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Solid-state nanopore fabrication by automated controlled breakdown

Nature Protocols volume 15, pages 122–143 (2020)Cite this article

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Abstract

Solid-state nanopores are now well established as single-biomolecule sensors that hold great promise as sensing elements in diagnostic and sequencing applications. However, until recently this promise has been limited by the expensive, labor-intensive, and low-yield methods used to fabricate low-noise and precisely sized pores. To address this problem, we pioneered a low-cost and scalable solid-state nanopore fabrication method, termed controlled breakdown (CBD), which is rapidly becoming the method of choice for fabricating solid-state nanopores. Since its initial development, nanopore research groups around the world have applied and adapted the CBD method in a variety of ways, with varying levels of success. In this work, we present our accumulated knowledge of nanopore fabrication by CBD, including a detailed description of the instrumentation, software, and procedures required to reliably fabricate low-noise and precisely sized solid-state nanopores with a yield of >85% in less than 1 h. The assembly instructions for the various custom instruments can be found in the Supplementary Manual, and take approximately a day to complete, depending on the unit that the user is building and their level of skill with mechanical and electrical assembly. Unlike traditional beam-based nanopore fabrication technologies, the methods presented here are accessible to non-experts, lowering the cost of, and technical barriers to, fabricating nanoscale pores in thin solid-state membranes.

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Fig. 1: Overview of the workflow for nanopore fabrication by CBD and beam-based methods.
Fig. 2: Overview of custom-built hardware.
Fig. 3: Overview of the disposable parts and hardware.
Fig. 4: Stage transition diagram for the nanopore automation algorithm.
Fig. 5: Overview of automated nanopore fabrication and conditioning processes.
Fig. 6: Overview of automated nanopore size and noise measurements.
Fig. 7: Typical pore fabrication and results.
Fig. 8: Results of noise, speed, and precision tests performed on several nanopores fabricated using the presented procedure.
Fig. 9: Results of a typical sensing experiment.

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Data availability

Data for all graphs is available from the corresponding author upon reasonable request.

Code availability

An executable version of the custom software presented (CBD Soft) is freely available for non-commercial uses, as outlined in the End-User License Agreement in the installer.

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Acknowledgements

We would like to acknowledge financial support from the Natural Sciences and Engineering Research Council of Canada (NSERC) i2i and CRD, as well as Ontario Centres of Excellence (OCE) VIP II grant programs. K.B. acknowledges the support of the Vanier Canadian Graduate Scholarship program. The authors would like to thank all members of the Tabard-Cossa and Godin laboratories at the University of Ottawa for their help in testing and troubleshooting the hardware, software, and protocols presented in this work.

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Author notes

  1. These authors contributed equally: Matthew Waugh, Kyle Briggs.

Authors and Affiliations

  1. Department of Physics, University of Ottawa, Ottawa, Ontario, Canada

    Matthew Waugh, Kyle Briggs, Dylan Gunn, Mathieu Gibeault, Simon King, Quinn Ingram, Aura Melissa Jimenez, Samuel Berryman, Dmytro Lomovtsev, Lukasz Andrzejewski & Vincent Tabard-Cossa

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Contributions

K.B. and V.T.-C. developed the CBD method. K.B. and M.W. designed the software protocols. M.G. wrote the LabView code of the software. D.G., M.W., S.K., Q.I., A.M.J., S.B., D.L., and L.A. designed and built the hardware. M.W. and K.B. developed the particular protocols provided. M.W., K.B., and V.T.-C. wrote the manuscript.

Corresponding author

Correspondence to Vincent Tabard-Cossa.

Ethics declarations

Competing interests

V.T.-C. and K.B. hold several patents related to CBD. All other authors have no competing interests.

Additional information

Peer review information Nature Protocols thanks Sébastien Balme and Ralph Scheicher for their contribution to the peer review of this work.

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Supplementary Manual

Component lists, assembly instructions, circuit diagrams and enlarged screenshots of our custom software

Reporting Summary

Supplementary Note 1

Detailed CAD designs and technical drawings

Supplementary Note 2

The PCB layout, in NI Ultiboard file format

Supplementary Software

An executable version of the custom software presented (CBD Soft). Updated versions will be available on our lab’s website (www.tcossalab.net/cbd)

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Waugh, M., Briggs, K., Gunn, D. et al. Solid-state nanopore fabrication by automated controlled breakdown. Nat Protoc 15, 122–143 (2020). https://doi.org/10.1038/s41596-019-0255-2

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