Skip to main content

Digital Microfluidics Assisted Sealing of Individual Magnetic Particles in Femtoliter-Sized Reaction Wells for Single-Molecule Detection

  • Protocol
  • First Online:
Microchip Diagnostics

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1547))

Abstract

Digital microfluidics has emerged in the last years as a promising liquid handling technology for a variety of applications. Here, we describe in detail how to build up an electrowetting-on-dielectric-based digital microfluidic chip with unique advantages for performing single-molecule detection. We illustrate how superparamagnetic particles can be printed with very high loading efficiency (over 98 %) and single-particle resolution in the microwell array patterned in the Teflon-AF® surface of the grounding plate of the chip. Finally, the potential of the device for its application to single-molecule detection is demonstrated by the ultrasensitive detection of the biotinylated enzyme β-Galactosidase captured on streptavidin-coated particles in the described platform.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 89.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 119.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 119.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Rondelez Y, Tresset G, Nakashima T (2005) Highly coupled ATP synthesis by F 1-ATPase single molecules. Nature 433:773–777

    Article  CAS  Google Scholar 

  2. Rissin DM, Walt DR (2006) Digital concentration readout of single enzyme molecules using femtoliter arrays Poisson statistics. Nano Lett 6:520–523

    Google Scholar 

  3. Rissin DM, Walt DR (2006) Digital readout of target binding with attomole detection limits via enzyme amplification in femtoliter arrays. J Am Chem Soc 128:6286–6287

    Article  CAS  Google Scholar 

  4. Rissin D, Kan C, Campbell T (2010) Single-molecule enzyme-linked immunosorbent assay detects serum proteins at subfemtomolar concentrations. Nat Biotechnol. 28:595–589

    Article  CAS  Google Scholar 

  5. Kan CW, Rivnak AJ, Campbell TG, Piech T, Rissin DM, Mösl M et al (2012) Isolationdetection of single molecules on paramagnetic beads using sequential fluid flows in microfabricated polymer array assemblies. Lab Chip 12:977–985

    Article  CAS  Google Scholar 

  6. Zhang H, Nie S, Etson CM, Wang RM, Walt DR (2012) Oil-sealed femtoliter fiber-optic arrays for single molecule analysis. Lab Chip 12:2229–2239

    Article  CAS  Google Scholar 

  7. Kim SH, Iwai S, Araki S, Sakakihara S, Iino R, Noji H (2012) Large-scale femtoliter droplet array for digital counting of single biomolecules. Lab Chip 12:4986–4991

    Article  CAS  Google Scholar 

  8. Chang L, Rissin DM, Fournier DR, Piech T, Patel PP, Wilson DH et al (2012) Single molecule enzyme-linked immunosorbent assays: theoretical considerations. J Immunol Methods 378:102–115

    Article  CAS  Google Scholar 

  9. Witters D, Knez K, Ceyssens F, Puers R, Lammertyn J (2013) Digital microfluidics-enabled single-molecule detection by printing and sealing single magnetic beads in femtoliter droplets. Lab Chip 13:2047–2054

    Article  CAS  Google Scholar 

  10. Pollack MG, AD S, RB F (2002) Electrowetting-based actuation of droplets for integrated microfluidics. Lab Chip 2:96–101

    Article  CAS  Google Scholar 

  11. Fair RB (2007) Digital microfluidics: is a true lab-on-a-chip possible? Microfluid Nanofluid 3:245–281

    Article  CAS  Google Scholar 

  12. Abdelgawad M, Wheeler AR (2009) The Digital Revolution: A New Paradigm for Microfluidics. Adv Mater 21:920–925

    Article  CAS  Google Scholar 

  13. Witters D, Vergauwe N, Vermeir S, Ceyssens F, Liekens S, Puers R et al (2011) Biofunctionalization of electrowetting-on-dielectric digital microfluidic chips for miniaturized cell-based applications. Lab Chip 16:2790–2794

    Article  Google Scholar 

  14. Vergauwe N, Witters D, Atalay YT, Verbruggen B, Vermeir S, Ceyssens F et al (2011) Controlling droplet size variability of a digital lab-on-a-chip for improved bio-assay performance. Microfluid Nanofluid 11:25–34

    Article  CAS  Google Scholar 

  15. Vergauwe N, Witters D, Ceyssens F, Vermeir S, Verbruggen B, Puers R et al (2011) A versatile electrowetting-based digital microfluidic platform for quantitative homogeneous and heterogeneous bio-assays. J Micromechanics Microengineering 21:054026

    Article  Google Scholar 

  16. Kühnemund M, Witters D, Nilsson M, Lammertyn J (2014) Circle-to-circle amplification on a digital microfluidic chip for amplified single molecule detection. Lab Chip 16:2983–2992

    Article  Google Scholar 

  17. KP T, Toffalini F, Witters D, Vermeir S, Rolland F, MLATM H et al (2014) Digital microfluidic chip technology for water permeability measurements on single isolated plant protoplasts. Sensors Actuators B Chem 199:479–487

    Article  Google Scholar 

  18. Witters D, Vergauwe N, Ameloot R, Vermeir S, De Vos D, Puers R et al (2012) Digital microfluidic high-throughput printing of single metal-organic framework crystals. Adv Mater 24:1316–1320

    Article  CAS  Google Scholar 

  19. Tewari KP, Vriens K, Cornaglia M, Gijs M, Kokalj T, Thevissen K et al (2015) Digital microfluidics for time-resolved cytotoxicity studies on single non-adherent yeast cells. Lab Chip 8:1852–1860

    Google Scholar 

  20. Kokalj T, Pérez-Ruiz E, Lammertyn J (2015) Building bio-assays with magnetic particles on a digital microfluidic platform. N Biotechnol 32(5):485–503

    Article  CAS  Google Scholar 

Download references

Acknowledgments

This research was financially supported by the KU Leuven Research Council (IDO-project 10/012, OT project 13/058 and Atheromix IOF-knowledge platform), the Agency for Innovation by Science and Technology in Flanders (IWT project 121615), and the Fund for Scientific Research Flanders—FWO (G.0997.11 and G.0861.14).

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Jeroen Lammertyn .

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2017 Springer Science+Business Media LLC

About this protocol

Cite this protocol

Decrop, D., Ruiz, E.P., Kumar, P.T., Tripodi, L., Kokalj, T., Lammertyn, J. (2017). Digital Microfluidics Assisted Sealing of Individual Magnetic Particles in Femtoliter-Sized Reaction Wells for Single-Molecule Detection. In: Taly, V., Viovy, JL., Descroix, S. (eds) Microchip Diagnostics. Methods in Molecular Biology, vol 1547. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6734-6_7

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-6734-6_7

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-6732-2

  • Online ISBN: 978-1-4939-6734-6

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics