Simulation design of wide-field temporal-focusing multiphoton excitation with a tunable excitation wavelength

Chi Hsiang Lien, Chun Yu Lin, Chia-Yuan Chang, Fan Ching Chien

Research output: Contribution to journalArticle

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 good 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 μ 3% was obtained at this wavelength range. By using the 830 lines/mm grating, a collimating lens with a 500mm 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 μm2, 2.6 ± 0.3 μm, and 71 μ 29%, respectively, were obtained in two-photon excitation fluorescence imaging at 700-1000 nm excitation wavelengths.

Original languageEnglish
Pages (from-to)1174-1187
Number of pages14
JournalOSA Continuum
Volume2
Issue number4
DOIs
Publication statusPublished - 2019 Apr 15

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Wavelength
wavelengths
excitation
simulation
gratings
Lenses
Microscopic examination
Lighting
Ultrafast lasers
grooves
Diffraction efficiency
Fluorophores
radiant flux density
Diffraction gratings
illumination
lenses
water immersion
microscopy
Photons
Fluorescence

All Science Journal Classification (ASJC) codes

  • Electrical and Electronic Engineering
  • Electronic, Optical and Magnetic Materials
  • Atomic and Molecular Physics, and Optics

Cite this

Lien, Chi Hsiang ; Lin, Chun Yu ; Chang, Chia-Yuan ; Chien, Fan Ching. / Simulation design of wide-field temporal-focusing multiphoton excitation with a tunable excitation wavelength. In: OSA Continuum. 2019 ; Vol. 2, No. 4. pp. 1174-1187.
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Simulation design of wide-field temporal-focusing multiphoton excitation with a tunable excitation wavelength. / Lien, Chi Hsiang; Lin, Chun Yu; Chang, Chia-Yuan; Chien, Fan Ching.

In: OSA Continuum, Vol. 2, No. 4, 15.04.2019, p. 1174-1187.

Research output: Contribution to journalArticle

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