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    • 9. 发明申请
    • Acid-labile isotope-coded extractant (ALICE) and its use in quantitative mass spectrometric analysis of protein mixtures
    • 酸不稳定同位素编码提取物(ALICE)及其在蛋白质混合物的定量质谱分析中的应用
    • US20050014278A1
    • 2005-01-20
    • US10919365
    • 2004-08-17
    • Yongchang QiuJack WangRodney Hewick
    • Yongchang QiuJack WangRodney Hewick
    • G01N27/62C07D405/12G01N1/28G01N30/46G01N30/72G01N30/88G01N33/532G01N33/58G01N33/68G01N33/00
    • G01N33/6848C07D405/12G01N30/461G01N30/463G01N30/468G01N30/7233G01N33/532G01N33/58G01N33/6803Y10T436/10Y10T436/105831Y10T436/13Y10T436/24Y10T436/25125B01D15/362B01D15/325
    • The method of the invention provides novel compounds, termed acid-labile isotope-coded extractants (ALICE), for quantitative mass spectrometric analysis of protein mixtures. The compounds contain a thiol-reactive group that is used to capture cysteine-containing peptides from all peptide mixtures, an acid-labile linker, and a non-biological polymer. One of the two acid-labile linkers is isotopically labeled and therefore enables the direct quantitation of peptides/proteins through mass spectrometric analysis. Because no functional proteins are required to capture peptides, a higher percentage of organic solvent can be used to solubilize the peptides, particularly hydrophobic peptides, through the binding, washing and eluting steps, thus permitting much better recover of peptides. Moreover, since the peptides are covalently linked to the non-biological polymer (ALICE), more stringent washing is allowed in order to completely remove non-specifically bound species. Finally, peptides captured by ALICE are readily eluted from the polymer support under mild acidic condition with high yield and permit the direct down stream mass spectrometric analysis without any further sample manipulation. In combination with our novel dual column two dimensional liquid chromatography- mass spectrometry (2D-LC-MS/MS) design, the ALICE procedure proves to a general approach for quantitative mass spectrometric analysis of protein mixtures with better dynamic range and sensitivity.
    • 本发明的方法提供了新的化合物,称为酸不稳定同位素编码的提取物(ALICE),用于蛋白质混合物的定量质谱分析。 该化合物含有硫醇反应性基团,其用于从所有肽混合物,酸不稳定接头和非生物聚合物捕获含半胱氨酸的肽。 两个酸不稳定接头之一是同位素标记的,因此能够通过质谱分析直接定量肽/蛋白质。 由于不需要功能蛋白来捕获肽,所以可以使用较高百分比的有机溶剂来通过结合,洗涤和洗脱步骤来增溶肽,特别是疏水性肽,因此可以更好地恢复肽。 此外,由于肽与非生物聚合物(ALICE)共价连接,所以允许更严格的洗涤以完全除去非特异性结合的物质。 最后,ALICE捕获的肽在温和的酸性条件下以高产率容易地从聚合物载体上洗脱,并允许直接的下游质谱分析,无需进一步的样品操作。 结合我们的双柱二维液相色谱 - 质谱(2D-LC-MS / MS)设计,ALICE方法证明了具有更好的动态范围和灵敏度的蛋白质混合物的定量质谱分析的一般方法。