Advances in frequency-domain flourometry; gigahertz instrumentation; time-dependent photo migration and flourescence lifetime imaging

Joseph R. Lakowicz

doi:10.1117/12.2322288

1 February 1992 Advances in frequency-domain flourometry; gigahertz instrumentation; time-dependent photo migration and flourescence lifetime imaging

Joseph R. Lakowicz

Author Affiliations +

Proceedings Volume 1599, Recent Advances in the Uses of Light in Physics, Chemistry, Engineering, and Medicine; (1992) https://doi.org/10.1117/12.2322288
Event: Recent Advances in the Uses of Light in Physics, Chemistry, Engineering, and Medicine, 1991, New York, NY, United States

Abstract

During the past seven years, there have been remarkable advances in the frequency—domain method for measurement of time—resolved emission or light scattering. In this presentation we describe the recent extension of the frequency range to 10 GHz using a specially designed microchannel plate PMT. Experimental data will be shown for measurement of picosecond rotational diffusion and for sub—picosecond resolution of time delays. The resolution of PS to ns timescale processes is not obtained at the expense of sensitivity, or is shown by measurements on the intrinsic tryptophan emission from hemoglobin. We also describe a time—resolved reflectance imaging experiment on a scattering medium containing an absorbing object. Time—resolved imaging of the back—scattered light is realized by means of a RF—phase—sensitive camera, synchronized to the laser pulses. By processing the stored images, a final image can be created, the contrast of which is based only on time differences of the back—scattered photons. This image reveals the presence and position of the absorber within the scattering medium. And finally, we describe a new methodology, fluorescence lifetime imaging (FLIM) , in which the contrast depends on the fluorescence lifetime at each point in a two—dimensional image, and not the local concentration and/or intensity of the fluorophore. We used FLIM to create lifetime images of NADH when free in solution and when bound to malate dehydrogenase. FLIM has numerous potential applications in cell biology and imaging.

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Joseph R. Lakowicz "Advances in frequency-domain flourometry; gigahertz instrumentation; time-dependent photo migration and flourescence lifetime imaging", Proc. SPIE 1599, Recent Advances in the Uses of Light in Physics, Chemistry, Engineering, and Medicine, (1 February 1992); https://doi.org/10.1117/12.2322288

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KEYWORDS

Modulation

Phase shift keying

Fluorescence lifetime imaging

Picosecond phenomena

Chemistry

Medicine

Photons

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