Regenerable immuno-biochip for screening ochratoxin A in green coffee extract using an automated microarray chip reader with chemiluminescence detection.

Ochratoxin A (OTA) can contaminate foodstuffs in the ppb to ppm range and once formed, it is difficult to remove. Because of its toxicity and potential risks to human health, the need exists for rapid, efficient detection methods that comply with legal maximum residual limits. In this work we have synthesized an OTA conjugate functionalized with a water-soluble peptide for covalent immobilization on a glass biochip by means of contact spotting. The chip was used for OTA determination with an indirect competitive immunoassay format with flow-through reagent addition and chemiluminescence detection, carried out with the stand-alone automated Munich Chip Reader 3 (MCR 3) platform. A buffer model and real green coffee extracts were used for this purpose. At the present, covalent conjugate immobilization allowed for at least 20 assay-regeneration cycles of the biochip surface. The total analysis time for a single sample, including measurement and surface regeneration, was 12 min and the LOQ of OTA in green coffee extract was 0.3 μg L(-1) which corresponds to 7 μg kg(-1).

[1]  Miriam M. Ngundi,et al.  Multiplexed detection of mycotoxins in foods with a regenerable array. , 2006, Journal of food protection.

[2]  Reinhard Niessner,et al.  Preparation and characterization of functional poly(ethylene glycol) surfaces for the use of antibody microarrays. , 2007, Analytical chemistry.

[3]  Sarah De Saeger,et al.  Rapid all-in-one three-step immunoassay for non-instrumental detection of ochratoxin A in high-coloured herbs and spices. , 2007, Talanta.

[4]  R. Linhardt,et al.  Poly(ethylene glycol)-based biosensor chip to study heparin-protein interactions. , 2005, Analytical biochemistry.

[5]  Rudolf Krska,et al.  Rapid test strips for analysis of mycotoxins in food and feed , 2009, Analytical and bioanalytical chemistry.

[6]  A. Lehmann,et al.  Determination of ochratoxin A in beer by LC–MS/MS ion trap detection , 2007 .

[7]  S. Saeger,et al.  Development of a new clean-up tandem assay column for the detection of ochratoxin A in roasted coffee , 2005 .

[8]  C. V. Van Peteghem,et al.  Optimization of solid-phase clean-up prior to liquid chromatographic analysis of ochratoxin A in roasted coffee. , 2002, Journal of chromatography. A.

[9]  G. Gauglitz,et al.  Characterisation of morphology of self-assembled PEG monolayers: a comparison of mixed and pure coatings optimised for biosensor applications , 2008, Analytical and bioanalytical chemistry.

[10]  R. Krska,et al.  Advances in the analysis of mycotoxins and its quality assurance , 2005, Food additives and contaminants.

[11]  Stability of Poly(ethylene glycol) Graft Coatings , 1998 .

[12]  A. Pittet,et al.  Liquid Chromatographic Determination of Ochratoxin A in Pure and Adulterated Soluble Coffee Using an Immunoaffinity Column Cleanup Procedure , 1996 .

[13]  Reinhard Niessner,et al.  Development of an open stand-alone platform for regenerable automated microarrays. , 2009, Biosensors & bioelectronics.

[14]  C. V. Van Peteghem,et al.  Application and validation of a clean-up tandem assay column for screening ochratoxin A in cocoa powder , 2007, Food additives and contaminants.

[15]  Juan C. Vidal,et al.  Ochratoxin A nanostructured electrochemical immunosensors based on polyclonal antibodies and gold nanoparticles coupled to the antigen , 2010 .

[16]  Reinhard Niessner,et al.  A regenerable immunochip for the rapid determination of 13 different antibiotics in raw milk. , 2009, The Analyst.

[17]  M. Váradi,et al.  Development of immunosensor based on OWLS technique for determining Aflatoxin B1 and Ochratoxin A. , 2007, Biosensors & bioelectronics.

[18]  F. Palmisano,et al.  Determination of ochratoxin A in green coffee beans by solid-phase microextraction and liquid chromatography with fluorescence detection. , 2008, Journal of chromatography. A.

[19]  Beatriz Prieto-Simón,et al.  Emerging biotools for assessment of mycotoxins in the past decade , 2007 .

[20]  Miriam M. Ngundi,et al.  Array biosensor for detection of ochratoxin A in cereals and beverages. , 2005, Analytical chemistry.

[21]  S. Saeger,et al.  Novel gel-based rapid test for non-instrumental detection of ochratoxin A in beer , 2008, Analytical and bioanalytical chemistry.

[22]  G. Mattson,et al.  A practical approach to crosslinking , 1993, Molecular Biology Reports.

[23]  S. Biselli.  Analytische Methoden für die Kontrolle von Lebens- und Futtermitteln auf Mykotoxine , 2006, Journal für Verbraucherschutz und Lebensmittelsicherheit.

[24]  S. Eremin,et al.  Fluorescence polarization immunoassay based on a monoclonal antibody for the detection of ochratoxin A , 2004 .

[25]  Frances S. Ligler,et al.  Immobilized biomolecules in analysis : a practical approach , 1998 .

[26]  Rudolf Krska,et al.  Mycotoxin analysis: An update , 2008, Food additives & contaminants. Part A, Chemistry, analysis, control, exposure & risk assessment.

[27]  Green Nm,et al.  Avidin and streptavidin. , 1990 .

[28]  B. Liedberg,et al.  A high-density poly(ethylene glycol) polymer brush for immobilization on glass-type surfaces. , 2000, Biosensors & bioelectronics.

[29]  R. Niessner,et al.  Efficient hybridoma screening technique using capture antibody based microarrays. , 2009, Analytical chemistry.

[30]  Reinhard Niessner,et al.  Automated analytical microarrays: a critical review , 2008, Analytical and bioanalytical chemistry.

[31]  A. Zherdev,et al.  Immunochemical methods of mycotoxin analysis (review) , 2010, Applied Biochemistry and Microbiology.

[32]  Linda Monaci,et al.  Determination of ochratoxin A in foods: state-of-the-art and analytical challenges , 2004, Analytical and bioanalytical chemistry.

[33]  S. Piletsky,et al.  Analytical methods for determination of mycotoxins: a review. , 2009, Analytica chimica acta.

[34]  M. Agut,et al.  Analysis of ochratoxin a in coffee by solid-phase cleanup and narrow-bore liquid chromatography-fluorescence detector-mass spectrometry. , 2003, Journal of agricultural and food chemistry.

[35]  M. Zheng,et al.  A Review of Rapid Methods for the Analysis of Mycotoxins , 2006, Mycopathologia.

[36]  F. Breitling,et al.  A novel glass slide-based peptide array support with high functionality resisting non-specific protein adsorption. , 2006, Biomaterials.

[37]  D. Ariza,et al.  Method validation for the determination of ochratoxin A in green and soluble coffee by immunoaffinity column cleanup and liquid chromatography , 2004, Mycotoxin Research.