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An Ultrasensitive Electrochemical Immunosensor for Alpha-Fetoprotein Using an Envision Complex-Antibody Copolymer as a Sensitive Label
The State Key Laboratory Base of Novel Functional Materials and Preparation Science, Faculty of Material Science and Chemical Engineering, Ningbo University, Ningbo 315211, China
Department of Laboratory Medicine, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China
* Authors to whom correspondence should be addressed.
Received: 21 August 2012; in revised form: 15 November 2012 / Accepted: 6 December 2012 / Published: 11 December 2012
Abstract: A novel strategy is presented for sensitive detection of alfa-fetoprotein (AFP), using a horseradish peroxidase (HRP)-functionalized Envision antibody complex (EVC) as the label. The Envision-AFP signal antibody copolymer (EVC-AFP Ab2) was composed of a dextran amine skeleton anchoring more than 100 molecules of HRP and 15 molecules of secondary antibody, and acted as a signal tag in the immunosensor. The sensor was constructed using the following steps: First, gold electrode (GE) was modified with nano-gold (AuNPs) by electro-deposition in HAuCl4 solution. The high affinity of the AuNPs surface facilitates direct formation of a self-assembled thiolated protein G layer. Next, the coated GE was incubated in a solution of AFP capture antibody (AFP Ab1); these antibodies attach to the thiolated protein G layer through their non-antigenic regions, leaving the antigen binding sites for binding of target analyte. Following a sandwich immunoreaction, an EVC-AFP Ab2-AFP-AFP Ab1 immunocomplex was formed on the electrode surface, allowing large amounts of HRP on the complex to produce an amplified electrocatalytic current of hydroquinone (HQ) in the presence of hydrogen peroxide (H2O2). Highly amplified detection was achieved, with a detection limit of 2 pg/mL and a linear range of 0.005–0.2 ng/mL for AFP in 10 μL undiluted serum; this is near or below the normal levels of most cancer biomarker proteins in human serum. Measurements of AFP in the serum of cancer patients correlated strongly with standard enzyme-linked immunosorbent assays. These easily fabricated EVC-modified immunosensors show excellent promise for future fabrication of bioelectronic arrays. By varying the target biomolecules, this technique may be easily extended for use with other immunoassays, and thus represents a versatile design route.
Keywords: envision antibody complex; alfa-fetoprotein; signal amplification; electrochemical immunosensor
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Xiong, P.; Gan, N.; Cao, Y.; Hu, F.; Li, T.; Zheng, L. An Ultrasensitive Electrochemical Immunosensor for Alpha-Fetoprotein Using an Envision Complex-Antibody Copolymer as a Sensitive Label. Materials 2012, 5, 2757-2772.
Xiong P, Gan N, Cao Y, Hu F, Li T, Zheng L. An Ultrasensitive Electrochemical Immunosensor for Alpha-Fetoprotein Using an Envision Complex-Antibody Copolymer as a Sensitive Label. Materials. 2012; 5(12):2757-2772.
Xiong, Ping; Gan, Ning; Cao, Yuting; Hu, Futao; Li, Tianhua; Zheng, Lei. 2012. "An Ultrasensitive Electrochemical Immunosensor for Alpha-Fetoprotein Using an Envision Complex-Antibody Copolymer as a Sensitive Label." Materials 5, no. 12: 2757-2772.