Novel In-Office Technique for Visual Confirmation of Demodex Infestation in Blepharitic Patients.

PURPOSE: To determine whether Demodex infestation in blepharitic patients can be confirmed by slit-lamp examination without the need for light microscopy. METHODS: Demodex infestation was evaluated in 16 patients presenting with blepharitis and cylindrical dandruff at a single medical center from November 2014 to February 2015. Two lashes with cylindrical dandruff were epilated from each lid (8 per patient, total 128), mounted on slides, and examined in the clinic under a slit lamp equipped with a 90D condensing lens followed by light microscopy in the pathology laboratory. All evaluations were performed by the same pathologist. Mites were identified by their characteristic morphology and movement patterns. Findings were compared between the 2 methods. RESULTS: The mean total Demodex count per lash was 1.5 ± 2.1 mites by using the slit lamp and 2 ± 2.9 mites by light microscopy. Corresponding counts per patient were 11.7 ± 9.4 and 16.1 ± 12.4. The correlation between the slit lamp and microscopy results was statistically significant, per lash (r = 0.922, P < 0.01) and per patient (r = 0.976, P < 0.01). On analysis by the more clinically relevant negative (no mites detected) or positive results (at least 1 mite detected), the accuracy of the slit-lamp examination for a single lash was 91.4% and the specificity and sensitivity were 89% and 94%, respectively; the negative predictive value was 93% [χ(1) = 87.94, P < 0.01)]. All 16 patients were positive for Demodex infestation by both methods (accuracy 100%). CONCLUSIONS: Demodex infestation in blepharitic patients with cylindrical dandruff can be confirmed using only a slit lamp and common eye clinic equipment.

Epitope-dependent blocking of the angiotensin-converting enzyme dimerization by monoclonal antibodies to the N-terminal domain of ACE: possible link of ACE dimerization and shedding from the cell surface.

In a biomembrane modeling system, reverse micelles, somatic ACE forms dimers via carbohydrate-mediated interaction, providing evidence for the existence of a carbohydrate-recognizing domain on the ACE molecule. We localized this putative region on the N-domain of ACE using monoclonal antibodies (mAbs) to seven different epitopes of ACE. Two mAbs, 9B9 and 3G8, directed to distinct, but overlapping, epitopes of the N-domain of ACE shielded the CRD. Only "simple" ACE-antibody complexes were found in the system. Five mAbs allowed the formation of "double" antibody-ACE-ACE-antibody complexes via carbohydrate-mediated interactions. The results were confirmed using the ACE N- and C-domains. Testicular ACE was unable to form carbohydrate-mediated ACE dimers in the reverse micelles, while the N-domain of ACE, obtained by limited proteolysis of the parent full-length ACE, retained the ability to form dimers. Furthermore, mAb 3G8, which blocked ACE dimerization in micelles, significantly inhibited ACE shedding from the surface of ACE-expressing cells. Galactose prevented ACE dimerization in reverse micelles and also affected antibody-induced ACE shedding in an epitope-dependent manner. Restricted glycosylation of somatic ACE, obtained by the treatment of CHO-ACE cells with the glucosidase inhibitor N-butyldeoxynojirimycin, significantly increased the rate of basal ACE shedding and altered antibody-induced ACE shedding. A chemical cross-linking approach was used to show that ACE is present (at least in part) as noncovalently linked dimers on the surface of CHO-ACE cells. These results suggest a possible link between putative ACE dimerization on the cell surface and the proteolytic cleavage (shedding) of ACE.