轨道轨道
化学
蛋白质组学
质谱法
选择性反应监测
定量蛋白质组学
三级四极质谱仪
工作流程
蛋白质组
计算生物学
色谱法
计算机科学
串联质谱法
数据库
生物化学
生物
基因
作者
Veronika Vidová,Zdeněk Spáčil
标识
DOI:10.1016/j.aca.2017.01.059
摘要
Mass spectrometry (MS) based proteomics have achieved a near-complete proteome coverage in humans and in several other organisms, producing a wealth of information stored in databases and bioinformatics resources. Recent implementation of selected/multiple reaction monitoring (SRM/MRM) technology in targeted proteomics introduced the possibility of quantitatively follow-up specific protein targets in a hypothesis-driven experiment. In contrast to immunoaffinity-based workflows typically used in biological and clinical research for protein quantification, SRM/MRM is characterized by high selectivity, large capacity for multiplexing (approx. 200 proteins per analysis) and rapid, cost-effective transition from assay development to deployment. The concept of SRM/MRM utilizes triple quadrupole (QqQ) mass analyzer to provide inherent reproducibility, unparalleled sensitivity and selectivity to efficiently differentiate isoforms, post-translational modifications and mutated forms of proteins. SRM-like targeted acquisitions such as parallel reaction monitoring (PRM) are pioneered on high resolution/accurate mass (HR/AM) platforms based on the quadrupole-orbitrap (Q-orbitrap) mass spectrometer. The expansion of HR/AM also caused development in data independent acquisition (DIA). This review presents a step-by-step tutorial on development of SRM/MRM protein assay intended for researchers without prior experience in proteomics. We discus practical aspects of SRM-based quantitative proteomics workflow, summarize milestones in basic biological and medical research as well as recent trends and emerging techniques.
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