Simulation approach to optimize fluorescence imaging performance of wide-field temporal-focusing microscopy with tunable wavelength excitation

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

Abstract

The optical parameters of temporal-focusing multiphoton excitation microscopy (TFMPEM), which is capable of achieving varying wavelength excitation for multiple fluorophore measurement, was systematically examined to have better excitation performance. For this purpose, the approaches were adopted to quantitatively evaluate the grating groove density, focal length of the collimating lens and objective, and different excitation wavelengths. A grating with a groove density of 830 lines/mm enables the TFMPEM system to achieve a wavelength range of 700-1000 nm by adjusting the incident angle of the ultrafast laser on the grating; a diffraction efficiency of 81 ± 3 % was obtained at this wavelength range. By using the 830 lines/mm grating, a collimating lens with a 500 mm focal length, and a 60× water immersion objective, we achieved a large excitation area and a better filling effect of the spectrum band of the pulse on the back focal plane of the objective; these parameters ensure high optical sectioning and small variation in the illumination power density within this wavelength range. The variation ranges of the excitation area, optical sectioning, and illumination power density of 4272 ± 798 m2, 2.6 ± 0.3 m, and 71 ± 29 %, respectively, were obtained in two-photon excitation fluorescence imaging at 700-1000 nm excitation wavelengths.

Original languageEnglish
Title of host publicationEuropean Conference on Biomedical Optics, ECBO_2019
PublisherOSA - The Optical Society
ISBN (Electronic)9781557528209
DOIs
StatePublished - 2019
EventEuropean Conference on Biomedical Optics, ECBO_2019 - Munich, Netherlands
Duration: 23 Jun 201925 Jun 2019

Publication series

NameOptics InfoBase Conference Papers
VolumePart F142-ECBO 2019

Conference

ConferenceEuropean Conference on Biomedical Optics, ECBO_2019
Country/TerritoryNetherlands
CityMunich
Period23/06/1925/06/19

Keywords

  • Optical sectioning
  • Temporal-focusing multiphoton excitation microscopy
  • Tunable wavelength excitation
  • Two-photon excitation fluorescence

Fingerprint

Dive into the research topics of 'Simulation approach to optimize fluorescence imaging performance of wide-field temporal-focusing microscopy with tunable wavelength excitation'. Together they form a unique fingerprint.

Cite this