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Microfluidics and Nanofluidics

, Volume 10, Issue 3, pp 685–690 | Cite as

Laser microstructuration of three-dimensional enzyme reactors in microfluidic channels

  • Monica Iosin
  • Teodora Scheul
  • Clément Nizak
  • Olivier Stephan
  • Simion Astilean
  • Patrice Baldeck
Short Communication

Abstract

In this paper, we report on the fabrication of three-dimensional (3D) enzymatic microreactors within polydimethylsiloxane microfluidic channels through a photocrosslinking mechanism mediated by the two-photon absorption process at the focal point of pulse lasers, i.e., a sub-nanosecond Nd:YAG microlaser or a femtosecond Ti:Sapphire laser. This approach allows the building of localized 3D trypsin structures with submicron resolution. The fabrication of two different trypsin structures was successfully demonstrated using Eosin Y and Flavin Adenine Dinucleotide as biological photosensitizers: (i) arrays of 3D cylindrical rows and (ii) 3D woodpile structure. The enzymatic activity of the fabricated structures was evaluated by fluorescence spectroscopy using BODIPY FL casein as fluorogenic substrate. The real time investigation of the peptide cleavage into the microfluidic channel demonstrated that the fabricated trypsin microstructures maintain their catalytic activity. This approach opens up the way to complex multistep enzymatic reactions in well-localized regions of microfluidic devices, with great importance in health screening and biomedical diagnostics.

Keywords

Laser microfabrication Two-photon absorption Microreactors Enzymatic activity Microfluidic channels 

Notes

Acknowledgment

This work was supported by Agence Universitaire de la Francophonie (AUF).

References

  1. Allen R, Nielson R, Wise DD, Shear JB (2005) Catalytic three-dimensional protein architectures. Anal Chem 77:5089–5095CrossRefGoogle Scholar
  2. Banu S, Greenway GM, McCreedy T, Shaddick R (2003) Microfabricated bioreactor chips for immobilised enzyme assays. Anal Chim Acta 486:149–157CrossRefGoogle Scholar
  3. Basu S, Campagnola PJ (2004) Enzymatic activity of alkaline phosphatase inside protein and polymer structures fabricated via multiphoton excitation. Biomacromolecules 5:572–576CrossRefGoogle Scholar
  4. Basu S, Cunningham LP, Pins GD et al (2005) Multiphoton excited fabrication of collagen matrixes cross-linked by a modified benzophenone dimer: bioactivity and enzymatic degradation. Biomacromolecules 6:1465–1474CrossRefGoogle Scholar
  5. Campagnola PJ, Delguidice DM, Epling GA et al (2000) 3-Dimensional submicron polymerization of acrylamide by multiphoton excitation of xanthene dyes. Macromolecules 33:1511–1513CrossRefGoogle Scholar
  6. Gao J, Xu J, Locascio LE, Lee CS (2001) Integrated microfluidic system enabling protein digestion, peptide separation, and protein identification. Anal Chem 73:2648–2655CrossRefGoogle Scholar
  7. Hashimoto M, Kaji H, Kemppinen ME, Nishizawa M (2008) Localized immobilization of proteins onto microstructures within a preassembled microfluidic device. Sens Actuators B 128:545–551CrossRefGoogle Scholar
  8. Heo J, Crooks RM (2005) Microfluidic biosensor based on an array of hydrogel-entrapped enzymes. Anal Chem 77:6843–6851CrossRefGoogle Scholar
  9. Hill RT, Shear JB (2006) Enzyme–nanoparticle functionalization of three-dimensional protein scaffolds. Anal Chem 78:7022–7026CrossRefGoogle Scholar
  10. Holden A, Jung SY, Cremer PS (2004) Patterning enzymes inside microfluidic channels via photoattachment chemistry. Anal Chem 76:1838–1843CrossRefGoogle Scholar
  11. Honda T, Miyazaki M, Yamaguchi Y, Nakamura H, Maeda H (2007) Integrated microreaction system for optical resolution of racemic amino acids. Lab Chip 7:366–372CrossRefGoogle Scholar
  12. Huber DL, Manginell RP, Samara MA, Kim Bl, Bunker BC (2003) Programmed adsorption and release of proteins in a microfluidic device. Science 301:352–354CrossRefGoogle Scholar
  13. Hui AY, Wang G, Lin B, Chan WT (2005) Microwave plasma treatment of polymer surface for irreversible sealing of microfluidic devices. Lab Chip 5:1173–1177CrossRefGoogle Scholar
  14. Iosin M, Stephan O, Astilean S, Dupperay A, Baldeck PL (2007) Microstructuration of protein matrices by laser-induced photochemistry. J Optoel Adv Mat 9:716–720Google Scholar
  15. Kaehr B, Allen R, Javier DJ, Currie J, Shear JB (2004) Guiding neuronal development with in situ microfabrication. PNAS 101:16104–16108CrossRefGoogle Scholar
  16. Koh G, Pishko M (2005) Immobilization of multi-enzyme microreactors inside microfluidic devices. Sens Actuators B 106:335–342CrossRefGoogle Scholar
  17. Křenková J, Foret F (2004) Immobilized microfluidic enzymatic reactors. Electrophoresis 25:3550–3563CrossRefGoogle Scholar
  18. Křenková J, Svec F (2009) Less common applications of monoliths: IV. Recent developments in immobilized enzyme reactors for proteomics and biotechnology. J Sep Sci 32:706–718Google Scholar
  19. Mao H, Yang T, Cremer PS (2002) Design and characterization of immobilized enzymes in microfluidic systems. Anal Chem 74:379–385CrossRefGoogle Scholar
  20. Peterson DS, Rohr T, Svec F, Fréchet JMJ (2002) Enzymatic microreactor-on-a-chip: protein mapping using trypsin immobilized on porous polymer monoliths molded in channels of microfluidic devices. Anal Chem 74:4081–4088CrossRefGoogle Scholar
  21. Pitts JD, Campagnola PJ, Epling GA, Goodman SL (2000) Submicron multiphoton free-form fabrication of proteins and polymers: studies of reaction efficiencies and applications in sustained release. Macromolecules 33:1514–1523CrossRefGoogle Scholar
  22. Pitts JD, Howell AR, Taboada R, Banerjee I, Wang J, Goodman SL, Campagnola PJ (2002) New photoactivators for multiphoton excited three-dimensional submicron cross-linking of proteins: bovine serum albumin and type 1 collagen. Photochem Photobiol 76:135–144CrossRefGoogle Scholar
  23. Sakai-Kato K, Kato M, Toyo’oka T (2002) On-line trypsin-encapsulated enzyme reactor by the sol–gel method integrated into capillary electrophoresis. Anal Chem 74:2943–2949CrossRefGoogle Scholar
  24. Uhlich T, Hubbell JA (1997) In: 23rd Annual meeting of the society for biomaterials transcripts, New Orleans, LAGoogle Scholar
  25. Wang C, Oleschuk R, Ouchen F, Li J, Thibault P, Harrison DJ (2000) Integration of immobilized trypsin bead beds for protein digestion within a microfluidic chip incorporating capillary electrophoresis separations and an electrospray mass spectrometry interface. Rapid Commun Mass Spectrom 14:1377–1383CrossRefGoogle Scholar
  26. Watts P, Haswell SJ (2005) The application of micro reactors for organic synthesis. Chem Soc Rev 34:235–246CrossRefGoogle Scholar
  27. Whitesides GM (2006) The origins and the future of microfluidics. Nature 442:368–373CrossRefGoogle Scholar
  28. Yang T, Jung SY, Mao H, Cremer PS (2001) Fabrication of phospholipid bilayer-coated microchannels for on-chip immunoassays. Anal Chem 73:165–169CrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2010

Authors and Affiliations

  • Monica Iosin
    • 1
    • 2
  • Teodora Scheul
    • 1
    • 2
  • Clément Nizak
    • 2
  • Olivier Stephan
    • 2
  • Simion Astilean
    • 1
  • Patrice Baldeck
    • 2
  1. 1.Nanobiophotonics Laboratory, Institute for Interdisciplinary Experimental ResearchBabes-Bolyai UniversityCluj-NapocaRomania
  2. 2.Laboratoire de Spectrométrie PhysiqueUniversité Joseph FourierSaint Martin d’Hères CedexFrance

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