[原发性肾病综合征患儿尿液蛋白质组学的预处理方法]
[Pretreatment methods of urine proteomics in children with primary nephrotic syndrome].
作者信息
Wang Qiu-Xia, Wang Jian-Hua, Cao Li, Niu Bo, Wang Guo-Liang, Zhong Ru-Gang
机构信息
Department of Nephrology, Teaching Hospital, Capital Institute of Pediatrics, Peking University, Beijing 100020, China.
出版信息
Zhongguo Dang Dai Er Ke Za Zhi. 2011 Feb;13(2):157-60.
OBJECTIVE
To optimize a pretreatment method of urine proteomics in children with primary nephrotic syndrome.
METHODS
Urine from children with primary nephrotic syndrome was treated in different pH and isolated by cold acetone precipitation for different durations. Then the amounts and kinds of proteins were compared by quantify, SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and two-dimensional electrophoresis (2-DE) in order to optimize a way to deal with urine protein.
RESULTS
Most proteins were obtained at pH 2.7. The amounts of protein precipitated by acetone for 0.5 hr was obviously less than those precipitated for 1 and 2 hrs (P<0.05), while there was no significant difference between the amount of protein precipitated for 1 and for 2 hrs. Protein precipitated by cold acetone for 1 hr at pH 2.7 was selected as the best pretreatment method. Satisfactory 2-DE maps can be acquired.
CONCLUSIONS
Urine protein can be best obtained at pH 2.7 and precipitated by cold acetone for 1 hr.
目的
优化原发性肾病综合征患儿尿蛋白质组学的预处理方法。
方法
对原发性肾病综合征患儿的尿液进行不同pH处理，并采用冷丙酮沉淀法进行不同时长的分离。然后通过定量、十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）和双向电泳（2-DE）比较蛋白质的量和种类，以优化尿蛋白处理方法。
结果
在pH 2.7时可获得最多蛋白质。丙酮沉淀0.5小时沉淀出的蛋白量明显少于沉淀1小时和2小时的（P<0.05），而沉淀1小时和2小时的蛋白量之间无显著差异。选择在pH 2.7时用冷丙酮沉淀1小时作为最佳预处理方法，可获得满意的双向电泳图谱。
结论
在pH 2.7时用冷丙酮沉淀1小时可最佳地获取尿蛋白。
Zotero 插件