Picoflow Liquid Chromatography-Mass Spectrometry for Ultrasensitive Bottom-Up Proteomics Using 2-μm-i.d. Open Tubular Columns.
Picoflow Liquid Chromatography-Mass Spectrometry for Ultrasensitive Bottom-Up Proteomics Using 2-μm-i.d. Open Tubular Columns.
复制标题
使用 2-μm-i.d. 进行超灵敏自下而上蛋白质组学的 Picoflow 液相色谱-质谱分析
DOI：
10.1021/acs.analchem.9b05639
复制
发表时间：
2020
影响因子：
7.4
通讯作者：
Liu,Shaorong
中科院分区：
文献类型：
--
作者：
Xiang,Piliang;Zhu,Ying;Yang,Yu;Zhao,Zhitao;Williams,SarahM;Moore,RonaldJ;Kelly,RyanT;Smith,RichardD;Liu,Shaorong
在许多应用领域，化学分离和分析的关键目标是最大限度地减少样品量，同时最大限度地获得化学信息。提高测量灵敏度对于蛋白质组学研究尤其重要，尤其是在处理痕量样品和需要多次测量时。已经开发了丰富的技术集合，但由此产生的灵敏度仍然不足以实现对小至单细胞的蛋白质组样品的深入覆盖。在这里，我们结合联合收割机皮升规模的液相色谱（picoLC）与质谱（MS）来解决这个问题。picoLC采用2 μ m-i. d.开放的管状柱，以减少所需的样品输入，大大提高了使用电喷雾电离（ESI）与MS实现的灵敏度。使用这种picoLC-MS系统，我们表明，我们可以可靠地识别1000种蛋白质，仅使用75 pg的胰蛋白酶肽，与最先进的液相色谱（LC）或毛细管电泳（CE）相比，灵敏度提高了10-100倍。MS方法。PicoLC-MS扩展了分离科学的极限，有望成为单细胞蛋白质组学的有力工具。
In many areas of application, key objectives of chemical separation and analysis are to minimize the sample quantity while maximizing the chemical information obtained. Increasing measurement sensitivity is especially critical for proteomics research, especially when processing trace samples and where multiple measurements are desired. A rich collection of technologies has been developed, but the resulting sensitivity remains insufficient for achieving in-depth coverage of proteomic samples as small as single cells. Here, we combine picoliter-scale liquid chromatography (picoLC) with mass spectrometry (MS) to address this issue. The picoLC employs a 2-μm-i.d. open tubular column to reduce the sample input needed to greatly increase the sensitivity achieved using electrospray ionization (ESI) with MS. With this picoLC-MS system, we show that we can identify ∼1000 proteins reliably using only 75 pg of tryptic peptides, representing a 10–100-fold sensitivity improvement compared with the state-of-the-art liquid chromatography (LC) or capillary electrophoresis (CE)-MS methods. PicoLC-MS extends the limit of separation science and is expected to be a powerful tool for single cell proteomics.