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BBB seminar: Joerg Bewersdorf

Pushing the limits of optical nanoscopy: Imaging whole cells at sub-20-nm 3D resolution and living cells in multiple colors

Joerg Bewersdorf
Department of Cell Biology and Department of Biomedical Engineering, Yale University, New Haven, CT, USA

Optical nanoscopy (super-resolution) techniques such as stimulated emission depletion (STED) microscopy and single-molecule switching (SMS) microscopy (e.g. FPALM, PALM or STORM) utilize either targeted or stochastic switching of fluorescent molecules to achieve ~25 nm spatial resolution – about 10-fold below the diffraction limit [1]. The last years have seen many improvements that make these technologies suitable for a rapidly expanding range of applications.

I will report about two recent breakthroughs that significantly expand the live-cell and 3D imaging capabilities of these techniques:

  • Optimized instrumentation and labeling approaches now allow multicolor live-cell STED microscopy as a general imaging technique [2] and allow the observation of morphological changes on the ~50-nm scale.
  • A new SMS nanoscope using two opposing objectives (4Pi-SMS/iPALM) which takes advantage of adaptive optics and novel data analysis approaches allows whole-cell imaging at sub-20-nm resolution in 3D [3].

I will present the scientific foundation, technical realization and application of these new techniques.

(Joerg Bewersdorf has financial interest in Hamamatsu and Bruker.)

 

References:

[1]   T.J. Gould, S.T. Hess, and J. Bewersdorf “Optical nanoscopy: from acquisition to analysis”, Annu. Rev. Biomed. Eng. 2012; 14:231-54.

[2]   F. Bottanelli, E.B. Kromann, E.S. Allgeyer, R.S. Erdmann, S. Wood Baguley, G. Sirinakis, A. Schepartz, D. Baddeley, D.K. Toomre, J.E. Rothman, and J. Bewersdorf “Two-color live-cell nanoscale imaging of intracellular targets”, Nat. Commun. 2016; 7:10778.

[3]   F. Huang, G. Sirinakis, E.S. Allgeyer, L.K. Schroeder, W.C. Duim, E.B. Kromann, T. Phan, F.E. Rivera-Molina, J.R. Myers, I. Irnov, M. Lessard, Y. Zhang, M.A. Handel, C. Jacobs-Wagner, C.P. Lusk, J.E. Rothman, D.K. Toomre, M.J. Booth, and J. Bewersdorf “Ultra-high resolution 3D imaging of whole cells”, Cell 2016; 166:1028-40.

 

Chairperson: Jaakko Saraste <jaakko.saraste@uib.no>, Department of Biomedicine