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PubMed · 8599631

Fluorescence lifetime imaging by asynchronous pump-probe microscopy.

C Y Dong·P T So·T French·E Gratton
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Abstract

We report the development of a scanning lifetime fluorescence microscope using the asynchronous, pump-probe (stimulated emission) approach. There are two significant advantages of this technique. First, the cross-correlation signal produced by overlapping the pump and probe lasers results in i) an axial sectioning effect similar to that in confocal and two-photon excitation microscopy, and ii) improved spatial resolution compared to conventional one-photon fluorescence microscopy. Second, the low-frequency, cross-correlation signal generated allows lifetime-resolved imaging without using fast photodetectors. The data presented here include 1) determination of laser sources' threshold powers for linearity in the pump-probe signal; 2) characterization of the pump-probe intensity profile using 0.28 microns fluorescent latex spheres; 3) high frequency (up to 6.7 GHz) lifetime measurement of rhodamine B in water; and 4) lifetime-resolved images of fluorescent latex spheres, human erythrocytes and a mouse fibroblast cell stained by rhodamine DHPE, and a mouse fibroblast labeled with ethidium bromide and rhodamine DHPE.

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AnimalsErythrocytesFibroblastsFluorescent DyesHumansLatexMiceMicroscopy, FluorescenceMicrospheresPhotonsTime FactorsJournal ArticleResearch Support, U.S. Gov't, P.H.S.

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C Y Dong, P T So, T French, E Gratton. 1995. Fluorescence lifetime imaging by asynchronous pump-probe microscopy.. https://doi.org/10.1016/s0006-3495(95)80148-7

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