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Proceedings Paper

Mapping molecular orientational distributions for biological sample in 3D (Conference Presentation)
Author(s): Wei HE; Patrick Ferrand; Benjamin Richter; Martin Bastmeyer; Sophie Brasselet

Paper Abstract

Measuring molecular orientation properties is very appealing for scientists in molecular and cell biology, as well as biomedical research. Orientational organization at the molecular scale is indeed an important brick to cells and tissues morphology, mechanics, functions and pathologies. Recent work has shown that polarized fluorescence imaging, based on excitation polarization tuning in the sample plane, is able to probe molecular orientational order in biological samples; however this applies only to information in 2D, projected in the sample plane. To surpass this limitation, we extended this approach to excitation polarization tuning in 3D. The principle is based on the decomposition of any arbitrary 3D linear excitation in a polarization along the longitudinal z-axis, and a polarization in the transverse xy–sample plane. We designed an interferometer with one arm generating radial polarization light (thus producing longitudinal polarization under high numerical aperture focusing), the other arm controlling a linear polarization in the transverse plane. The amplitude ratio between the two arms can vary so as to get any linear polarized excitation in 3D at the focus of a high NA objective. This technique has been characterized by polarimetry imaging at the back focal plane of the focusing objective, and modeled theoretically. 3D polarized fluorescence microscopy is demonstrated on actin stress fibers in non-flat cells suspended on synthetic polymer structures forming supporting pillars, for which heterogeneous actin orientational order could be identified. This technique shows a great potential in structural investigations in 3D biological systems, such as cell spheroids and tissues.

Paper Details

Date Published: 27 April 2016
PDF: 1 pages
Proc. SPIE 9711, Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues IX, 97111M (27 April 2016); doi: 10.1117/12.2211078
Show Author Affiliations
Wei HE, Institut Fresnel (France)
Cell and Neurobiology Institute, Karlsruhe Institute of Technology (Germany)
Patrick Ferrand, Institute Fresnel (France)
Benjamin Richter, Cell and Neurobiology Institute, Karlsruhe Institute of Technology (Germany)
Martin Bastmeyer, Cell and Neurobiology Institute, Karlsruhe Institute of Technology (Germany)
Sophie Brasselet, Institute Fresnel (France)


Published in SPIE Proceedings Vol. 9711:
Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues IX
Daniel L. Farkas; Dan V. Nicolau; Robert C. Leif, Editor(s)

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