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Updated: Jan 2, 2026
In Vivo Confocal Microscopy: A Standard Operating Procedure for the Detection of Demodex Mites at the Eyelid Margin
Published on: July 3, 2025
Improved Demodex diagnosis in the clinical setting using a novel in situ technique
Alex Muntz1, Christine Purslow2, James S Wolffsohn3
1Department of Ophthalmology, New Zealand National Eye Centre, The University of Auckland, New Zealand.
Purpose:
To compare existing and novel diagnostic techniques for confirming ocular Demodex infestation and to recommend the most reliable method for routine use by eye care practitioners, based on yield and clinical applicability.
Methods:
Fifteen participants with a prior Demodex blepharitis diagnosis or featuring typical cylindrical dandruff (CD) collarettes, and seven healthy controls were enrolled. Demodex presence was assessed using five techniques, applied consecutively, on a minimum of two different eyelashes on each eyelid of every participant, for each test, in situ: 1. using fine-point forceps and 25-40x biomicroscopy magnification, by eyelash rotation as proposed by Mastrota (ROT); 2. by removing cylindrical dandruff and exposing the eyelash insertion point at the lid margin (CDR); and 3. by laterally tensioning the eyelash (LET) following CDR. The typical appearance of cigar-shaped mite tails protruding from each assessed eyelash follicle was observed, and mite tails counted and averaged per participant for each assessment technique. 4. Lash epilation, and mite presence evaluated using bright-field microscopy at 10-40x magnification (EPI). 5. Finally, eyelash follicles were imaged using in vivo confocal microscopy (IVCM) and the images visually inspected for mite presence.
Results:
In the Demodex group, the highest numbers of mites/eyelash were identified by LET (3.8 ± 1.4), versus CDR (2.4 ± 1.6) and ROT (1.1 ± 1.2), alone (all p < 0.002). An average of 1.0 ± 0.8 mites/lash was identified by EPI. IVCM failed to offer unequivocal evidence of Demodex presence even in confimed cases.
Conclusions:
A novel technique for the clinical diagnosis and grading of Demodex in situ is described. By removing cylindrical dandruff and applying static, lateral tension to the eyelash without epilation, large numbers of mites are visible at the exposed eyelash follicle. The proposed method is convenient and clinically applicable, requiring only forceps and 25-40x biomicroscope magnification, and allowing rapid, efficient evaluation of large numbers of eyelashes.
