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首页> 外文期刊>Mass Spectrometry Letters >An Automated High Throughput Proteolysis and Desalting Platform for Quantitative Proteomic Analysis
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An Automated High Throughput Proteolysis and Desalting Platform for Quantitative Proteomic Analysis

机译：用于定量蛋白质组学分析的自动化高通量蛋白水解和脱盐平台

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Proteomics for biomarker validation needs high throughput instrumentation to analyze huge set of clinical samples for quantitative and reproducible analysis at a minimum time without manual experimental errors. Sample preparation, a vital step in proteomics plays a major role in identification and quantification of proteins from biological samples. Tryptic digestion a major check point in sample preparation for mass spectrometry based proteomics needs to be more accurate with rapid processing time. The present study focuses on establishing a high throughput automated online system for proteolytic digestion and desalting of proteins from biological samples quantitatively and qualitatively in a reproducible manner. The present study compares online protein digestion and desalting of BSA with conventional off-line (in-solution) method and validated for real time sample for reproducibility. Proteins were identified using SEQUEST data base search engine and the data were quantified using IDEALQ software. The present study shows that the online system capable of handling high throughput samples in 96 well formats carries out protein digestion and peptide desalting efficiently in a reproducible and quantitative manner. Label free quantification showed clear increase of peptide quantities with increase in concentration with much linearity compared to off line method. Hence we would like to suggest that inclusion of this online system in proteomic pipeline will be effective in quantification of proteins in comparative proteomics were the quantification is really very crucial.
机译：用于生物标志物验证的蛋白质组学需要高通量仪器，以便在最短的时间内分析大量临床样品，以进行定量和可重复的分析，而没有人工实验错误。样品制备是蛋白质组学中至关重要的一步，在鉴定和定量生物样品中蛋白质方面起着重要作用。胰蛋白酶消化是基于质谱的蛋白质组学样品制备中的一个主要检查要点，它需要在快速处理时间内更准确。本研究的重点是建立高通量的自动化在线系统，以可再现的方式定量和定性地从生物样品中进行蛋白水解消化和脱盐。本研究比较了常规脱机（溶液中）方法对BSA的在线蛋白质消化和脱盐效果，并验证了实时样品的可重复性。使用SEQUEST数据库搜索引擎识别蛋白质，并使用IDEALQ软件对数据进行定量。本研究表明，能够处理96孔格式高通量样品的在线系统以可再现和定量的方式有效地进行蛋白质消化和肽脱盐。与离线方法相比，无标记物定量显示肽浓度随浓度的增加而明显增加，并且线性很大。因此，我们想建议在蛋白质组学流水线中包含此在线系统将对比较蛋白质组学中的蛋白质定量有效，因为该定量确实非常关键。

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