Detection and Quantification of Carbohydrate-Deficient Transferrin by MALDI-Compatible Protein Chips Prepared by Ambient Ion Soft Landing
Detection and Quantification of Carbohydrate-Deficient Transferrin by MALDI-Compatible Protein Chips Prepared by Ambient Ion Soft Landing
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DOI：
10.1373/clinchem.2017.285452
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发表时间：
2018-09-01
影响因子：
9.3
通讯作者：
Pompach, Petr
中科院分区：
文献类型：
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作者：
Darebna, Petra;Spicka, Jan;Pompach, Petr
背景：转铁蛋白在肝脏中合成，是人体内最重要的铁转运载体。严重饮酒会导致转铁蛋白糖基化的改变。由于转铁蛋白异构体具有不同的分子质量，质谱仪可以提供快速检测和定量的方法。本研究将抗体芯片与MALDI-TOF MS联用，用于转铁蛋白异构体的检测和定量。方法：利用电喷雾抗转铁蛋白抗体的环境离子软着陆，将氧化铟锡玻璃功能化，制备蛋白质芯片。将两微升患者血清涂抹在抗体修饰的斑点上，经孵育、洗涤和基质沉积后，用MALDI-TOF MS检测转铁蛋白亚型，用每种转铁蛋白形式的峰强度计算总糖缺乏性转铁蛋白(CDT)。结果：芯片MALDI-TOF MS法可用于人血清中CDT的富集和检测。对186例患者的样本队列进行了分析。在这些样本中，44个样本被MALDI芯片方法确定为阳性，而142个样本为阴性。毛细管电泳法与芯片法检测结果的相关系数r=0.986，95%CI。结论：利用环境离子软着陆法制备的抗转铁蛋白抗体修饰的功能化芯片成功地用于人血清中转铁蛋白的检测和定量。(C)2018年美国临床化学协会。
BACKGROUND: Transferrin is synthetized in the liver and is the most important iron-transport carrier in the human body. Severe alcohol consumption leads to alterations in glycosylation of transferrin. Mass spectrometry can provide fast detection and quantification of transferrin isoforms because they have different molecular masses. In this study, we used antibody chips in combination with MALDI-TOF MS for the detection and quantification of transferrin isoforms.METHODS: Protein chips were prepared by functionalization of indium tin oxide glass using ambient ion soft landing of electrosprayed antitransferrin antibody. Two microliters of patient serum was applied on the antibody-modified spots, and after incubation, washing, and matrix deposition, transferrin isoforms were detected by MALDI-TOF MS. Peak intensities of each transferrin form were used to calculate total carbohydrate-deficient transferrin (CDT). The CDT values obtained by the MALDI chip method were compared with the results obtained by a standard capillary electrophoresis (CE).RESULTS: The chip-based MALDI-TOF MS method was used for enrichment and detection of CDT from human serum. A sample cohort from 186 patients was analyzed. Of these samples, 44 were positively identified as belonging to alcoholic patients, whereas 142 were negative by the MALDI chip approach. The correlation of the data obtained by the CE and the chip-based MALDI was r = 0.986, 95% CI.CONCLUSIONS: Functionalized MALDI chips modified by antitransferrin antibody prepared by ambient ion soft landing were successfully used for detection and quantification of CDT from human sera. (c) 2018 American Association for Clinical Chemistry.