Vera J C, Rivas C
Instituto de Bioquimica, Universidad Austral de Chile, Valdivia.
Anal Biochem. 1988 Sep;173(2):399-404. doi: 10.1016/0003-2697(88)90206-0.
Proteins blotted on nitrocellulose were stained with either 5-dimethylamino-1-naphthalene-sulfonylchloride (dansyl chloride) or fluorescein isothiocyanate. In both cases the staining procedure can be completed in less than 30 min. The sensitivity for detecting fluorescent-labeled proteins on nitrocellulose was 0.5 ng using a dot test. This was accomplished by transparentizing the nitrocellulose with either immersion oil or toluene. Dansylated proteins were successfully utilized for optimizing the electroblotting procedure. In the presence of 0.2% sodium dodecyl sulfate and 20% methanol the distribution of proteins on the nitrocellulose was an exact replica of the protein pattern seen in the polyacrylamide gel. The fluorescent labeling did not affect the antigenic properties of proteins allowing the subsequent probing with antisera. For this procedure, only one set of samples is needed to obtain accurate photographic records of the gel, the nitrocellulose blot, and the probed blot.
转移至硝酸纤维素膜上的蛋白质用5-二甲基氨基-1-萘磺酰氯（丹磺酰氯）或异硫氰酸荧光素进行染色。在这两种情况下，染色过程均可在30分钟内完成。使用斑点试验检测硝酸纤维素膜上荧光标记蛋白质的灵敏度为0.5纳克。这是通过用浸油或甲苯使硝酸纤维素膜透明化来实现的。丹磺酰化的蛋白质成功用于优化电转印程序。在存在0.2%十二烷基硫酸钠和20%甲醇的情况下，蛋白质在硝酸纤维素膜上的分布与聚丙烯酰胺凝胶中所见的蛋白质条带模式完全一致。荧光标记不影响蛋白质的抗原特性，从而允许随后用抗血清进行检测。对于此程序，仅需一组样品即可获得凝胶、硝酸纤维素膜印迹和检测后的印迹的准确照片记录。
