高壓輔助in-gel蛋白酶解：定性和定量方面

    胰蛋白酶解仍然是一個(gè)蛋白質(zhì)組學(xué)的基本工作流程。zui近幾年,一些提高蛋白酶解的方法已經(jīng)開(kāi)發(fā)起來(lái)。盡管如此，仍然存在著新蛋白酶解方法的開(kāi)發(fā)空間，以滿足蛋白質(zhì)組學(xué)中定性和定量的需求。另一個(gè)蛋白質(zhì)組學(xué)的組成部分是蛋白質(zhì)的分離作用。所有蛋白質(zhì)組學(xué)中的分離技術(shù)大致可分為三組：凝膠基（gel-based），液相基（liquid phase-based）和混合型液相凝膠基（hybrid liquid phase-gel-based）。zui近，我們制訂了一個(gè)綜合方法: 利用直接目標(biāo)胰蛋白酶解結(jié)合反相液相色譜法（RP-HPLC）分 離蛋白質(zhì)（ 2 ） 。在這里，我們報(bào)告利用壓力循環(huán)技術(shù)（ PCT ）結(jié)合in-gel胰蛋白酶解和評(píng)估利用1DE來(lái)這個(gè)方法應(yīng)用中以1DE分離的標(biāo)記蛋白質(zhì)進(jìn)行定量。 

High pressure-assisted in-gel tryptic digestion: qualitative and quantitative aspects

Tryptic digestion of proteins continues to be a fundamental part of any proteomics workflow. A number of enhanced digestion protocols have been developed in recent years （1,2）. Nonetheless, a need still exists for new digestion approaches that meet the demands of qualitative and quantitative proteomics. Another integral part of proteomics is separation of proteins. All separation techniques in proteomics may be broadly classified under three groups: gel-based, liquid phase-based and hybrid liquid phase-gel- based. Recently we have developed an integrated approach combining RP-HPLC separation of proteins with direct on target tryptic digestion （2）. Here we report an application of pressure cycling technology （PCT） with tryptic in-gel- digestion of proteins and an evaluation of the application of this method to label-free quantitation of proteins separated by 1DE.

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