Journal Article

Real-time high dynamic range laser scanning microscopy

David Rousso, Shawn Stapleton, Ralph Weissleder, Claudio Vinegoni, Randy J. Giedt, C. Leon Swisher, P. Fumene Feruglio

2016

Overview

In conventional confocal/multiphoton fluorescence microscopy, images are typically acquired under ideal settings and after extensive optimization of parameters for a given structure or feature, often resulting in information loss from other image attributes. To overcome the problem of selective data display, we developed a new method that extends the imaging dynamic range in optical microscopy and improves the signal-to-noise ratio. Here we demonstrate how real-time and sequential high dynamic range microscopy facilitates automated three-dimensional neural segmentation. We address reconstruction and segmentation performance on samples with different size, anatomy and complexity. Finally, in vivo real-time high dynamic range imaging is also demonstrated, making the technique particularly relevant for longitudinal imaging in the presence of physiological motion and/or for quantification of in vivo fast tracer kinetics during functional imaging. Confocal and multiphoton fluorescence microscopy often suffers from low dynamic range. Here the authors develop a high dynamic range, laser scanning fluorescence technique by simultaneously recording different light intensity ranges. The method can be adapted to commercial systems.

• Publisher: Springer Science and Business Media LLC,Nature Publishing Group UK,Nature Publishing Group,Nature Portfolio
• Subject: 14/63 / 14/69 / 639/624/1107/328 / Algorithms / Animals / Brain / Brain - pathology / Confocal microscopy / Dynamic range / Fluorescence / Fluorescence microscopy / Humanities and Social Sciences / Image acquisition / Image processing / Image reconstruction / Image segmentation / imaging dynamic range; optical microscopy; signal-to-noise ratio; real-time; sequential high dynamic range microscopy; in vivo; functional imaging / In vivo methods and tests / Light microscopy / Mice / Microscopes / Microscopy / Microscopy, Confocal / Microscopy, Confocal - methods / Microscopy, Fluorescence, Multiphoton / multidisciplinary / Optical microscopy / Optimization / Q / Real time / Scanning microscopy / Science / Science (multidisciplinary) / Signal-To-Noise Ratio