Handen J S, Rosenberg H F
The Laboratory of Host Defenses, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Building 10, Room 11N104, 9000 Rockville Pike, Bethesda, Maryland 20892, USA.
Front Biosci. 1997 Jun 15;2:c9-11. doi: 10.2741/A166.
We have developed a modified Southwestern blotting technique which utilizes broad-spectrum protease inhibitors during nuclear protein extraction and a procedure for radiolabelling an oligonucleotide probe to a high specific activity. These modifications have resulted in minimal protein degradation during nuclear protein isolation and have permitted room temperature hybridizations, improving both the facility and sensitivity of the standard Southwestern assay. This technique was used in our laboratory to visualize NFAT-1 consensus sequence binding proteins in nuclear extracts of human promyelocytic HL-60 cells.
我们开发了一种改良的蛋白质印迹技术，该技术在核蛋白提取过程中使用广谱蛋白酶抑制剂，并采用一种将寡核苷酸探针放射性标记至高比活性的方法。这些改进使得在核蛋白分离过程中蛋白质降解降至最低，并允许在室温下进行杂交，提高了标准蛋白质印迹检测的便利性和灵敏度。我们实验室使用该技术来可视化人早幼粒白血病HL-60细胞的核提取物中与NFAT-1共有序列结合的蛋白质。
