Construction of an electrochemical probe for on chip type flow immunoassay

Miniaturized immunosensors, that combine the analytical power of microfluidic devices with the high specificity of biomolecules, have been intensively studied for clinical analyses, environmental immunoassays and biochemical studies. Protein A has the ability to bind Fc-binding site of immunoglobulin G (IgG), and can be used as the detection probe in immunoassays. Ferrocenecarboaldehyde (Fc-CHO) was labeled to protein A as an electrochemical probe for on chip type amperometric flow immunoassay. When labeling Fc-CHO onto protein A via formation of Schiff-base and its reduction, mean number of nine Fc-CHO labeled to individual protein A molecules. Protein A labeled with Fc-CHO exhibited reversible redox properties of ferrocene in cyclic voltammograms and its peak currents increased with number of labeled ferrocenes. The current response of protein A labeled with Fc-CHO well correlated with its concentration when applying 390 V versus Ag/AgCl. Protein A labeled with Fc-CHO showed sufficient electrochemical properties that are useful in a miniaturized electrochemical immunoassays.

[1]  Joseph Wang,et al.  Microchip‐based amperometric immunoassays using redox tracers , 2002, Electrophoresis.

[2]  Jun Wang,et al.  Amplified voltammetric detection of DNA hybridization via oxidation of ferrocene caps on gold nanoparticle/streptavidin conjugates. , 2003, Analytical chemistry.

[3]  Adam Heller,et al.  Direct electrical communication between chemically modified enzymes and metal electrodes. I. Electron transfer from glucose oxidase to metal electrodes via electron relays, bound covalently to the enzyme , 1987 .

[4]  K R Rogers,et al.  Thick-film electrochemical immunosensor based on stripping potentiometric detection of a metal ion label. , 1998, Analytical chemistry.

[5]  Isao Karube,et al.  Electrochemical protein chip with arrayed immunosensors with antibodies immobilized in a plasma-polymerized film. , 2003, Analytical chemistry.

[6]  D. Chan,et al.  Immunosensors--principles and applications to clinical chemistry. , 2001, Clinica chimica acta; international journal of clinical chemistry.

[7]  T. Matsunaga,et al.  Miniaturized amperometric flow immunoassay system using a glass fiber membrane modified with anion. , 2002, Biotechnology and bioengineering.

[8]  Takehiko Kitamori,et al.  Microchip‐based immunoassay system with branching multichannels for simultaneous determination of interferon‐γ , 2002, Electrophoresis.

[9]  Lydia L. Sohn,et al.  Direct detection of antibody–antigen binding using an on-chip artificial pore , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[10]  H. Kondo,et al.  DNA sensing on a DNA probe-modified electrode using ferrocenylnaphthalene diimide as the electrochemically active ligand. , 2000, Analytical chemistry.

[11]  Adam Heller,et al.  Electrical communication between redox centers of glucose oxidase and electrodes via electrostatically and covalently bound redox polymers , 1989 .

[12]  B Guss,et al.  Complete sequence of the staphylococcal gene encoding protein A. A gene evolved through multiple duplications. , 1984, The Journal of biological chemistry.

[13]  C. Padeste,et al.  Ferrocene-avidin conjugates for bioelectrochemical applications. , 2000, Biosensors & bioelectronics.

[14]  Kyuwon Kim,et al.  Enzyme-amplified electrochemical detection of DNA using electrocatalysis of ferrocenyl-tethered dendrimer. , 2003, Analytical chemistry.

[15]  G. Jin,et al.  A label-free multisensing immunosensor based on imaging ellipsometry. , 2003, Analytical chemistry.

[16]  Chunhai Fan,et al.  Electrochemical interrogation of conformational changes as a reagentless method for the sequence-specific detection of DNA , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[17]  Chen-zhong Li,et al.  A comparison of electron-transfer rates of ferrocenoyl-linked DNA. , 2003, Journal of the American Chemical Society.

[18]  H. Hill,et al.  Homogeneous ferrocene-mediated amperometric immunoassay. , 1986, Analytical chemistry.

[19]  Adam Heller,et al.  Direct electrical communication between chemically modified enzymes and metal electrodes. 2. Methods for bonding electron-transfer relays to glucose oxidase and D-amino-acid oxidase , 1988 .

[20]  A. Lapolla,et al.  Diabetes and mass spectrometry , 2001, Diabetes/metabolism research and reviews.

[21]  Tae-Kyu Lim,et al.  Microfabricated on-chip-type electrochemical flow immunoassay system for the detection of histamine released in whole blood samples. , 2003, Analytical chemistry.

[22]  H. Kondo,et al.  Electrochemically active DNA probes: detection of target DNA sequences at femtomole level by high-performance liquid chromatography with electrochemical detection. , 1994, Analytical biochemistry.