iFASP: combining isobaric mass tagging with filter-aided sample preparation.
iFASP: combining isobaric mass tagging with filter-aided sample preparation.
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DOI：
10.1021/pr400032m
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发表时间：
2013-08-02
影响因子：
4.4
通讯作者：
Steen H
中科院分区：
文献类型：
--
作者：
McDowell GS;Gaun A;Steen H
仔细、清洁和受控的质谱蛋白质组学样品制备对于获得可重复和可靠的数据至关重要。这在通过化学同量异位素标记（又名串联质量标记）进行定量蛋白质组学时尤其重要，因为差异标记的样品在样品处理期间相当晚才组合。为了满足定量蛋白质组学对稳健可靠的样品处理的需求，我们在这里描述了iFASP，一种简单的协议，用于将同量异位素质量标记与最近引入的过滤辅助样品制备（FASP）方法相结合。iFASP提供了一种快速、简单和有效的方法，用于获得干净的样品，确保有效的消化，并为定量蛋白质组学实验提供出色的标记产率。我们已经进行了我们的iFASP协议，使用几个高度复杂的非洲爪蟾卵和胚胎裂解物，并比较了标记产率和高置信度的肽鉴定的数量，以标准的溶液中消化和标记协议。虽然两种技术的标记效率都在99+%的范围内，但与相应的非基于fasp的方法相比，iFASP的1%错误发现率鉴定的肽数量和相应的定量肽光谱匹配数量增加了一倍。
Careful, clean and controlled preparation of samples for mass spectrometry proteomics is crucial to obtain reproducible and reliable data. This is especially important when carrying out quantitative proteomics by chemical isobaric labeling (aka tandem mass tagging) since the differentially labeled samples are combined quite late during the sample processing. Addressing this need for robust and reliable sample processing for quantitative proteomics, we describe here iFASP, a simple protocol for combining isobaric mass tagging with the recently introduced Filter-Aided Sample Preparation (FASP) method. iFASP provides a quick, simple and effective method for obtaining clean samples, ensuring efficient digestion and providing excellent labeling yields for quantitative proteomics experiments. We have carried out our iFASP protocol using several highly complex Xenopus laevis egg and embryo lysates and compared the labeling yields and number of high-confidence peptide identifications to a standard in-solution digestion and labeling protocol. Although the labeling efficiency with both techniques is in the 99+% range, the number of peptides identified with a 1% false discovery rate and the corresponding number of quantified peptide spectral matches are as much as doubled with iFASP compared to the corresponding non-FASP-based method.
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影响因子：
7
作者：
Ross, PL;Huang, YLN;Pappin, DJ
通讯作者：
Pappin, DJ
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