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Continuous-flow bioseparation using microfabricated anisotropic nanofluidic sieving structures

Nature Protocols volume 4, pages 1681–1698 (2009)Cite this article

Abstract

The anisotropic nanofluidic-filter (nanofilter) array (ANA) is a unique molecular-sieving structure for separating biomolecules. In this protocol we describe the fabrication of planar and vertical ANA chips and how to perform continuous-flow bioseparation using them. This protocol is most useful for bioengineers who are interested in developing automated multistep chip-based bioanalysis systems and assumes previous cleanroom microfabrication knowledge. The ANA consists of a two-dimensional periodic nanofilter array, and the designed structural anisotropy of ANA causes different-sized or charged biomolecules to follow distinct trajectories under applied electric fields, leading to efficient continuous-flow separation. Using microfluidic channels surrounding the ANA, the fractionated biomolecule streams are collected and routed to different fluid channels or reservoirs for convenient sample recovery and downstream bioanalysis. The ANA is physically robust and can be reused repeatedly. Compared with the conventional gel-based separation techniques, ANA offers the potential for faster separation, higher throughput and more convenient sample recovery.

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Figure 1: Schematic of the fabrication process for both the planar and vertical ANA devices.
Figure 2: Structure of the microfabricated devices incorporating the ANA structure.
Figure 3: Experimental setup for separation of biomolecules through the ANA.

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Acknowledgements

The authors acknowledge financial support from the National Institute of Health (EB005743), Korea Institute of Science and Technology–Intelligent Microsystems Center (KIST-IMC), and the Singapore-MIT Alliance (SMA-II, CE program). We also thank J. Yoo for his contribution in the experimental setup and H. Bow and S. Reto for helpful discussions. The MIT Microsystems Technology Laboratories is acknowledged for support in microfabrication.

Author information

Author notes

  1. Jianping Fu

    Present address: Current address: Department of Mechanical Engineering, University of Michigan, Ann Arbor, Michigan, USA.,

Authors and Affiliations

  1. Research Laboratories of Electronics, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA

    Jianping Fu

  2. Department of Electrical Engineering and Computer Science, Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA

    Jianping Fu & Jongyoon Han

  3. Department of Mechanical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA

    Pan Mao

Authors
  1. Jianping Fu
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  2. Pan Mao
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Contributions

J.F., P.M. and J.H. conceived and designed the ANA chips. J.F. and P.M. fabricated the ANA chips. J.F. and P.M. designed and performed experiments, and analyzed data. J.F. and P.M. wrote manuscript.

Corresponding author

Correspondence to Jianping Fu.

Supplementary information

Supplementary Figure 1

Cleanroom equipments used for fabrication of the ANA structures (PDF 509 kb)

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Fu, J., Mao, P. & Han, J. Continuous-flow bioseparation using microfabricated anisotropic nanofluidic sieving structures. Nat Protoc 4, 1681–1698 (2009). https://doi.org/10.1038/nprot.2009.176

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