Department of Analytical Chemistry, Faculty of Chemistry, University Complutense of Madrid, 28040 Madrid, Spain.
Biosens Bioelectron. 2014 Feb 15;52:98-104. doi: 10.1016/j.bios.2013.08.021. Epub 2013 Aug 29.
An amperometric immunosensor for the determination of the hormone insulin-like growth factor 1 (IGF1) is reported for the first time in this work. As electrochemical transducer, a multiwalled carbon nanotubes-modified glassy carbon electrode on which poly(pyrrole propionic acid) was electropolymerized was prepared. This approach provided a high content of surface confined carboxyl groups suitable for direct covalent binding of anti-IGF1 monoclonal antibody. A sandwich-type immunoassay using a polyclonal antibody labeled with peroxidase, hydrogen peroxide as the enzyme substrate and catechol as redox mediator was employed to monitor the affinity reaction. All the variables involved in the preparation of the modified electrode were optimized and the electrodes were characterized by electrochemical impedance spectroscopy and cyclic voltammetry. Moreover, the different experimental variables affecting the amperometric response of the immunosensor were also optimized. The calibration graph for IGF1 showed a range of linearity extending from 0.5 to 1000 pg/mL, with a detection limit, 0.25 pg/mL, more than 100 times lower than the lowest values reported for the ELISA immunoassays available for IGF1 (30 pg/mL, approximately). Excellent reproducibility for the measurements carried out with different immunosensors and selectivity against other hormones were also evidenced. A commercial human serum spiked with IGF1 at different levels between 0.01 and 10.0 ng/mL was analyzed with good results.
本文首次报道了一种用于测定激素胰岛素样生长因子 1(IGF1)的安培免疫传感器。作为电化学传感器,制备了一种多壁碳纳米管修饰的玻碳电极,其上电聚合了聚(吡咯丙酸)。这种方法提供了高含量的表面受限的羧基基团,适合于直接共价结合抗 IGF1 单克隆抗体。采用一种用过氧化物酶标记的多克隆抗体、过氧化氢作为酶底物和儿茶酚作为氧化还原介体的三明治型免疫测定法来监测亲和反应。优化了制备修饰电极过程中涉及的所有变量,并通过电化学阻抗谱和循环伏安法对电极进行了表征。此外,还优化了影响免疫传感器安培响应的不同实验变量。IGF1 的校准曲线呈现出从 0.5 到 1000 pg/mL 的线性范围,检测限为 0.25 pg/mL,比现有的 IGF1 ELISA 免疫分析方法(30 pg/mL,约)报告的最低值低 100 多倍。还证明了不同免疫传感器测量的出色重现性和对其他激素的选择性。对含有不同水平 IGF1 的商业人血清(0.01 至 10.0 ng/mL)进行了分析,结果良好。