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Development of a multichannel flow-through chemiluminescence microarray chip for parallel calibration and detection of pathogenic bacteria

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Abstract

Pathogen detection is important for health and safety reasons. Several outbreaks all over the world have shown the need for rapid, qualitative, quantitative, and, particularly, multianalyte detection systems. Hence, a multichannel flow-through chemiluminescence microarray chip for parallel detection of pathogenic bacteria was developed. The disposable chip made of acrylonitrile–butadiene–styrene (ABS) copolymer was devised as a support for a multiplexed sandwich immunoassay. Calibration and measurement was possible in one experiment, because the developed chip contains six parallel flow-through microchannels. Polyclonal antibodies against the pathogenic bacteria Escherichia coli O157:H7, Salmonella typhimurium, and Legionella pneumophila were immobilized on the chip by microcontact printing in order to use them as specific receptors. Detection of the captured bacteria was carried out by use of specific detection antibodies labelled with biotin and horseradish peroxidase (HRP)–streptavidine conjugates. The enzyme HRP generates chemiluminescence after adding luminol and hydrogen peroxide. This signal was observed by use of a sensitive CCD camera. The limits of detection are 1.8 × 104 cells mL−1 for E. coli O157:H7, 7.9 × 104 cells mL−1 for L. pneumophila, and 2.0 × 107 cells mL−1 for S. typhimurium. The overall assay time for measurement and calibration is 18 min, enabling very fast analysis.

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Acknowledgments

We gratefully thank York Karsunke (Softwarebüro Karsunke, Wolnzach, Germany) for the free license of SIP 0.4. Special thanks to Sebastian Wiesemann for the fabrication of the readout system and the ABS chips. We also thank Dr Sören Schubert from the Max-Pettenkofer-Institut (Munich, Germany) and Dr Rossella Lelli from the Instituto Zooprofilattico Sperimentale Dell’Abruzzo (Italy) for the heat-inactivated bacteria and Adhesive Research Ireland Ltd (Limerick, Ireland) for the adhesive film.

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Authors and Affiliations

  1. Institute of Hydrochemistry and Chair of Analytical Chemistry, Technische Universität München, Marchioninistrasse 17, 81377, München, Germany

    Xaver Y. Z. Karsunke, Reinhard Niessner & Michael Seidel

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  1. Xaver Y. Z. Karsunke
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  2. Reinhard Niessner
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  3. Michael Seidel
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Correspondence to Michael Seidel.

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Karsunke, X.Y.Z., Niessner, R. & Seidel, M. Development of a multichannel flow-through chemiluminescence microarray chip for parallel calibration and detection of pathogenic bacteria. Anal Bioanal Chem 395, 1623–1630 (2009). https://doi.org/10.1007/s00216-009-2905-7

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  • DOI: https://doi.org/10.1007/s00216-009-2905-7

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