Department of Chemistry, Capital Normal University, Beijing 100048, China.
Department of Chemistry, Capital Normal University, Beijing 100048, China.
Biosens Bioelectron. 2014 May 15;55:343-9. doi: 10.1016/j.bios.2013.12.037. Epub 2013 Dec 27.
In this work, a novel and sensitive multiplexed immunoassay protocol for simultaneous electrochemical determination of alpha-fetoprotein (AFP) and carcinoembryonic antigen (CEA) was designed using functionalized chitosan composites. The immunosensing platform was prepared via immobilizing capture anti-AFP and anti-CEA on chitosan-Au nanoparticles (AuNPs) through EDC/NHS linking. The signal tags were fabricated by immobilizing electroactive redox probes - Prussian blue (PB) and ferrocenecarboxylic acid (Fc) on chitosan (CHIT), following by absorbing AuNPs to immobilize labeled anti-AFP and anti-CEA, respectively. A sandwich-type immunoassay format was employed for the simultaneous detection of AFP and CEA. The assay was based on the electrochemical oxidation/reduction of the redox species in signal tags, which has a relationship with the concentration of analytes. Experimental results revealed that the multiplexed electrochemical immunoassay enabled the simultaneous monitoring of AFP and CEA with a wide range of 0.05-100 ng mL(-1) for both AFP and CEA. The detection limits (LOD) was 0.03 ng mL(-1) for AFP and 0.02 ng mL(-1) for CEA (S/N=3). The assay results of serum samples with the proposed method were in a good agreement with the reference values from standard ELISA method. And the negligible cross-reactivity between the two analytes makes it possesses potential promise in clinical diagnosis.
在这项工作中，设计了一种新颖而灵敏的基于功能化壳聚糖复合材料的电化学多重免疫分析方法，用于同时电化学测定甲胎蛋白（AFP）和癌胚抗原（CEA）。免疫传感平台通过壳聚糖-Au 纳米粒子（AuNPs）上的 EDC/NHS 连接固定捕获抗 AFP 和抗 CEA 来制备。信号标签通过将电化学活性氧化还原探针普鲁士蓝（PB）和二茂铁羧酸（Fc）固定在壳聚糖（CHIT）上，然后分别吸附 AuNPs 来固定标记的抗 AFP 和抗 CEA 来制备。采用三明治型免疫分析形式同时检测 AFP 和 CEA。该测定法基于信号标签中氧化还原物质的电化学氧化/还原，这与分析物的浓度有关。实验结果表明，该多重电化学免疫分析能够同时监测 AFP 和 CEA，AFP 和 CEA 的检测范围分别为 0.05-100ngmL(-1)。AFP 的检测限（LOD）为 0.03ngmL(-1)，CEA 的检测限（LOD）为 0.02ngmL(-1)（S/N=3）。与标准 ELISA 方法相比，该方法对血清样本的检测结果具有良好的一致性。并且两种分析物之间的交叉反应可以忽略不计，这使其在临床诊断中具有潜在的应用前景。
