Wide-field three-photon excitation in biological samples
Author(s)
Fukumura, Dai; Jain, Rakesh K; Rowlands, Christopher; Park, Demian; Bruns, Oliver Thomas; Piatkevich, Kiryl; Bawendi, Moungi G; Boyden, Edward; So, Peter T. C.; ... Show more Show less
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Three-photon wide-field depth-resolved excitation is used to overcome some of the limitations in conventional point-scanning two- and three-photon microscopy. Excitation of chromophores as diverse as channelrhodopsins and quantum dots is shown, and a penetration depth of more than 700 μm into fixed scattering brain tissue is achieved, approximately twice as deep as that achieved using two-photon wide-field excitation. Compatibility with live animal experiments is confirmed by imaging the cerebral vasculature of an anesthetized mouse; a complete focal stack was obtained without any evidence of photodamage. As an additional validation of the utility of wide-field three-photon excitation, functional excitation is demonstrated by performing three-photon optogenetic stimulation of cultured mouse hippocampal neurons expressing a channelrhodopsin; action potentials could reliably be excited without causing photodamage.
Date issued
2017-05Department
Massachusetts Institute of Technology. Department of Biological Engineering; Massachusetts Institute of Technology. Department of Brain and Cognitive Sciences; Massachusetts Institute of Technology. Department of Chemistry; Massachusetts Institute of Technology. Department of Mechanical Engineering; Massachusetts Institute of Technology. Media Laboratory; McGovern Institute for Brain Research at MIT; Massachusetts Institute of Technology. Center for Neurobiological EngineeringJournal
Light: Science & Applications
Publisher
Springer Nature
Citation
Rowlands, Christopher J et al. “Wide-Field Three-Photon Excitation in Biological Samples.” Light: Science & Applications 6, 5 (November 2016): e16255 © 2017 The Author(s)
Version: Author's final manuscript
ISSN
2047-7538