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Catalytic nanocrystalline coordination polymers as an efficient peroxidase mimic for labeling and optical immunoassays

Abstract

We report that nanocrystalline Prussian blue of the type Fe4[Fe(CN)6]3 is a powerful peroxidase mimic for use in labeling of biomolecules. The cubic nanocrystals typically have a diameter of 15 nm and are capable of catalyzing the oxidation of colorless 3,3′,5,5′-tetramethylbenzidine in the presence of H2O2 to form an intensively colored product with an absorption maximum at 662 nm. The determined pseudo turnover number is ~20,000 s−1 which is the highest value reported for nanoparticles of a size comparable to common proteins. We also present a method for the biotinylation of the surface of these nanocrystals, and show their use in competitive bioaffinity based assays of biotin and human serum albumin. The limits of detection are 0.35 and 0.27 μg mL−1, respectively. The results prove the applicability of coordination polymers for signal amplification and also their compatibility with the format of enzyme linked immunosorbent assays.

Nanocrystalline Prussian blue catalyzes the oxidation of colorless 3,3′,5,5′-tetramethylbenzidine to form intensely colored products. Biotinylated cubic Prussian blue nanoparticles (biotin-NPs) facilitated the immunoassay of human serum albumin (HSA) utilizing biotinylated anti-human serum albumin antibody (biotin-IgG).

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Acknowledgments

The authors acknowledge funding by the Program of “Employment of Newly Graduated Doctors of Science for Scientific Excellence” (Grant CZ.1.07/2.3.00/30.0009), cofinanced by the European Social Fund and the state budget of the Czech Republic. The work was also supported by the CEITEC - Central European Institute of Technology (CZ.1.05/1.1.00/02.0068) from the European Regional Development Fund, COST CZ LD15023 from “The Ministry of Education, Youth and Sports” of the Czech Republic and by funds from the Faculty of Medicine of the Masaryk University (MUNI/A/1558/2014). Access to the core facilities of CEITEC CryoEM is acknowledged.

Author contributions

The manuscript was written through contributions of all authors. All authors have given approval to the final version of the manuscript. Veronika Čunderlová and Antonín Hlaváček contributed equally.

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Correspondence to Antonín Hlaváček.

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Čunderlová, V., Hlaváček, A., Horňáková, V. et al. Catalytic nanocrystalline coordination polymers as an efficient peroxidase mimic for labeling and optical immunoassays. Microchim Acta 183, 651–658 (2016). https://doi.org/10.1007/s00604-015-1697-z

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Keywords

  • Enzyme mimic
  • Immunoassay
  • Nanoparticle
  • Prussian blue
  • Biotinylation
  • Transmission electron microscopy