色谱

色谱 ›› 2014, Vol. 32 ›› Issue (11): 1271-1274.DOI: 10.3724/SP.J.1123.2014.09019

• 技术与应用 • 上一篇    下一篇

基于大体积循环进样的低丰度蛋白质富集

张权青1, 张磊2, 高小迪2, 张维冰2, 张庆合1   

  1. 1. 中国计量科学研究院, 北京 100029;
    2. 华东理工大学, 上海市功能性材料化学重点实验室, 上海 200237
  • 收稿日期:2014-09-11 修回日期:2014-09-29 出版日期:2014-11-08 发布日期:2014-10-31
  • 通讯作者: 张庆合,E-mail:Qhzhang204@gmail.com.
  • 基金资助:

    国家重大科学仪器设备开发专项(2012YQ09016703,2012YQ120044);国家自然科学基金重点项目(21235005).

Protein enrichment based on large volume recycling injection

ZHANG Quanqing1, ZHANG Lei2, GAO Xiaodi2, ZHANG Weibing2, ZHANG Qinghe1   

  1. 1. National Institute of Metrology, Beijing 100029, China;
    2. East China University of Science and Technology, Shanghai Key Laboratory of Functional Materials Chemistry, Shanghai 200237, China
  • Received:2014-09-11 Revised:2014-09-29 Online:2014-11-08 Published:2014-10-31

PDF

133

被引次数

1

摘要：

发展了一种大体积循环进样方法,用于富集低丰度蛋白质。在优化的色谱分离条件下,通过增加蛋白质样品的上样体积提高低丰度蛋白质的绝对含量;进一步采用增加样品进样循环次数的方法提高蛋白质的富集效率。以猪肝提取蛋白质为样品,每次上样量500 μL的大体积11次循环进样。根据色谱峰的信号强弱,选择了在原始谱图中看不到色谱峰、有较少小峰和有较多小峰出现的时间段等有代表性的馏分进行研究。在中等极性的组分中,保留时间为11.38 min和12.58 min组分的富集效率分别提高了52倍和61倍,实验结果与理论富集效率相近。所发展的方法为生物蛋白质样品研究提供了一种新的富集制备及检测方法。

关键词: 大体积进样, 蛋白质, 低丰度, 富集, 循环, 猪肝

Abstract:

Biological proteins have great differences in size, hydrophobicity, pH and relative abundance. Especially, the relative contents of high-abundance proteins and low abundance proteins can reach 11 orders of magnitude or more in blood. However, the contents of proteins which have an important impact on the biological function and have related to major diseases are usually very low in body fluids or tissues. So, it is very meaningful to detect the low abundance proteins in biological samples. A method of large volume recycling injection was developed as a new sample enrichment technology in this study. The low absolute contents of proteins in a sample were improved by increasing the volume of sample, and the enrichment efficiency was also increased with the addition of injection times. With the pork liver protein as an example, under the optimized conditions, the representative contents which were no peaks or little peaks and more peaks with low signals were studied. The contents of peaks at the retention times of 11.38 min and 12.58 min were enriched by 11 time injections of 500 μL each time. The enrichment factors respectively reached 52 and 61 which were closed to theoretical values. This method can be used for the enrichment of biological protein samples.

Key words: enrichment, large volume injection, low abundance, pork liver, proteins, recycling

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