title: An easy and inexpensive method for quantitative analysis of endothelial damage by using vital dye staining and Adobe Photoshop software.
abstract: OBJECTIVE
  We developed a simple, practical, and inexpensive technique to analyze areas of endothelial cell loss and/or damage over the entire corneal area after vital dye staining by using a readily available, off-the-shelf, consumer software program, Adobe Photoshop. The purpose of this article is to convey a method of quantifying areas of cell loss and/or damage.
  
  
  METHODS
  Descemet-stripping automated endothelial keratoplasty corneal transplant surgery was performed by using 5 precut corneas on a human cadaver eye. Corneas were removed and stained with trypan blue and alizarin red S and subsequently photographed. Quantitative assessment of endothelial damage was performed by using Adobe Photoshop 7.0 software.
  
  
  RESULTS
  The average difference for cell area damage for analyses performed by 1 observer twice was 1.41%. For analyses performed by 2 observers, the average difference was 1.71%. Three masked observers were 100% successful in matching the randomized stained corneas to their randomized processed Adobe images.
  
  
  CONCLUSIONS
  Vital dye staining of corneal endothelial cells can be combined with Adobe Photoshop software to yield a quantitative assessment of areas of acute endothelial cell loss and/or damage. This described technique holds promise for a more consistent and accurate method to evaluate the surgical trauma to the endothelial cell layer in laboratory models. This method of quantitative analysis can probably be generalized to any area of research that involves areas that are differentiated by color or contrast.
authors.affiliation: Devers Eye Institute, Portland, OR 97210, USA.
authors.forename: Hisham A
authors.initials: HA
authors.lastname: Saad
authors.forename: Mark A
authors.initials: MA
authors.lastname: Terry
authors.forename: Neda
authors.initials: N
authors.lastname: Shamie
authors.forename: Edwin S
authors.initials: ES
authors.lastname: Chen
authors.forename: Daniel F
authors.initials: DF
authors.lastname: Friend
authors.forename: Jeffrey D
authors.initials: JD
authors.lastname: Holiman
authors.forename: Christopher
authors.initials: C
authors.lastname: Stoeger
mesh_terms: D000880:Anthraquinones; D002470:Cell Survival; D004396:Coloring Agents; D016039:Corneal Transplantation / Q000473:pathology*; D003886:Descemet Membrane / Q000601:surgery; D004728:Endothelium, Corneal / Q000473:pathology* / Q000637:transplantation; D006801:Humans; D007091:Image Processing, Computer-Assisted / Q000379:methods*; D010781:Photography / Q000379:methods*; D013194:Staining and Labeling / Q000379:methods; D014343:Trypan Blue
chemical_list: D000880:Anthraquinones; D004396:Coloring Agents; C004468:Alizarin Red S; D014343:Trypan Blue