gms | German Medical Science

Measurement of time-resolved auto-fluorescence according to the principle of time-correlated single photon counting is an extremely sensitive method in which application all disturbing influences should be estimated. In a first approximation, the eye can be assumed as a 2- layer model, in which the anterior parts, especially the crystalline lens, and the fundus exhibit fluorescent properties. It was to investigate, in which degree fundus lifetime measurements are influenced by auto-fluorescence of the lens.

Theoretically, the influence of light, diffusely reflected at anterior parts of the eye is reduced by factor 225, assuming 3 mm as diameter of the aperture diaphragm and 0.2 mm as diameter of the confocal field stop. This consideration does not account for increasing absorption and scattering of short wave excitation light in the ageing lens, resulting in a reduced excitation at the fundus. Furthermore considerable different concentration-thickness products have to be considered between lens and fundus in fluorescence measurements. Also in case of equal concentration of fluorophore, only a suppressing factor of 30 is calculated if the retinal thickness is assumed with 0.5 mm and the thickness of the lens is 4 mm and additionally an ocular transmission of 20% is assumed for a 70-year-old person. Measurements on healthy persons show a tendency to longer lifetimes with increasing age, but in case of artificial eye lenses, shorter lifetimes are measurable again. A method will be explained, permitting a correction of influence of auto-fluorescence of lens on fundus lifetime measurements, using multiple lifetime measurements under different experimental conditions.

Using the developed method, the dynamic auto-fluorescence of the lens can be separately be determined from fundus fluorescence. So, the fundus fluorescence lifetime can more exactly be calculated. The method is also convenient for determination of lifetimes of single fluorophores based on measurements of superposed sum lifetimes on fundus.

The dynamic auto-fluorescence measured both from the whole eye and on the fundus is an superposition of different lifetimes. Applying the explained method, an experimental and calculated separation of the contribution of several fluorophores is possible. By this means, the assessment of the metabolic state by lifetime mapping will considerably be improved.