Label-free Quantification of Proteins in Single Embryonic Cells with Neural Fate in the Cleavage-Stage Frog (Xenopus laevis) Embryo using Capillary Electrophoresis Electrospray Ionization High-Resolution Mass Spectrometry (CE-ESI-HRMS)
Label-free Quantification of Proteins in Single Embryonic Cells with Neural Fate in the Cleavage-Stage Frog (Xenopus laevis) Embryo using Capillary Electrophoresis Electrospray Ionization High-Resolution Mass Spectrometry (CE-ESI-HRMS)
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DOI：
10.1074/mcp.m115.057760
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发表时间：
2016-08-01
影响因子：
7
通讯作者：
Nemes, Peter
中科院分区：
文献类型：
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作者：
Lombard-Banek, Camille;Reddy, Sushma;Nemes, Peter
单细胞中蛋白质表达的定量有望推进对正常发育的系统水平理解。使用自下而上的蛋白质组学工作流程和多重量化串联质量标签，我们最近证明了相对量化的单个胚胎细胞（卵裂球）在青蛙（非洲爪蟾）胚胎。在这项研究中，我们尽量减少衍生化步骤，以提高分析灵敏度，并使用无标记定量（LFQ）的单个非洲爪蟾细胞。该技术基于MaxLFQ（MaxQuant）定制设计的毛细管电泳微流电喷雾电离高分辨率质谱平台和LFQ。通过明智地定制性能的肽分离，电离，和数据依赖性的采集，我们证明了一个类似的75-阿莫尔（类似于11 nM）的检测和定量下限的蛋白质在复杂的细胞膜。该平台通过测量16 ng蛋白质消化物，或
Quantification of protein expression in single cells promises to advance a systems-level understanding of normal development. Using a bottom-up proteomic workflow and multiplexing quantification by tandem mass tags, we recently demonstrated relative quantification between single embryonic cells (blastomeres) in the frog (Xenopus laevis) embryo. In this study, we minimize derivatization steps to enhance analytical sensitivity and use label-free quantification (LFQ) for single Xenopus cells. The technology builds on a custom-designed capillary electrophoresis microflow-electrospray ionization high-resolution mass spectrometry platform and LFQ by MaxLFQ (MaxQuant). By judiciously tailoring performance to peptide separation, ionization, and data-dependent acquisition, we demonstrate an similar to 75-amol (similar to 11 nM) lower limit of detection and quantification for proteins in complex cell digests. The platform enabled the identification of 438 nonredundant protein groups by measuring 16 ng of protein digest, or