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Microfabrication of human organs-on-chips

Nature Protocols volume 8, pages 2135–2157 (2013)Cite this article

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Abstract

'Organs-on-chips' are microengineered biomimetic systems containing microfluidic channels lined by living human cells, which replicate key functional units of living organs to reconstitute integrated human organ-level pathophysiology in vitro. These microdevices can be used to test efficacy and toxicity of drugs and chemicals, and to create in vitro models of human disease. Thus, they potentially represent low-cost alternatives to conventional animal models for pharmaceutical, chemical and environmental applications. Here we describe a protocol for the fabrication, microengineering and operation of these microfluidic organ-on-chip systems. First, microengineering is used to fabricate a multilayered microfluidic device that contains two parallel elastomeric microchannels separated by a thin porous flexible membrane, along with two full-height, hollow vacuum chambers on either side; this requires ∼3.5 d to complete. To create a 'breathing' lung-on-a-chip that mimics the mechanically active alveolar-capillary interface of the living human lung, human alveolar epithelial cells and microvascular endothelial cells are cultured in the microdevice with physiological flow and cyclic suction applied to the side chambers to reproduce rhythmic breathing movements. We describe how this protocol can be easily adapted to develop other human organ chips, such as a gut-on-a-chip lined by human intestinal epithelial cells that experiences peristalsis-like motions and trickling fluid flow. Also, we discuss experimental techniques that can be used to analyze the cells in these organ-on-chip devices.

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Figure 1: Mechanically active organ-on-chip microdevice with compartmentalized 3D microarchitecture.
Figure 2: Fabrication of the upper microchannels of the lung-on-a-chip.
Figure 3: Fabrication of porous PDMS membranes.
Figure 4: Alignment, bonding and chemical etching of the lung-on-a-chip microdevice.
Figure 5: Microfabrication of gut-on-a-chip.
Figure 6: A multilayered 3D microfluidic device for the production of the human breathing lung-on-a-chip.
Figure 7: Production and microfluidic engineering of the alveolar epithelium and microvascular endothelium in the lung-on-a-chip microdevice.
Figure 8: Intestinal epithelial cell culture and spontaneous villus morphogenesis in the gut-on-a-chip microdevice.

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Acknowledgements

We thank D. Levner and C. Hinojosa for their assistance in preparing the protocols for device fabrication. This work was supported by the Wyss Institute for Biologically Inspired Engineering at Harvard University and grants from US National Institutes of Health (NIH) NIEHS (ES016665-01A1) and the NIH Common Fund (U01 NS073474) through the Division of Program Coordination, Planning, and Strategic Initiatives (DPCPSI), Office of the Director, NIH and the US Food and Drug Administration (FDA). Additional funds were provided by the Defense Advanced Research Projects Agency (DARPA) under Cooperative Agreement Number W911NF-12-2-0036, and FDA contract HHSF223201310079C. The content of the information does not necessarily reflect the position or the policy of DARPA or the US Government, and no official endorsement should be inferred.

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Authors and Affiliations

  1. Wyss Institute for Biologically Inspired Engineering at Harvard University, Boston, Massachusetts, USA

    Dongeun Huh, Hyun Jung Kim, Jacob P Fraser, Daniel E Shea, Mohammed Khan, Anthony Bahinski, Geraldine A Hamilton & Donald E Ingber

  2. Department of Bioengineering, University of Pennsylvania, Philadelphia, Pennsylvania, USA

    Dongeun Huh

  3. School of Engineering and Applied Sciences, Harvard University, Cambridge, Massachusetts, USA

    Donald E Ingber

  4. Departments of Pathology & Surgery, Vascular Biology Program, Children's Hospital Boston and Harvard Medical School, Boston, Massachusetts, USA

    Donald E Ingber

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  1. Dongeun Huh
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Contributions

D.H. led development of the lung-on-a-chip, performed experiments, analyzed data and prepared the manuscript. H.J.K. led development of the gut-on-a-chip, performed experiments, analyzed data and contributed to preparation of the manuscript. J.P.F., D.E.S., M.K., A.B. and G.A.H. provided assistance in experiments, data analysis and manuscript preparation. D.E.I. led the organ-on-chip effort, assisted in experimental design and analysis and helped in writing of the manuscript.

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Correspondence to Dongeun Huh or Donald E Ingber.

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Huh, D., Kim, H., Fraser, J. et al. Microfabrication of human organs-on-chips. Nat Protoc 8, 2135–2157 (2013). https://doi.org/10.1038/nprot.2013.137

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