In recent years, various enzymatic microreactors and on-line enzyme digestion strategies have been widely applied in high throughput proteome analysis. However, the incomplete and irreproducible digestion would introduce some unexpected variations in comparative proteome quantification when the samples are digested and then chemically isotope labeled in different aliquots. To address these problems, we developed an integrated platform for high throughput proteome quantification with combination of on-line low miss-cleavage protein digestion by an ultra-performance immobilized enzymatic reactor, on-line dimethyl labeling onto a C18 precolumn, peptide separation by two-dimensional nano liquid chromatography and MS detection. Compared to traditional off-line method, such a platform exhibits obvious advantages such as high sensitivity, throughput, accuracy, precision and ease of automation. All these results demonstrated that such a platform might become a promising technique for the quantitative proteome analysis.

中文翻译：

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结合在线蛋白质消化、同位素二甲基标记和多维肽分离的集成平台，用于高通量蛋白质组定量

近年来，各种酶促微反应器和在线酶消化策略已广泛应用于高通量蛋白质组分析。然而，当样品被消化，然后在不同的等分试样中进行化学同位素标记时，不完全和不可重复的消化会在比较蛋白质组定量中引入一些意想不到的变化。为了解决这些问题，我们开发了一个用于高通量蛋白质组定量的集成平台，结合了超高性能固定酶反应器的在线低错切蛋白质消化、C18 预柱上的在线二甲基标记、两个肽分离维纳液相色谱和质谱检测。与传统的离线方法相比，这种平台具有明显的优势，如灵敏度高、通量大、准确性、精密度和易于自动化。所有这些结果表明，这样的平台可能成为定量蛋白质组分析的有前途的技术。