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Immunosensor based on fluorescence quenching matrix of the conducting polymer polypyrrole

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Abstract

In this study, the combination of autofluorescent proteins and fluorescence quenching polymers was shown to be a design which can increase the selectivity and sensitivity of immunosensors. With this objective, the conducting polymer polypyrrole (Ppy) was used as a matrix for immobilization of proteins, which enables biological recognition of the analyte, and as a fluorescence quencher, which increases the selectivity of fluorescence-based detection. In this study, bovine leukemia virus proteins gp51 were immobilized within the Ppy matrix and formed a polymeric layer with affinity for antibodies against protein gp51 (anti-gp51). The anti-gp51 antibodies are present at high levels in the blood serum of cattle infected by bovine leukemia virus. Secondary antibodies labeled with horseradish peroxidase (HRP) were used as specific fluorescent probes for detection of a particular target, because the fluorescence of HRP was readily detectable at the required sensitivity. The Ppy was used as fluorescent background, because its fluorescence was almost undetectable when excited by near UV light at 325 nm. Moreover the Ppy quenched the fluorescence of some fluorescent agents including fluorescein-5(6)-isothiocyanate (fluorescein), rhodamine B, and HRP by almost 100% when these fluorescent agents were adsorbed on the surface of Ppy. It is predicted that Ppy-induced fluorescence quenching could be used in the design of immunosensors to increase selectivity and sensitivity.

 

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Acknowledgement

This work was partially financially supported by a postdoctoral fellowship which is being funded by the European Union Structural Funds project “Postdoctoral Fellowship Implementation in Lithuania” within the framework of the Measure for Enhancing the Mobility of Scholars and Other Researchers and the Promotion of Student Research (VP1-3.1-SMM-01) of the Program of Human Resources Development Action Plan. The authors are grateful to PhD student D. Barauskas-Memenas for support in the editing of the manuscript.

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Authors and Affiliations

  1. Nanotechnas—Centre of Nanotechnology and Material Science, Faculty of Chemistry, Vilnius University, Naugarduko 24, 03225, Vilnius 6, Lithuania

    A. Ramanavicius, N. Ryskevic, Y. Oztekin, A. Kausaite-Minkstimiene, J. Baniukevic, J. Kirlyte, U. Bubniene & A. Ramanaviciene

  2. Institute of Chemistry, State Research Institute Centre for Physical and Technological Sciences, A. Gostauto g. 11, 01108, Vilnius, Lithuania

    A. Ramanavicius

  3. Faculty of Science, Department of Chemistry, Selcuk University, 42070, Konya, Turkey

    Y. Oztekin

  4. Institute of Applied Research, Vilnius University, Sauletekio 9-III, Vilnius, 10222, Lithuania

    N. Ryskevic & S. Jursenas

  5. Laboratory of Immunotechnology, State Research Institute Center for Innovative Medicine, Moletu pl. 29, 08409, Vilnius 21, Lithuania

    A. Ramanaviciene

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  1. A. Ramanavicius
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  2. N. Ryskevic
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Correspondence to A. Ramanavicius.

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Ramanavicius, A., Ryskevic, N., Oztekin, Y. et al. Immunosensor based on fluorescence quenching matrix of the conducting polymer polypyrrole. Anal Bioanal Chem 398, 3105–3113 (2010). https://doi.org/10.1007/s00216-010-4265-8

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  • DOI: https://doi.org/10.1007/s00216-010-4265-8

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