Abstract
This paper describes the development of a biosensor for the detection of anti-hemagglutinin antibodies against the influenza virus hemagglutinin. The steps of biosensor fabrications are as follows: (i) creation of a mixed layer containing the thiol derivative of dipyrromethene and 4-mercapto-1-butanol, (ii) complexation of Cu(II) ions, (iii) oriented immobilization of the recombinant histidine-tagged hemagglutinin, and (iv) filling free spaces with bovine serum albumin. The interactions between recombinants hemagglutinin from the highly pathogenic avian influenza virus type H5N1 and anti-hemagglutinin H5 monoclonal antibodies were explored with Osteryoung square-wave voltammetry. The biosensor displayed a good detection limit of 2.4 pg/mL, quantification limit of 7.2 pg/mL, and dynamic range from 4.0 to 100.0 pg/mL in buffer. In addition, this analytical device was applied for the detection of antibodies in hen sera from individuals vaccinated and non-vaccinated against the avian influenza virus type H5N1. The limit of detection for the assay was the dilution of sera 1: 7 × 106, which is about 200 times better than the enzyme-linked immunosorbent assay.
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Acknowledgments
The authors are grateful to Violetta Cecuda-Adamczewska and Grażyna Płucienniczak from Institute of Biotechnology and Antibiotics (Warsaw, Poland) for the hybridoma culture producing Mab 6-9-1. This work was supported by the National Centre for Research and Development (NCBiR) under grant no. PBS2/A7/14/2014, grant no. 679/N-BELGIA/2010/0, COST Action CM10005 “Supramolecular Chemistry in Water” and Institute of Animal Reproduction and Food Research, Polish Academy of Sciences, Olsztyn, Poland. Wim Dehaen thanks the University of Leuven, the FWO-Vlaanderen, and the Ministerie voor Wetenschapsbeleid for financial support.
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Jarocka, U., Sawicka, R., Stachyra, A. et al. A biosensor based on electroactive dipyrromethene-Cu(II) layer deposited onto gold electrodes for the detection of antibodies against avian influenza virus type H5N1 in hen sera. Anal Bioanal Chem 407, 7807–7814 (2015). https://doi.org/10.1007/s00216-015-8949-y
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DOI: https://doi.org/10.1007/s00216-015-8949-y