Ere consecutively recruited from the outpatient clinic of the department of

Ere consecutively recruited from the outpatient clinic of the department of neurology at the Heinrich-Heine University Dusseldorf, Germany. ?Optical Coherence TomographyThe details of the principles of spectral-domain OCT have been described elsewhere [28]. Using a Spectralis OCT device (Heidelberg Engineering, Heidelberg, Germany) with image alignment eye tracking-software (TruTrack, Heidelberg Engineering, Heidelberg, Germany), we obtained perifoveal volumetric retinal scans consisting of 25 single vertical axial scans (scanning area: 6 66 mm, centered at the fovea). To assess the peripapillary RNFL, a circular scan with a diameter of approximately 3.4 mm was performed after manually positioning the center on the middle of the optic disc (figure 1 A). Furthermore, we performed highresolution horizontal scans through the middle of the fovea. All of the scans were performed using the eye-tracking system. The RNFL measurements and high-resolution single horizontal scans were 298690-60-5 site averaged from 100 images and measurements for volumetric calculations were averaged from 10 scans (Automatic Real Time, ART). All of the scans were of sufficient or good quality (.20 DB) and fulfilled OSCAR-IB quality control criteria for OCT scans [29]. While the results of the RNFL- and paramacular volumetric measurements were automatically analyzed by the Heidelberg Eye explorer software, the paramacular retinal layers were manually segmented as previously described [18,30]. In brief, the segmentation of the different retinal layers in the single horizontal foveal scans was performed manually by repositioning the measurement lines (the white dotted lines in figure 2 A) onFigure 1. RNFL and mean macular thickness are reduced in Wilson’s disease. A The areas of measurement are marked in an image of the fundus. The RNFL was measured in a circular scan centered on the optic disc and the macular thickness in scans around the fovea. The manual segmentation of the retinal layers was performed in a horizontal scan through the center of the fovea. The scatter plots display the thickness of the RNFL (B) and the total macular thickness (C). Each point represents the mean of the two eyes of one patient. The mean of all DprE1-IN-2 biological activity patients is indicated by a horizontal bar. Significant differences are indicated by asterisks (p,0.05, two-tailed t-test). doi:10.1371/journal.pone.0049825.gthe borders between the different layers and measuring the layers at their thickest points nasally and temporally of the macula. In most subjects, the outer nuclear layer (ONL) presented only one central thickest point rather than nasal and temporal peaks as in the other layers. We therefore used the thickness at this point for analysis (the central vertical line in figure 2 A). In the few subjects with a nasal and temporal ONL thickness peaks, we used the higher value.Visual Evoked Potentials (VEPs)VEPs were recorded 1326631 in 27 Wilson’s disease patients and 26 control probands using monocular stimulation with a fullfield black and white checkerboard generated on a TV monitor and reversed in contrast at a rate of 3.1/s. The square size was 12 mm, which subtended a visual angle of 41 min at the subject’s eyes. VEPs were recorded from the scalp using pin electrodes positioned at Oz (active) and Fz (reference) sites. A total of 150 responses were averaged in each recording and two trials were performed for each eye. The peak latencies of N75, P100 and N140 and peak-to-peakamplitudes from P100 to N140 were measured.Ere consecutively recruited from the outpatient clinic of the department of neurology at the Heinrich-Heine University Dusseldorf, Germany. ?Optical Coherence TomographyThe details of the principles of spectral-domain OCT have been described elsewhere [28]. Using a Spectralis OCT device (Heidelberg Engineering, Heidelberg, Germany) with image alignment eye tracking-software (TruTrack, Heidelberg Engineering, Heidelberg, Germany), we obtained perifoveal volumetric retinal scans consisting of 25 single vertical axial scans (scanning area: 6 66 mm, centered at the fovea). To assess the peripapillary RNFL, a circular scan with a diameter of approximately 3.4 mm was performed after manually positioning the center on the middle of the optic disc (figure 1 A). Furthermore, we performed highresolution horizontal scans through the middle of the fovea. All of the scans were performed using the eye-tracking system. The RNFL measurements and high-resolution single horizontal scans were averaged from 100 images and measurements for volumetric calculations were averaged from 10 scans (Automatic Real Time, ART). All of the scans were of sufficient or good quality (.20 DB) and fulfilled OSCAR-IB quality control criteria for OCT scans [29]. While the results of the RNFL- and paramacular volumetric measurements were automatically analyzed by the Heidelberg Eye explorer software, the paramacular retinal layers were manually segmented as previously described [18,30]. In brief, the segmentation of the different retinal layers in the single horizontal foveal scans was performed manually by repositioning the measurement lines (the white dotted lines in figure 2 A) onFigure 1. RNFL and mean macular thickness are reduced in Wilson’s disease. A The areas of measurement are marked in an image of the fundus. The RNFL was measured in a circular scan centered on the optic disc and the macular thickness in scans around the fovea. The manual segmentation of the retinal layers was performed in a horizontal scan through the center of the fovea. The scatter plots display the thickness of the RNFL (B) and the total macular thickness (C). Each point represents the mean of the two eyes of one patient. The mean of all patients is indicated by a horizontal bar. Significant differences are indicated by asterisks (p,0.05, two-tailed t-test). doi:10.1371/journal.pone.0049825.gthe borders between the different layers and measuring the layers at their thickest points nasally and temporally of the macula. In most subjects, the outer nuclear layer (ONL) presented only one central thickest point rather than nasal and temporal peaks as in the other layers. We therefore used the thickness at this point for analysis (the central vertical line in figure 2 A). In the few subjects with a nasal and temporal ONL thickness peaks, we used the higher value.Visual Evoked Potentials (VEPs)VEPs were recorded 1326631 in 27 Wilson’s disease patients and 26 control probands using monocular stimulation with a fullfield black and white checkerboard generated on a TV monitor and reversed in contrast at a rate of 3.1/s. The square size was 12 mm, which subtended a visual angle of 41 min at the subject’s eyes. VEPs were recorded from the scalp using pin electrodes positioned at Oz (active) and Fz (reference) sites. A total of 150 responses were averaged in each recording and two trials were performed for each eye. The peak latencies of N75, P100 and N140 and peak-to-peakamplitudes from P100 to N140 were measured.

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