title: Cell-by-cell alignment of repeated specular microscopy images from the same eye.
abstract: OBJECTIVE
  Modern specular microscopes (SM) robustly depict the same central area of the corneal endothelium at different time points through a built-in fixation light. However, repeated image acquisitions slightly shift and rotate because of minute changes in head position in the chin and forehead rest. This prevents the manual retrieval of individual corneal endothelial cells (CECs) in repeated measurements because SM images usually lack obvious landmarks. We devised and validated an image registration algorithm that aligns SM images from the same eye to make corresponding CECs coincide.
  
  
  METHODS
  We retrospectively selected 27 image pairs for the presence of significant image overlap. Each image pair had been recorded on the same day and of the same eye. We applied our registration method in each image pair. Two observers independently validated, by means of alternation flicker, that the image pairs had been correctly aligned. We also repeatedly applied our registration method on unrelated image pairs by randomly drawing images and making certain that the images did not originate from the same eye. This was done to assess the specifity of our method.
  
  
  RESULTS
  All automated registrations of the same-day and same-eye image pairs were accurate. However, one single image incorrectly failed to trigger the non-match diagnosis twice in 81 registration attempts between unrelated images. As it turned out, this particular image depicted only 73 CECs. The average number of CECs was 253 (range 73-393).
  
  
  CONCLUSIONS
  Repeated non-contact SM images can be automatedly aligned so that the corresponding CECs coincide. Any successful alignment can be considered as proof of the retrieval of identical CECs as soon as at least 100 CEC centroids have been identified. We believe our method is the first to robustly confirm endothelial stability in individual eyes.
authors.affiliation: University Eye Hospital, Cornea Reading Center, Freiburg, Germany. daniel.boehringer@uniklinik-freiburg.de
authors.forename: Daniel
authors.initials: D
authors.lastname: Böhringer
authors.forename: Stefan
authors.initials: S
authors.lastname: Lang
authors.forename: Thomas
authors.initials: T
authors.lastname: Reinhard
mesh_terms: D000465:Algorithms; D042783:Endothelial Cells / Q000502:physiology; D004728:Endothelium, Corneal / Q000502:physiology*; D006801:Humans; D008853:Microscopy / Q000379:methods*; D012189:Retrospective Studies