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PALM imaging and cluster analysis of protein heterogeneity at the cell surface

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2010

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Owen, Dylan M.
Rentero, Carles
Magenau, Astrid
Williamson, David
Rodriguez, Macarena
Gaus, Katharina

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Published

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Journal of Biophotonics. 2010, 3(7), pp. 446-454. ISSN 1864-063X. eISSN 1864-0648. Available under: doi: 10.1002/jbio.200900089

Zusammenfassung

The authors employed photoactivatable localization microscopy (PALM) and direct stochastic optical reconstruction microscopy (dSTORM) imaging and image analysis based on Ripley's K-function to quantify the distribution and heterogeneity of proteins at the cell plasma membrane. The membrane targeting sequence of the N-terminal region of the T cell receptor-pathway kinase Lck fused to the photo-convertible fluorescent protein tdEos (LckN10-tdEos), clusters into sub-100 nm regions which cover approximately 7% of the cell surface. 2-channel PALM imaging of LckN10-tdEos and the N-terminus of the kinase Src (SrcN15-PS-CFP2) are demonstrated. Finally, T cell microclusters at the immune synapse are imaged at super-resolution using dSTORM, showing that conventional TIRF images contain unresolved, small clusters. These methods are generally applicable to other cell and fluorophore systems to quantify 2-D molecular clustering at nanometer scales.

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570 Biowissenschaften, Biologie

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ISO 690OWEN, Dylan M., Carles RENTERO, Jérémie ROSSY, Astrid MAGENAU, David WILLIAMSON, Macarena RODRIGUEZ, Katharina GAUS, 2010. PALM imaging and cluster analysis of protein heterogeneity at the cell surface. In: Journal of Biophotonics. 2010, 3(7), pp. 446-454. ISSN 1864-063X. eISSN 1864-0648. Available under: doi: 10.1002/jbio.200900089
BibTex
@article{Owen2010-07imagi-43213,
  year={2010},
  doi={10.1002/jbio.200900089},
  title={PALM imaging and cluster analysis of protein heterogeneity at the cell surface},
  number={7},
  volume={3},
  issn={1864-063X},
  journal={Journal of Biophotonics},
  pages={446--454},
  author={Owen, Dylan M. and Rentero, Carles and Rossy, Jérémie and Magenau, Astrid and Williamson, David and Rodriguez, Macarena and Gaus, Katharina}
}
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