Characterization of human plasma lipoproteins using anion exchange fast protein liquid chromatography and targeted mass spectrometry assay.
Characterization of human plasma lipoproteins using anion exchange fast protein liquid chromatography and targeted mass spectrometry assay.
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用阴离子交换快速蛋白液相色谱和靶向质谱法测定人血浆脂蛋白的特性。
作者：
我们描述了一种基于选择反应监测（SRM）的靶向质谱法，用于测定血浆脂蛋白中的5种载脂蛋白（apoA 1、apoB、apoJ、apoD和apoE），该方法通过阴离子交换快速蛋白液相色谱法分离，仅使用100 μL血浆。我们对SRM测定进行了分析表征，并评价了整个工作流程的重现性。apoA 1、apoB、apoD、apoJ和apoE的检测限分别为0.6、4.6、0.8、1.2和0.7 nM;除apoD（4.2 nM）外，所有肽的定量限均为8.3 nM。SRM测定在0.4至1667 nM范围内呈线性。对于所有5种肽，测定的日内和日间以及总重复性（CV%）范围为2.2%至21.7%。对于分级高密度脂蛋白、低密度脂蛋白和IDL中的所有五种肽，整个工作流程的日内和日间以及总再现性范围为12.2%至43.9%。在未来，我们将应用这个工作流程来研究分级血浆脂蛋白与淀粉样蛋白沉积和阿尔茨海默病背景下认知变化的关联。
We described a targeted mass spectrometry assay based on selected reaction monitoring (SRM) for five apolipoproteins (apoA1, apoB, apoJ, apoD, and apoE) in plasma lipoproteins isolated by anion exchange fast protein liquid chromatography using only 100 μL of plasma. We performed analytical characterization of the SRM assay and evaluated reproducibility of the entire workflow. The limit of detections for apoA1, apoB, apoD, apoJ, and apoE were 0.6, 4.6, 0.8, 1.2, and 0.7 nM, respectively; the limit of quantifications was 8.3 nM for all peptides except apoD (4.2 nM). The SRM assay was linear from 0.4 to 1667 nM. The intra-day and inter-day and total repeatability (CV%) of the assay ranged from 2.2% to 21.7% for all five peptides. The intra-day and inter-day and total reproducibility of the entire workflow ranged from 12.2% to 43.9% for all five peptides in fractionated high-density lipoprotein, low-density lipoprotein, and IDL. In the future, we will apply this workflow to investigate the association of fractionated plasma lipoproteins with amyloid deposition and cognitive changes in the context of Alzheimer’s disease.
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