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Microfluidic control of cell pairing and fusion

Nature Methods volume 6, pages 147–152 (2009)Cite this article

Abstract

Cell fusion has been used for many different purposes, including generation of hybridomas and reprogramming of somatic cells. The fusion step is the key event in initiation of these procedures. Standard fusion techniques, however, provide poor and random cell contact, leading to low yields. We present here a microfluidic device to trap and properly pair thousands of cells. Using this device, we paired different cell types, including fibroblasts, mouse embryonic stem cells and myeloma cells, achieving pairing efficiencies up to 70%. The device is compatible with both chemical and electrical fusion protocols. We observed that electrical fusion was more efficient than chemical fusion, with membrane reorganization efficiencies of up to 89%. We achieved greater than 50% properly paired and fused cells over the entire device, fivefold greater than with a commercial electrofusion chamber and observed reprogramming in hybrids between mouse embryonic stem cells and mouse embryonic fibroblasts.

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Figure 1: Microfluidic device for cell capture and pairing.
Figure 2: Three-step cell-loading protocol.
Figure 3: Timecourse of chemical– and electric field–induced fusion for different cell pairs.
Figure 4: Comparison of fusion efficiencies.
Figure 5: Functionality of fused cells.

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Acknowledgements

This research was supported by the US National Aeronautics and Space Administration and the National Institutes of Health (EB007278 and EB008550 to JV and 5-R37CA084198 and 5-R01-HDO45022 to R.J.). All microfluidic devices were constructed in the Microsystems Technology Laboratory at MIT. We thank S. Desai (MIT) for providing the eGFP and DsRed mouse 3T3s, R. Foreman (MIT) for providing Oct4-GFP MEFs, and N. Kunst (Fraunhofer Institute) and T. van Boxtel (MIT) for fruitful discussions.

Author information

Author notes

  1. Alison M Skelley and Oktay Kirak: These authors contributed equally to this work.

Authors and Affiliations

  1. Research Laboratory of Electronics, 50 Vassar Street, Massachusetts Institute of Technology (MIT), Cambridge, 02139, Massachusetts, USA

    Alison M Skelley & Joel Voldman

  2. Microsystems Technology Laboratory, 60 Vassar Street, MIT, Cambridge, 02139, Massachusetts, USA

    Alison M Skelley & Joel Voldman

  3. Whitehead Institute for Biomedical Research, Nine Cambridge Center, Cambridge, 02142, Massachusetts, USA

    Oktay Kirak, Heikyung Suh & Rudolf Jaenisch

  4. Department of Biology, MIT, 77 Massachusetts Avenue, Cambridge, 02139, Massachusetts, USA

    Rudolf Jaenisch

  5. Electrical Engineering and Computer Science Department, 77 Massachusetts Avenue, MIT, Cambridge, 02139, Massachusetts, USA

    Joel Voldman

Authors
  1. Alison M Skelley
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  2. Oktay Kirak
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  3. Heikyung Suh
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  4. Rudolf Jaenisch
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  5. Joel Voldman
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Contributions

A.M.S. and O.K. designed, performed and analyzed experiments, and drafted the manuscript. H.S. generated the GFP-positive cells and mice, and performed experiments. R.J. and J.V. participated in experimental design, analyzed data and drafted the manuscript.

Corresponding authors

Correspondence to Rudolf Jaenisch or Joel Voldman.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–10, Supplementary Methods, Supplementary Discussion (PDF 3065 kb)

Supplementary Video 1

Parallel cell transfer. Video of fluorescent 3T3s manually transferred from the backside cup to the front side cup. Images were taken every 250 ms, and the video is playing in real time. (MOV 1165 kb)

Supplementary Video 2

PEG flowing past immobilized 3T3 cells. Images were taken every 100 ms, and the video is playing at 4 frames/s. (MOV 410 kb)

Supplementary Video 3

Fusion of 3T3s with multiple doses of PEG. Images were taken every 2.5 minutes, and the video is playing at 2 frames/s. (MOV 1215 kb)

Supplementary Video 4

Closeup of electrofusion. Video shows fusion of 3T3s with a single electrofusion pulse, including cells swelling in hypoosmolar buffer before fusion pulse and then membrane fusion after media is applied. Images were taken every 2.5 minutes, and the video is playing at 2 frames/s. (MOV 219 kb)

Supplementary Video 5

Electrofusion over entire field. Video shows fusion of 3T3s with a single electrofusion pulse over entire field of view at 10× magnification. Images were taken every 5 minutes, and the video is playing at 2 frames/s. (MOV 1234 kb)

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Skelley, A., Kirak, O., Suh, H. et al. Microfluidic control of cell pairing and fusion. Nat Methods 6, 147–152 (2009). https://doi.org/10.1038/nmeth.1290

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