Electrochemical Immunochip Sensor for Aflatoxin M1 Detection

• Published in: Analytical chemistry (Washington) 2009-07, Vol.81 (13), p.5291-5298
• Main Authors: Parker, Charlie O, Lanyon, Yvonne H, Manning, Mary, Arrigan, Damien W. M, Tothill, Ibtisam E
• Format: Article
• Language: English
• Subjects: Aflatoxin M1 - analysis; Analytical chemistry; Animals; Antibodies, Immobilized - chemistry; Antibodies, Immobilized - immunology; Biosensing Techniques - methods; Cattle; Chemistry; Electrochemical methods; Electrochemistry; Enzyme-Linked Immunosorbent Assay - instrumentation; Enzyme-Linked Immunosorbent Assay - methods; Exact sciences and technology; General, instrumentation; Gold - chemistry; Microelectrodes; Milk - chemistry; Miscellaneous
• ISSN: 0003-2700; 1520-6882
• DOI: 10.1021/ac900511e

An investigation into the fabrication, electrochemical characterization, and development of a microelectrode array (MEA) immunosensor for aflatoxin M1 is presented in this paper. Gold MEAs (consisting of 35 microsquare electrodes with 20 μm × 20 μm dimensions and edge-to-edge spacing of 200 μm) together with on-chip reference and counter electrodes were fabricated using standard photolithographic methods. The MEAs were then characterized by cyclic voltammetry, and the behavior of the on-chip electrodes were evaluated. The microarray sensors were assessed for their applicability to the development of an immunosensor for the analysis of aflatoxin M1 directly in milk samples. Following the sensor surface silanization, antibodies were immobilized by cross-linking with 1,4-phenylene diisothiocyanate (PDITC). Surface characterization was conducted by electrochemistry, fluorescence microscopy, scanning electron microscopy (SEM), and atomic force microscopy (AFM). A competitive enzyme linked immunosorbent assay (ELISA) assay format was developed on the microarray electrode surface using the 3,3,5′,5′-tetramethylbenzidine dihyrochloride (TMB)/H2O2 electrochemical detection scheme with horseradish peroxidase (HRP) as the enzyme label. The performance of the assay and the microarray sensor were characterized in pure buffer conditions before applying to the milk samples. With the use of this approach, the detection limit for aflatoxin M1 in milk was estimated to be 8 ng L−1, with a dynamic detection range of 10−100 ng L−1, which meets present legislative limits of 50 ng L−1. The milk interference with the sensor surface was also found to be minimal. These devices show high potential for development of a range of new applications which have previously only been detected using elaborate instrumentation.