Multiphoton excitation fluorescence imaging applied to the study of embryo development

David L. Wokosin; John G. White

doi:10.1117/12.312334

8 July 1998 Multiphoton excitation fluorescence imaging applied to the study of embryo development

David L. Wokosin, John G. White

Proceedings Volume 3269, Commercial Applications of Ultrafast Lasers; (1998) https://doi.org/10.1117/12.312334
Event: Optoelectronics and High-Power Lasers and Applications, 1998, San Jose, CA, United States

Abstract

The use of fluorescent probes is a powerful technique for the study of living specimens. Unfortunately, living tissues are vulnerable to photodamage from the excitation illumination and they make poor optical specimens due to their light-scattering nature. Multiphoton (two or more photon) excitation imaging offers significant advantages compared to laser-scanning confocal fluorescence microscopy for fluorescence microscopy of live specimens: considerable reduction in total sample fluorophore excitation and hence less photodamage, increased depth penetration due to increased tolerance for scattering, and increased detection sensitivity as more signal photons can be used for imaging. These advantages become more significant if 3D or 4D (multifocal plane, time-lapse) imaging is undertaken. In addition, multiphoton excitation imaging allows UV excited probes such as DAPI or INDO I or endogenous fluorophores such as NAD(P)H and serotonin to be imaged without UV excitation. We, and others, have been evaluating the potential of multi-photon excitation imaging for biological microscopy and have found all of the aforementioned advantages particularly significant for laser-scanning fluorescence imaging of developing embryos; a summary of currently pursued developmental biology applications will be presented. The current status of all-solid-state ultrafast lasers as excitation sources will also be reviewed since these lasers offer tremendous potential for affordable, reliable, 'turnkey' multiphoton imaging systems. The combination of demonstrated applications, simple ultrafast laser sources, and affordable commercial systems may promote a revolution in the study of embryogenesis with the light microscope.

Citation Download Citation

David L. Wokosin and John G. White "Multiphoton excitation fluorescence imaging applied to the study of embryo development", Proc. SPIE 3269, Commercial Applications of Ultrafast Lasers, (8 July 1998); https://doi.org/10.1117/12.312334

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