Background/aims: To evaluate in vivo three-dimensional (3-D) confocal laser scanning microscopy (CLSM) as a technique for visualising the corneal surface and epithelium.
Methods: Ten human corneas (three from healthy volunteers, three with bullous keratopathy, three from patients following penetrating keratoplasty, and one with corneal erosion) were examined by 3-D CLSM. A novel polymethyl methacrylate (PMMA) contact cap was designed to minimise artefacts due to applanation pressure.
Results: 3-D reconstruction and different visualisation techniques (volume rendering, cross-section, en face view, oblique section and surface reconstruction) were performed to demonstrate alterations to corneal surface and epithelium. Image quality (cell identification, motion blur, absence of compression artefacts, imaging of superficial structures and of subepithelial nerve plexus) was considerably superior to that obtained using a conventional contact cap with a planar surface.
Conclusions: 3-D CLSM permits in vivo visualisation and analysis of the corneal surface and of spatial arrangement at the cellular level in epithelium in normal and pathological corneas. The novel design of the contact cap minimises artefacts due to applanation pressure and improves the image quality of epithelial structures. The method provides a basis for further in vivo studies of alterations to corneal surface structure and its cellular arrangement.
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Competing interests: AZ, JS and RFG are consultants to Heidelberg Engineering, Heidelberg, Germany.
Funding: The study was supported in part by the DFG (Transregio 37, Micro- und Nanosystems in Medicine—Reconstruction of Biological Functions) and by an internal grant from the University of Rostock (FORUN N 889009). The PMMA caps were manufactured in collaboration with Acri.Tec AG (Berlin, Germany).
Ethics approval: Ethics approval was provided by the Ethics Committee of the Medical Faculty, University of Rostock.
Patient consent: Obtained.