title: Mapping retinal fluorescein leakage with confocal scanning laser fluorometry of the human vitreous.
abstract: OBJECTIVE
  To demonstrate an objective, quantitative, and sensitive method of mapping retinal fluorescein leakage into the vitreous while simultaneously imaging the retina.
  
  
  METHODS
  A prototype Zeiss confocal scanning laser ophthalmoscope was modified to obtain fluorometric measurements from 18 optical planes across the retina and cortical vitreous, separated from each other by 150 microm, and parallel to the retinal surface. After intravenous administration of fluorescein, an axial graphic of equivalent fluorescein concentration in the vitreous may be obtained from any region of interest. After correcting for fluorescence levels in the retina and choroid and plasma levels of free fluorescein, permeability values of the blood-retinal barrier to fluorescein were obtained from 1512 regions measuring 75 x 75 microm, from a total 3150 x 2700-microm area of the fundus, generating a detailed map of retinal fluorescein leakage. The method was assessed in vitro and in 7 healthy subjects who underwent scans during separate visits. Depth resolution and influence of chorioretinal fluorescence were further tested in 2 patients with multiple drusen and in 2 eyes after vitrectomy. Fourteen eyes from 7 patients with diabetes and nonproliferative retinopathy were also examined. Lateral resolution was tested in 3 diabetic eyes that underwent focal photocoagulation. Four eyes from 2 patients with diabetes and minimal retinopathy were examined at 3-month intervals. All eyes examined had less than 2 diopters of astigmatism.
  
  
  RESULTS
  Characteristics of the modified confocal scanning laser fluorometer included a lower limit of detection equal to 0.40 Eq ng/mL and depth precision of +/-15 microm. Values for the blood-retinal barrier permeability index in healthy subjects, measured 30 minutes after a single intravenous pulse of fluorescein (14 mg/kg), ranged from 1.3 +/- 0.4 x 10(-6) cm/s over the foveal avascular zone to 2.2 +/- 0.6 x 10(-6) cm/s over vessels in the retina. Diabetic eyes with retinopathy showed higher values, ranging from 1.4 to 15.0 x 10(-6) cm/s. Vitrectomized eyes and eyes with multiple drusen showed the validity of the correction algorithm demonstrating that measurements of fluorescence in the vitreous are not influenced by the chorioretinal fluorescence level. Argon laser photocoagulation burns placed in the diabetic retina demonstrated a lateral resolution on the order of 75 to 100 microm. Intravisit and intervisit reproducibility was +/-10.2% and +/-13%, respectively.
  
  
  CONCLUSIONS
  This new method measures localized alterations of the blood-retinal barrier and allows for direct correlation with retinal anatomy. Its most interesting feature is the ability to map retinal fluorescein leakage while simultaneously imaging the retina. This capability is expected to improve our understanding and management of retinal disease.
mesh_terms: D000328:Adult; D001813:Blood-Retinal Barrier; D002199:Capillary Permeability; D003930:Diabetic Retinopathy / Q000378:metabolism / Q000473:pathology / Q000601:surgery; D005260:Female; D019793:Fluorescein / Q000378:metabolism*; D005451:Fluorescein Angiography; D015590:Fluorophotometry / Q000379:methods*; D006801:Humans; D017075:Laser Coagulation; D008297:Male; D019721:Ophthalmoscopes; D015203:Reproducibility of Results; D015593:Retinal Drusen / Q000378:metabolism / Q000473:pathology / Q000601:surgery; D012171:Retinal Vessels / Q000378:metabolism* / Q000473:pathology; D014821:Vitrectomy; D014822:Vitreous Body / Q000378:metabolism*