Combined spectrally-resolved multiphoton microscopy and transmission microscopy employing a high-sensitivity electron-multiplying CCD camera

Valeric? Raicu; Russell Fung; Mike Melnichuk; Anurag Chaturvedi; Devin Gillman

doi:10.1117/12.705920

10 February 2007 Combined spectrally-resolved multiphoton microscopy and transmission microscopy employing a high-sensitivity electron-multiplying CCD camera

Valeric? Raicu, Russell Fung, Mike Melnichuk, Anurag Chaturvedi, Devin Gillman

Author Affiliations +

Proceedings Volume 6442, Multiphoton Microscopy in the Biomedical Sciences VII; 64420M (2007) https://doi.org/10.1117/12.705920
Event: SPIE BiOS, 2007, San Jose, California, United States

Abstract

We have developed a fast, sensitive multiphoton microscope employing a tunable femtosecond laser and an electron-multiplying CCD camera (EMCCD) in a non-descanned detection scheme. In one configuration, the microscope provides "standard" multiphoton fluorescence images with a relatively high acquisition rate (limited only by the speed of the scanners) over a broad band of emission wavelengths. In a second configuration, the use of a diffractive optic element allows acquisition of one full set of spectrally-resolved images after only one complete scan. Increased speed is achieved in this configuration by setting the integration time of the camera such that an entire line is scanned at once, with the spectrum at each point being perpendicular to the scanned line. In both configurations, a suitable optical arrangement permits acquisition of high resolution images for both multiphoton excitation of fluorescence and widefield transmission microscopy. To illustrate the performances of our microscope we present spectrally-resolved images of individual fluorescent molecules spread over a surface as well as fluorescence images of yeast cells expressing a membrane receptor tagged with a variant of the green fluorescent protein.

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Valeric? Raicu, Russell Fung, Mike Melnichuk, Anurag Chaturvedi, and Devin Gillman "Combined spectrally-resolved multiphoton microscopy and transmission microscopy employing a high-sensitivity electron-multiplying CCD camera", Proc. SPIE 6442, Multiphoton Microscopy in the Biomedical Sciences VII, 64420M (10 February 2007); https://doi.org/10.1117/12.705920

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