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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 anddoi:10.1117/1.2136313 pmid:16409080 fatcat:sd6ctlxrxbgtrf5hmcgymupyde