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D. Schweitzer, F. Schweitzer, M. Hammer, S. Schenke, S. Richter; Time–Resolved Autofluorescence Yields More Information Than Intensity Measurements of Fundus Autofluorescence . Invest. Ophthalmol. Vis. Sci. 2005;46(13):227.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: To evaluate first in vivo measurements of time–resolved fundus autofluorescence in healthy subjects and AMD patients. Methods: Time–resolved autofluorescence was measured using a modified laser scanning ophthalmoscope. Autofluorescence was excited by a laser diode at 446 nm, 100 ps FWHM (LDH 440, Picoquant, Berlin, Germany) and detected in single photon counting technique (SPC730, B&H, Berlin, Germany) for wavelength > 475 nm. For image–registration, reflectance images were detected simultaneously with fluorescence images. Time–resolved autofluorescence was measured in 34 healthy and in 16 AMD eyes. Intensity images were determined by adding photons in all time–channels of each pixel. Applying SPCImage 2.6 software (B&H), dynamic fluorescence was approximated bi– and tri–exponentially, resulting in lifetime images of tau 1, 2, 3, of amplitudes a 1, 2, 3 and in images of relative contribution of components q 1, 2, 3. Results: The fluorescence of the crystalline lens contributes long decay components in fundus autofluorescence. For that reason, relative changes in images of dynamic autofluorescence of phacic and pseudophacic eyes are considered. In bi–exponential approximation, in dry AMD macular lifetime tau 2 is shorter and a2 is lower than in surrounding tissue. In contrast, in healthy subjects, macular tau 2 is longer and a2 is greater than in surroundings. In drusen, the contribution of the short component q1 is low, comparable with q1 in the optic disc. Selecting specific intervals, small areas of changed autofluorescence can be found in the macula in AMD. Comparing the local distribution of relative contributions q1, q2, and q3 with retinal anatomy, q1 might be determined by RPE, q2 by neural retina, and q3 by connective tissue. Lifetime measurements on porcine fundi (retina removed) (tau 1 = 150 ps, a1= 94%, tau 2 = 1500 ps, a2 = 6%) and isolated retina (tau = 380 ps) confirm this assumption. Conclusions: Whereas images of fluorescence intensity show only local differences in grey values, dynamic fluorescence permits determination of contribution of single components in fluorescence images.
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