Toward Systeme International d'Unite-traceable protein quantification: From amino acids to proteins
Toward Systeme International d'Unite-traceable protein quantification: From amino acids to proteins
复制标题
DOI：
10.1016/j.ab.2008.02.010
复制
发表时间：
2008-05-15
影响因子：
2.9
通讯作者：
O'Connor, Gavin
中科院分区：
文献类型：
--
作者：
Burkitt, William I.;Pritchard, Caroline;O'Connor, Gavin
在这里，我们展示了原理证明，即其他蛋白质混合物中蛋白质的绝对浓度可以通过 SI 溯源性进行测量。所使用的方法基于蛋白质的胰蛋白酶消化，然后使用双精确匹配同位素稀释质谱 (IDMS) 对释放的肽进行定量。为了提供对蛋白质浓度测量的完整 SI 可追溯性，我们演示了一种 SI 可追溯肽定量方法，其中使用的肽标准品通过氨基酸分析方法进行定量，该方法结合了双重精确匹配的 IDMS 和已知纯度的氨基酸标准品。因此，蛋白质的浓度是根据胰蛋白酶肽的浓度来确定的，而胰蛋白酶肽又根据氨基酸标准品进行了定量。这使得可以对蛋白质浓度进行完全可追踪的 SI 测量。还讨论了实施此方法时的重要注意事项，并举例说明了这些注意事项如何对测量产生不利影响。 (C) 2008 Elsevier Inc. 保留所有权利。
Here we present a demonstration of the proof of principle that absolute concentration of a protein within a mixture of other proteins can be measured with SI traceability. The method used was based on tryptic digestion of a protein followed by quantification using double exact matching isotope dilution mass spectrometry (IDMS) of the peptides released. To provide full SI traceability to measurements of protein concentration we demonstrated a method of SI traceable peptide quantification in which the peptide standards used were quantified by an amino acid analysis method that incorporated double exact matching IDMS and amino acid standards of known purity. The concentration of the protein was therefore determined based upon the concentration of tryptic peptides, which in turn had been quantified based upon amino acid standards. This allowed fully SI-traceable measurements of protein concentration to be made. Important caveats in the implementation of this approach are also discussed and examples of how these can have detrimental effects on the measurements are shown. (C) 2008 Elsevier Inc. All rights reserved.