSA, IgG, ConA, and Ferritin were obtained from Sigma Chemical Co. (St. Louis, MO) and used without further purification. The proteins (10 mg portions) were labeled by adding a 100-fold molar excess of the Ru-NHS ester in 50 jLI of DMF to 1 ml of stirred protein solution (0.2 M carbonate buffer, pH 8.3-9.1), followed by a 2-6 h incubation and purification of the labeled protein by gel filtration chromatography on Sephadex G-25 or G-50, using 0.1 M PBS, pH 7.2. |
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