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Biomedical Microdevices

, 19:11 | Cite as

Yeasts identification in microfluidic devices using peptide nucleic acid fluorescence in situ hybridization (PNA-FISH)

  • André M. Ferreira
  • Daniela Cruz-Moreira
  • Laura Cerqueira
  • João M. Miranda
  • Nuno F. Azevedo
Article

Abstract

Peptide nucleic acid fluorescence in situ hybridization (PNA-FISH) is a highly specific molecular method widely used for microbial identification. Nonetheless, and due to the detection limit of this technique, a time-consuming pre-enrichment step is typically required before identification. In here we have developed a lab-on-a-chip device to concentrate cell suspensions and speed up the identification process in yeasts. The PNA-FISH protocol was optimized to target Saccharomyces cerevisiae, a common yeast that is very relevant for several types of food industries. Then, several coin-sized microfluidic devices with different geometries were developed. Using Computational fluid dynamics (CFD), we modeled the hydrodynamics inside the microchannels and selected the most promising options. SU-8 structures were fabricated based on the selected designs and used to produce polydimethylsiloxane-based microchips by soft lithography. As a result, an integrated approach combining microfluidics and PNA-FISH for the rapid identification of S. cerevisiae was achieved. To improve fluid flow inside microchannels and the PNA-FISH labeling, oxygen plasma treatment was applied to the microfluidic devices and a new methodology to introduce the cell suspension and solutions into the microchannels was devised. A strong PNA-FISH signal was observed in cells trapped inside the microchannels, proving that the proposed methodology works as intended. The microfluidic designs and PNA-FISH procedure described in here should be easily adaptable for detection of other microorganisms of similar size.

Keywords

PNA-Fish Microfluidics Modelling Fluid mechanics Oxygen plasma treatment 

Notes

Acknowledgements

This work was financially supported by: 1) POCI-01-0145-FEDER-006939 (Laboratory for Process Engineering, Environment, Biotechnology and Energy – UID/EQU/00511/2013) funded by the European Regional Development Fund (ERDF), through COMPETE2020 - Programa Operacional Competitividade e Internacionalização (POCI) and by national funds, through FCT - Fundação para a Ciência e a Tecnologia; 2) NORTE‐01‐0145‐FEDER‐000005 – LEPABE-2-ECO-INNOVATION, supported by North Portugal Regional Operational Programme (NORTE 2020), under the Portugal 2020 Partnership Agreement, through the European Regional Development Fund (ERDF) and 3) FCT (Scholarship SFRH/BPD/98525/2013) and Project NanoDiaBac (ENMed/0003/2014).

Compliance with ethical standards

Ethical statements

All authors declare that they have no conflict of interest.

This article does not contain any studies with human participants or animals performed by any of the authors.

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Copyright information

© Springer Science+Business Media New York 2017

Authors and Affiliations

  • André M. Ferreira
    • 1
    • 2
  • Daniela Cruz-Moreira
    • 1
    • 2
  • Laura Cerqueira
    • 1
    • 3
  • João M. Miranda
    • 2
  • Nuno F. Azevedo
    • 1
  1. 1.LEPABE– Laboratory for Process Engineering, Environment, Biotechnology and Energy, Department of Chemical EngineeringFaculty of Engineering of University of PortoPortoPortugal
  2. 2.CEFT–Transport Phenomena Research Center, Department of Chemical EngineeringFaculty of Engineering of University of PortoPortoPortugal
  3. 3.Biomode 2BragaPortugal

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