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Amperometric immunoassay for the obesity biomarker amylin using a screen printed carbon electrode functionalized with an electropolymerized carboxylated polypyrrole
Microchimica Acta ( IF 5.3 ) Pub Date : 2018-06-09 , DOI: 10.1007/s00604-018-2863-x
Gonzalo Martínez-García , Esther Sánchez-Tirado , Araceli González-Cortés , Paloma Yáñez-Sedeño , José M. Pingarrón

AbstractAmylin (the islet amyloid polypeptide) is a hormone related to adiposity, hunger and satiety. It is co-secreted with insulin from pancreatic B-cells. An amperometric immunosensor is presented here for the determination of amylin. It is making use of a screen printed carbon electrode (SPCE) functionalized with electropolymerized poly(pyrrole propionic acid) (pPPA) with abundant carboxyl groups that facilitate covalent binding of antibody against amylin. A competitive immunoassay was implemented using biotinylated amylin and streptavidin labeled with horse radish peroxidase (HRP-Strept) as the enzymatic tracer. The amperometric detection of H2O2 mediated by hydroquinone was employed as an electrochemical probe to monitor the affinity reaction. The variables involved in the preparation and function of the immunosensor were optimized and the electrodes were characterized by electrochemical impedance spectroscopy and cyclic voltammetry. The calibration graph for amylin, obtained by amperometry at −200 mV vs Ag pseudo-reference electrode, showed a range of linearity extending from 1.0 fg∙mL−1 to 50 pg∙mL−1, with a detection limit of 0.92 fg∙mL−1. This is approximately 7000 times lower than the minimum detectable concentration reported for the ELISA immunoassays available for amylin. The assay has excellent reproducibility and good selectivity over potential interferents. Graphical abstractSchematic of an amperometric competitive immunoassay for the obesity biomarker amylin using a poly(pyrrole propionic acid)-modified screen-printed electrode. The detection limit is 0.92 fg∙mL-1 amylin. The method provides excellent reproducibility for the measurements, good selectivity and successful applicability to human urine and serum samples.

中文翻译:

使用电聚合羧化聚吡咯功能化的丝网印刷碳电极对肥胖生物标志物胰淀素进行电流免疫测定

摘要胰岛淀粉样蛋白(胰岛淀粉样多肽)是一种与肥胖、饥饿和饱腹感相关的激素。它与胰腺 B 细胞的胰岛素共同分泌。这里介绍了一种用于测定胰淀素的电流免疫传感器。它利用丝网印刷碳电极 (SPCE) 功能化的电聚合聚(吡咯丙酸)(pPPA)具有丰富的羧基,促进抗体与胰淀素的共价结合。使用以辣根过氧化物酶 (HRP-Strept) 标记的生物素化胰淀素和链霉亲和素作为酶促示踪剂进行竞争性免疫测定。由氢醌介导的 H2O2 的电流检测被用作电化学探针来监测亲和反应。优化了免疫传感器的制备和功能中涉及的变量,并通过电化学阻抗谱和循环伏安法对电极进行了表征。通过在 -200 mV 与 Ag 伪参比电极的电流分析法获得的胰淀素校准图显示线性范围从 1.0 fg∙mL−1 到 50 pg∙mL−1,检测限为 0.92 fg∙mL -1。这比可用于胰淀素的 ELISA 免疫测定报告的最低可检测浓度低约 7000 倍。该测定对潜在干扰物具有出色的重现性和良好的选择性。图形摘要使用聚(吡咯丙酸)修饰的丝网印刷电极对肥胖生物标志物胰淀素进行安培竞争性免疫测定的示意图。检测限为 0.92 fg∙mL-1 胰淀素。
更新日期:2018-06-09
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