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Triggering cell detachment from patterned electrode arrays by programmed subcellular release

Nature Protocols volume 5, pages 1273–1280 (2010)Cite this article

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Abstract

Programmed subcellular release is an in vitro technique for the quantitative study of cell detachment. The dynamics of cell contraction are measured by releasing cells from surfaces to which they are attached with spatial and temporal control. Release of subcellular regions of cells is achieved by plating cells on an electrode array created by standard microfabrication methods. The electrodes are then biochemically functionalized with an arginine-glycine-aspartic acid (RGD)-terminated thiol. Application of a voltage pulse results in electrochemical desorption of the RGD-terminated thiols, triggering cell detachment. This method allows for the study of the full cascade of events from detachment to subsequent subcellular reorganization. Fabrication of the electrode arrays may take 1–2 d. Preparation for experiments, including surface functionalization and cell plating, can be completed in 10 h. A series of cell release experiments on one device may last several hours.

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Figure 1: Schematic illustration of the concept of programmed subcellular release.
Figure 2: Experimental setup for programmed subcellular release.
Figure 3: Cyclic voltammograms of electrodes.
Figure 4: Programmed subcellular release.
Figure 5: Immunofluorescence staining of a cell during cell release.
Figure 6: Effect of blebbistatin concentration on cell contraction.
Figure 7: Live cell imaging of Lifeact-GFP actin during release.

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Acknowledgements

This work was supported in part by NIH Grants R21EB008259 and U54CA143868. B.W. acknowledges support from the Achievement Awards for College Scientists (ARCS) Foundation. We thank members of the Wirtz and Searson labs for technical advice and reagents.

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Authors and Affiliations

  1. Department of Materials Science and Engineering, Johns Hopkins University, Baltimore, Maryland, USA

    Bridget Wildt, Denis Wirtz & Peter C Searson

  2. Department of Chemical and Biomolecular Engineering, Johns Hopkins University, Baltimore, Maryland, USA

    Denis Wirtz & Peter C Searson

  3. Johns Hopkins Physical Science Oncology Center and Institute for Nanobiotechnology, Johns Hopkins University, Baltimore, Maryland, USA

    Denis Wirtz & Peter C Searson

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  1. Bridget Wildt
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Contributions

B.W. conducted experiments. B.W., D.W. and P.C.S. designed experiments, analyzed results and wrote the paper.

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Correspondence to Denis Wirtz or Peter C Searson.

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The authors declare no competing financial interests.

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Wildt, B., Wirtz, D. & Searson, P. Triggering cell detachment from patterned electrode arrays by programmed subcellular release. Nat Protoc 5, 1273–1280 (2010). https://doi.org/10.1038/nprot.2010.42

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