Efficient Molecular Imprinting Strategy for Quantitative Targeted Proteomics of Human Transferrin Receptor in Depleted Human Serum

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• 作者：Liang Liu ; Ting Zhong ; Qingqing Xu ; Yun Chen
• 刊名：Analytical Chemistry
• 出版年：2015
• 出版时间：November 3, 2015
• 年：2015
• 卷：87
• 期：21
• 页码：10910-10919
• 全文大小：482K
• ISSN：1520-6882

文摘

Soluble transferrin receptor (sTfR) in serum has been suggested as a marker for breast cancer diagnosis, monitoring and treatment. However, sTfR levels in some situations could be far below the limit of quantification (LOQ) of most assays. Thus, an efficient sample pretreatment strategy is required. In this study, molecularly imprinted polymers (MIPs) were developed and coupled with liquid chromatography鈥搕andem mass spectrometry (LC-MS/MS)-based targeted proteomics for sTfR measurement. The key to this effort was that the same surrogate peptide of sTfR (VEYHFLSPYVSPK, VK13) was employed in both the enrichment by MIPs and the quantification by targeted proteomics. Specifically, three peptide templates with different lengths were evaluated for the synthesis of MIPs, and the imprinting conditions were optimized. The characteristics of MIPs, including the adsorption capacity, adsorption kinetics, and binding selectivity, were also investigated. As a result, a 鈭?2-fold enhancement of sensitivity was achieved using MIPs. An LOQ of 200 ng路mL^鈥? was obtained. The intra- and interday precision were <10.7 and 7.8%, respectively. The accuracy was 7.5% at the lower limit of quantification (LLOQ) and <8.4% for the other QC levels. After validation, the assay was applied to determine the sTfR levels in breast cancer patients (n = 20) and healthy volunteers (n = 20) using the standard addition method. The corresponding levels of sTfR were 1.59 卤 0.36 渭g路mL^鈥? (range: 0.96鈥?.34 渭g路mL^鈥?) in the volunteers and 1.82 卤 0.42 渭g路mL^鈥? (range: 0.95鈥?.47 渭g路mL^鈥?) in the patients. This study is among the first to combine MIPs and LC-MS/MS targeted proteomics for protein quantification at the peptide level.