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Quantitative optical trapping on single organelles in cell extract

Nature Methods volume 10, pages 68–70 (2013)Cite this article

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Abstract

We have developed an optical trapping method to precisely measure the force generated by motor proteins on single organelles of unknown size in cell extract. This approach, termed VMatch, permits the functional interrogation of native motor complexes. We apply VMatch to measure the force, number and activity of kinesin-1 on motile lipid droplets isolated from the liver of normally fed and food-deprived rats.

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Figure 1: The VMatch method for quantitative optical trapping of cellular organelles.
Figure 2: Motility and force measurement on lipid droplets extracted from rat liver.

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Acknowledgements

We thank A. Rai and R. Jha for discussions and help and K. Verhey (University of Michigan) for kinesin-1 antibody. R.M. acknowledges funding through an International Senior Research Fellowship from the Wellcome Trust, UK (grant WT079214MA).

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Authors and Affiliations

  1. Department of Biological Sciences, Tata Institute of Fundamental Research, Mumbai, India

    Pradeep Barak, Ashim Rai, Priyanka Rai & Roop Mallik

Authors
  1. Pradeep Barak
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  2. Ashim Rai
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  3. Priyanka Rai
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  4. Roop Mallik
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Contributions

R.M. designed the experiments. P.B., A.R., P.R. and R.M. performed the experiments. R.M., P.B. and A.R. analyzed the data and wrote the paper.

Corresponding author

Correspondence to Roop Mallik.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–5 and Supplementary Notes 1–3 (PDF 928 kb)

The microtubule is out of focus and therefore not visible clearly.

The lipid droplet moves towards microtubule plus end. The microtubule minus end was labeled using avidin coated magnetic beads (not visible in this field of view). Movie runs in real-time. Field of view is 10 microns horizontally. (AVI 884 kb)

The polarity label on minus-end of microtubule is not visible in this field of view.

Movie runs in real-time. Field of view is 5 microns horizontally. (AVI 2002 kb)

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Barak, P., Rai, A., Rai, P. et al. Quantitative optical trapping on single organelles in cell extract. Nat Methods 10, 68–70 (2013). https://doi.org/10.1038/nmeth.2287

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