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Proceedings Paper

Development and application of 2-color live-cell STED nanoscopy (Conference Presentation)
Author(s): Edward S. Allgeyer; Francesca Bottanelli; Emil B. Kromann; Xiang Hao; Joerg Bewersdorf

Paper Abstract

Stimulated emission depletion (STED) microscopy has been established as an important technique for imaging below the diffraction limit facilitating new discoveries in an array of biological systems. In STED microscopy a “donut-shaped” laser focus is super-imposed onto the diffraction-limited focus of an excitation laser. The dounut-shaped beam suppresses fluorescence in the periphery of the excitation spot, reducing the effective point spread function to a sub-diffraction size. However, the application of multicolor STED microscopy in living cells poses a number of challenges. Here we detail a novel STED system specifically designed for two-color STED applications. Our system employs FPGA-based gated detection and fast beam scanning to reduce pixel dwell time and photobleaching. We demonstrate the instrument’s capability with two-color continuous imaging of intracellular targets below the diffraction limit allowing observation of rare events within live-cells.

Paper Details

Date Published: 27 April 2016
PDF: 1 pages
Proc. SPIE 9714, Single Molecule Spectroscopy and Superresolution Imaging IX, 97140D (27 April 2016); doi: 10.1117/12.2213728
Show Author Affiliations
Edward S. Allgeyer, Yale School of Medicine (United States)
Francesca Bottanelli, Yale School of Medicine (United States)
Emil B. Kromann, Yale School of Medicine (United States)
Xiang Hao, Yale School of Medicine (United States)
Joerg Bewersdorf, Yale School of Medicine (United States)

Published in SPIE Proceedings Vol. 9714:
Single Molecule Spectroscopy and Superresolution Imaging IX
Jörg Enderlein; Ingo Gregor; Zygmunt Karol Gryczynski; Rainer Erdmann; Felix Koberling, Editor(s)

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