Spatiotemporal focusing-based widefield multiphoton microscopy for fast optical sectioning of thick tissues

Li Chung Cheng*, Chia Yuan Chang, Wei Chung Yen, Shean-Jen Chen

*Corresponding author for this work

Research output: Chapter in Book/Report/Conference proceedingConference contributionpeer-review

2 Scopus citations

Abstract

Conventional multiphoton microscopy employs beam scanning; however, in this study a microscope based on spatiotemporal focusing offering widefield multiphoton excitation has been developed to provide fast optical sectioning images. The microscope integrates a 10 kHz repetition rate ultrafast amplifier featuring strong instantaneous peak power (maximum 400 μJ/pulse at 90 fs pulse width) with a TE-cooled, ultra-sensitive photon detecting, electron multiplying charge-coupled device camera. This configuration can produce multiphoton excited images with an excitation area larger than 200 × 100 μm2 at a frame rate greater than 100 Hz. Brownian motions of fluorescent microbeads as small as 0.5 μm have been instantaneously observed with a lateral spatial resolution of less than 0.5 μm and an axial resolution of approximately 3.5 μm. Moreover, we combine the widefield multiphoton microscopy with structure illuminated technique named HiLo to reject the background scattering noise to get better quality for bioimaging.

Original languageEnglish
Title of host publicationUnconventional Imaging and Wavefront Sensing 2012
DOIs
StatePublished - 1 Dec 2012
EventUnconventional Imaging and Wavefront Sensing 2012 - San Diego, CA, United States
Duration: 13 Aug 201214 Aug 2012

Publication series

NameProceedings of SPIE - The International Society for Optical Engineering
Volume8520
ISSN (Print)0277-786X

Conference

ConferenceUnconventional Imaging and Wavefront Sensing 2012
CountryUnited States
CitySan Diego, CA
Period13/08/1214/08/12

Keywords

  • Brownian motion
  • Second harmonic Generation
  • Spatiotemporal focusing
  • Widefield multiphoton microscopy

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