Purpose
FAF images are currently analyzed descriptively and qualitatively. The purpose is to assess the reliability of a new standardized measurement technique designed to quantitatively analyze fundus autofluorescence (FAF) images.
Methods
FAF photos were captured in eyes of healthy subjects using a Canon CX-1 Hybrid Mydriatic/Non-mydriatic retinal camera. Single static images were captured using a flash power of 300 w/sec with a wide band excitation and barrier filter (535-585 nm and 605-715 nm, respectively) and a 45 degree field of view (FOV). The zone between the temporal edge of the optic nerve (ON) and macula (M), designated as the papillo-macular zone (P), of each image was chosen and analyzed by assigning a grayscale pixel intensity, ranging from 0 (black) to 256 (white). A grayscale pixel score was determined for each image (Figure 1). To reduce background noise, the same images were then “standardized” using a technique, which adjusted a square area (1.80 x 1.80mm) overlying the ON to an intensity of 0 (Figure 2). In order to determine the consistency of the scoring, the standardized images were analyzed twice, blindly on two separate occasions by one observer. Means, standard deviations (SD), t-tests, and an intra-class correlation (ICC) were performed.
Results
In total, 96 images were analyzed (from the left eye of 32 subjects, mean age=23 yrs). Two eyes had poor quality FAF images and were excluded from the analysis. In the non-standardized image readings, the mean pixel intensity for zone P was 84.64±29.27. After standardization to reduce background noise, the mean pixel intensity for zone P was highly comparable between the two readings: 38.80±12.81 on the first read (R1) and 38.78 ± 12.84 on the second read (R2) (p=0.99). The intra-class correlation between the two standardized readings was 0.99.
Conclusions
This novel standardized technique applied to FAF images was strongly reliable and reproducible and can yield scores that quantify autofluorescence in a preselected zone in the fundus. Clinical applications will allow quantitative scoring of FAF and follow-up in clinical therapeutic and research settings. A larger study testing this quantitative scoring system in different demographic populations are needed to further characterize the full utility of this modality.