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Journal of Biomedical Optics

Loss of image quality in photobleaching during microscopic imaging of fluorescent probes bound to chromatin
Author(s): Tytus Bernas; Bartlomiej P. Rajwa; Elikplimki K. Asem; Joseph Paul Robinson
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Paper Abstract

Prolonged excitation of fluorescent probes leads eventually to loss of their capacity to emit light. A decrease in the number of detected photons reduces subsequently the resolving power of a fluorescence microscope. Adverse effects of fluorescence intensity loss on the quality of microscopic images of biological specimens have been recognized, but not determined quantitatively. We propose three human-independent methods of quality determination. These techniques require no reference images and are based on calculation of the actual resolution distance, information entropy, and signal-to-noise ratio (SNR). We apply the three measures to study the effect of photobleaching in cell nuclei stained with propidium iodide (PI) and chromomycin A3 (CA3) and imaged with fluorescence confocal microscopy. We conclude that the relative loss of image quality is smaller than the corresponding decrease in fluorescence intensity. Furthermore, the extent of quality loss is related to the optical properties of the imaging system and the noise characteristics of the detector. We discuss the importance of these findings for optimal registration and compression of biological images.

Paper Details

Date Published: 1 November 2005
PDF: 9 pages
J. Biomed. Opt. 10(6) 064015 doi: 10.1117/1.2136313
Published in: Journal of Biomedical Optics Volume 10, Issue 6
Show Author Affiliations
Tytus Bernas, Purdue Univ. (United States)
Bartlomiej P. Rajwa, Purdue Univ. (United States)
Elikplimki K. Asem, Purdue Univ. (United States)
Joseph Paul Robinson, Purdue Univ. (United States)


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