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  Optical sectioning fluorescence spectroscopy in a Programmable Array Microscope

Hanley, Q. S., Verveer, P. J., & Jovin, T. M. (1998). Optical sectioning fluorescence spectroscopy in a Programmable Array Microscope. Applied Spectroscopy, 52, 783-789.

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Item Permalink: http://hdl.handle.net/11858/00-001M-0000-0012-FCE5-2 Version Permalink: http://hdl.handle.net/11858/00-001M-0000-0027-E5E7-7
Genre: Journal Article

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 Creators:
Hanley, Q. S.1, Author              
Verveer, P. J.1, Author              
Jovin, T. M.1, Author              
Affiliations:
1Department of Molecular Biology, MPI for biophysical chemistry, Max Planck Society, ou_578628              

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Free keywords: Confocal microscopy; Confocal spectroscopy; Hadamard; Imaging spectroscopy; Fluorescence microscopy; Spatial light modulator
 Abstract: We report the use of a programmable array microscope (PAM) for the acquisition of spectrally resolved and high throughput optical sections. The microscope is based on the use of a spatial light modulator for defining patterns of illumination and/or detection. To obtain optically sectioned spectral images, the entrance slit of an imaging spectrograph and a line illumination pattern defined with a spatial light modulator are placed in conjugate optical positions. Compared to wide-field illumination, optical sectioning led to greater than 3x improvement in the rejection of out-of-focus fluorescence emission and nearly 6x greater peak-to-background ratios in biological specimens, yielding better contrast and spectral characterization. These effects resulted from a reduction in the artifacts arising from spectral contributions of structures outside the region of interest. We used the programmable illumination capability of the spectroscopic system to explore a variety of excitation/detection patterns for increasing the throughput of optical sectioning microscopes. A Sylvester type Hadamard construction was particularly efficient, performing optical sectioning while maintaining a 50% optical throughput. These results demonstrate the feasibility of full-field highly multiplexed confocal spectral imaging.

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Language(s): eng - English
 Dates: 2005-07-061998
 Publication Status: Published in print
 Pages: -
 Publishing info: -
 Table of Contents: -
 Rev. Method: -
 Identifiers: eDoc: 226078
Other: 629
 Degree: -

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Title: Applied Spectroscopy
Source Genre: Journal
 Creator(s):
Affiliations:
Publ. Info: -
Pages: - Volume / Issue: 52 Sequence Number: - Start / End Page: 783 - 789 Identifier: -