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R.N. Ebner, P. Rozenfeld; Enzymatic Diagnosis of Fabry Disease in Tears Sampled on Filter Paper. Observations in a Series of Non–Treated Patients, Treated Patients and Controls . Invest. Ophthalmol. Vis. Sci. 2005;46(13):1842.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: To analyze the AGA levels in tears, obtained from non–treated patients, treated patients and controls. Methods: Tears samples were taken from the conjunctival sac in: 21 Fabry patients, 3 patients undergoing replacement therapy with agalidase–A (ReplagalTM), and 28 normal controls. The filter paper used was Whatman® #41, standard for Shirmer test. A 5–mm diameter circle paper was placed into a well and 70 µl of the reaction mixture containing 3.57M 4–MU–α–galactoside and 0.07M N–acetilgalactosamine in acetate buffer pH=4,5 was added. After incubation at 37°C for 3 hrs, the stop solution was added, and the fluorescence was measured. Results: The kinetics of the reaction was linear. The mean values obtained from normal controls (n=28) was 40.61 µmoles/l.h.(± 24.99), for Fabry non–treated patients (n=21) the value obtained was 5.25 µmoles/l.h.(± 4.43) and for treated patients (n=3) was 14.61 µmoles/l.h .(± 4.99). The differences between non–treated patients and control groups were statistically significative ( p=0.0001). When compared the treated patients vs. control groups, the difference was not significative ( p = 0.130). Statistics were calculated using ANOVA test, when applying Bartlett's test the variants were homogenous (p<0.00001) showing that they differ significantly, this obliges to use the non–parametric Kruskal–Wallis test to compare the 3 groups. This test was also very significant (p<0.000001) demonstrating that groups strongly differ between them. The software used was: Primer for Biostatistics, v. 4.0 and EpiInfo, v. 5.1 (WHO). Conclusions: The determination of AGA activity in patients with Fabry disease with and without treatment, and controls were easy to obtain. The differences among groups were statistically significant when compared non–treated patients and controls, on the contrary there were non significant differences when the statistical analysis compared treated patients and controls. This method helped, first; with the diagnostic process among heterozygote–hemyzigote patients with Fabry's and second; to monitor the presence of enzyme higher levels in the tears obtained from treated patients. The higher AGA values observed in tears, among patients under replacement therapy, was an important finding in the process to monitor the patient’s response to the drug, this has been unreported at present. The data obtained are useful to consider this procedure as a diagnostic and follow–up tool.
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