Measuring forces from the piconewton to millinewton range is of great importance for the study of living systems from a biophysical perspective. The use of flexible micropipettes as highly sensitive force probes has become established in the biophysical community, advancing our understanding of cellular processes and microbial behavior. The micropipette force sensor (MFS) technique relies on measurement of the forces acting on a force-calibrated, hollow glass micropipette by optically detecting its deflections. The MFS technique covers a wide micro- and mesoscopic regime of detectable forces (tens of piconewtons to millinewtons) and sample sizes (micrometers to millimeters), does not require gluing of the sample to the cantilever, and allows simultaneous optical imaging of the sample throughout the experiment. Here, we provide a detailed protocol describing how to manufacture and calibrate the micropipettes, as well as how to successfully design, perform, and troubleshoot MFS experiments. We exemplify our approach using the model nematode Caenorhabditis elegans, but by following this protocol, a wide variety of living samples, ranging from single cells to multicellular aggregates and millimeter-sized organisms, can be studied in vivo, with a force resolution as low as 10 pN. A skilled (under)graduate student can master the technique in ~1–2 months. The whole protocol takes ~1–2 d to finish.
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3D mechanical characterization of single cells and small organisms using acoustic manipulation and force microscopy
Nature Communications Open Access 10 May 2021
Communications Materials Open Access 15 September 2020
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The data presented in this protocol are available from the corresponding authors upon request.
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M.B. gratefully acknowledges support from the Academy of Finland (Centres of Excellence Programme (2014–2019, grant agreement no. 272361) and the Postdoctoral Researcher Project (grant agreement no. 309237)). O.B. acknowledges funding from the German Research Foundation (DFG) under grant BA3406/2. The authors are deeply grateful to K. Dalnoki-Veress for inspiring discussions and support. R.D. Schulman, C.T. Kreis, M.M. Makowski, T. Böddeker, and Q. Magdelaine are acknowledged for sharing their hands-on experiences and for valuable technical suggestions regarding improvements to the protocol.
The authors declare no competing interests.
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Key reference using this protocol
Backholm, M., Ryu, W. S. & Dalnoki-Veress, K. Proc. Natl Acad. Sci. USA 110, 4528–4533 (2013): http://www.pnas.org/content/110/12/4528
Backholm, M., Kasper, A. K. S., Schulman, R. D., Ryu, W. S. & Dalnoki-Veress, K. Phys. Fluids 27, 091901 (2015): https://aip.scitation.org/doi/10.1063/1.4931795
Kreis, C. T., Le Blay, M., Linne, C., Makowski, M. M. & Bäumchen, O. Nat. Phys. 14, 45–49 (2018): https://www.nature.com/articles/nphys4258
Real-time video from an MFS measurement of a swimming C. elegans nematode (in a 10% (wt/vol) PEO-M9 buffer solution) in which a three-dimensional micropipette is used to measure both lateral and propulsive drag forces.
Matlab code ‘calibration.m’ for determining the pipette spring constant in Calibration Option A.
Matlab code ‘deflection.m’ for analysing the pipette deflection via a cross-correlation approach.
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Backholm, M., Bäumchen, O. Micropipette force sensors for in vivo force measurements on single cells and multicellular microorganisms. Nat Protoc 14, 594–615 (2019). https://doi.org/10.1038/s41596-018-0110-x
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