Mariano Cozzi

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	By Mariano Cozzi and Dr Giovanni Staurenghi

	Most ocular structures become luminescent for a short time when exposed to a light source of appropriate wavelength; this phenomenon is known as autofluorescence (AF). AF is due to the presence of fluorescent substances called fluorophores. Of special importance for diagnostic purposes is the autofluorescence of the ocular fundus (fundus autofluorescence; FAF) caused by the presence of lipofuscin, the most important fluorophore of the retinal pigment epithelium (RPE).

	Lipofuscin is the product of degradation of the outer segment disks of photoreceptors, which in normal conditions are phagocytised by RPE cells. Many retinal pathologies cause accumulation or depletion of lipofuscin, and consequently, an increase or a reduction of autofluorescence. FAF is increased in the following diseases: age-related macular degeneration; Stargardt disease; Best disease; and adult-onset foveomacular vitelliform macular dystrophy. Levels are also raised in the drusen of the sub-pigment epithelial space and in choroidal tumours, such as nevi and melanomas.

	Reduced or absent FAF occurs in cases of damage of the RPE, as in geographic atrophy, cystoid macular oedema, and in recent retinal haemorrhages. Sometimes, the alterations of FAF precede the anatomic alterations, hence FAF imaging is being considered as increasingly important for the evaluation of patients with fundus alterations.

The confocal retinal imager (EIDON AF, CenterVue Spa)

 is a true-colour, last-generation confocal scanner with unique characteristics. Thanks to its confocal optics, 

 collects only the light reflected by the focal plan, excluding the components of the light originated by other eye structures. 

 is able to capture sharp and well-defined images, with a higher contrast compared with non-confocal imagers. This is achieved without averaging multiple images to increase the signal-to-noise ratio, as other confocal scanners do.

 can capture high-quality images even in conditions of poor mydriasis (pupil diameter of 2.5 mm) and can explore an area of the fundus of 60° (h) by 55° (w). It is noteworthy that, thanks to its 15 MP sensor, 

 obtains the images with the highest pixel resolution among scanning laser ophthalmoscopes.

captures three different types of images:

	·       those in true colours, obtained through a white light source (LED, 440-650 nm);

	·       images in black and white, obtained using an infra-red source (LED, 825-870 nm) to examine choroid and the deepest layers of the retina;

	·       and red-free images to enhance the visibility of the vessels and nerve fiber layer.

	In addition to the above-mentioned modes, the imager uses a blue light source (LED 450 nm) together with a barrier filter (500 nm) to capture FAF.

 can operate in full-automatic mode for the capture of true colour, infra-red and autofluorescent images. In automatic mode, 

 controls the alignment and focus, and obtains a snapshot of both eyes in less than a minute. The system can also operate in manual and semi-automatic modes.

	We had the opportunity to test the new autofluorescence function of 

 in our Center of Medical Retina at the Ophthalmologic Clinic of Hospital Sacco (Milan University).

	Autofluorescence is a diagnostic exam that, today, should be used routinely for more accurate diagnosis and easier differential diagnosis. Blue-light autofluorescence is of special importance because it highlights the macular pigment, hence playing a crucial role in diagnosis of macular alterations. Blue-light FAF, however, requires confocal scanners. 

 is the possibility to obtain, in automatic mode, FAF images of a field of 60° even in case of poor dilatation of the pupil. We report below the case of an eight-year-old patient, where the capability to capture a field of 60° showed its importance in the diagnosis.

	The images (Figure 1) show bilateral alterations of the retina, consisting in hyperfluorescent areas known as ‘flecks’, a typical lesion of Stargardt disease.

	The above-mentioned lesions can extend beyond the retinal arcades; it is therefore very important to capture an image that includes the mid periphery of the retina; this to observe all lesions of the fundus in the same frame. Despite the non-collaborative behaviour of the patient (due to the young age) and a severe photosensitivity, it was possible to capture high-quality images of the retina.

	The FAF examination is also of importance in patients affected by lamellar hole, as shown in 

	The lesion, clearly visible in autofluorescence, involves mainly the foveal zone, with a retinal delamination, generally between the outer plexiform layer and the outer nuclear layer. The autofluorescence is also of importance to define the differential diagnosis and the adequate treatment in case of pseudo-hole, as such patients do not have loss of retinal tissue. 

	Finally, one of the most frequent cases where the autofluorescence exam demonstrates its importance is probably in geographic atrophy 

	To evaluate the retinal damage in this disease, it is better to rely on autofluorescence imaging rather than on visual acuity. As a matter of fact, autofluorescence proved to be more accurate than true colour photos in the evaluation of the lesions’ borders. Geographic atrophy can be the outcome of different pathologies, evolving into similar retinal lesions.

	Today it is paramount to identify and differentiate the patterns of posterior pole atrophy. There are many clinical trials aimed at the evaluation of efficacy and safety of new pharmacologic treatments; the availability of high-quality documentation of the lesions has direct consequences on the therapy of the patients. A continued follow up of patients by means of autofluorescence images is of great help in following their evolution and the progression of the pathologies.

 make it a valuable tool for ophthalmologists, both in the hospital and in private practice. 

 can capture autofluorescence images of the ocular fundus on a 60° field, also in myosis.

	In addition, the capacity to obtain images with only one snapshot, without averaging multiple images, allows one to examine photophobic patients. The differential diagnosis in retinal pathologies is, without any doubt, made easier by a multimodal approach.

	Autofluorescence plays a fundamental role by enabling the identification of lipofuscin deposits in hereditary degenerative diseases or in melanomas to monitor the evolution of geographic atrophies; supporting the differential diagnosis between lamellar hole and pseudo-hole; and allowing the presence of macular pigment for the diagnosis of juxtafoveolar retinal telangiectasia to be assessed. 

Articles

Most ocular structures become luminescent for a short time when exposed to a light source of appropriate wavelength; this phenomenon is known as autofluorescence (AF). AF is due to the presence of fluorescent substances called fluorophores.

Author | Mariano Cozzi

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