title: A comparing study of quantitative staining techniques for retinal neovascularization in a mouse model of oxygen-induced retinopathy.
abstract: OBJECTIVE
  To explore an efficient, practical and objective quantitative method to evaluate the retinal neovascularization in mouse model of oxygen induced retinopathy (OIR).
  
  
  METHODS
  Thirty C57BL/6J mice were explored in OIR model procedure. Eyes were removed for different staining methods including: (1) HE staining; (2) immunohistochemistry with Griffonia Simplicifolia Lectin(GSL); (3) Immunofluorescence with FITC labeled CD31 antibody; (4) Two-step immunofluorescence with purified-CD31 antibody; (5) FITC-Dextran perfusion combined with two-step purified-CD31immunofluorescence. Images of the retinal vasculature were analyzed by imaging software.
  
  
  RESULTS
  GSL immunohistochemistry could clearly demonstrate the deep and superficial capillary beds. FITC labeled CD31 Immunofluorescence was blurring with high fluorescence background which was hard to distinguish retinal neovascularization in some area. Excellent detail of neovascularization and preexistent retinal vessels was provided in two-step Purified-CD31 immunofluorescence group.
  
  
  CONCLUSIONS
  GSL immunohistochemistry can clearly demonstrate neovascularization tufts in deep and superficial capillary beds. Immunofluorescence of specific antigen CD31 on vascular endothelium can selectively label the neovascularization of mouse retina. When combined with computer analysis software, it is an effective and objective quantitative method to evaluate the retinal neovascularization in OIR mouse model.
authors.affiliation: Key Lab of Ophthal Ministry of Edu, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou 510060, Guangdong Province, China.
authors.forename: Xiao-Ling
authors.initials: XL
authors.lastname: Liang
authors.forename: Jie
authors.initials: J
authors.lastname: Li
authors.forename: Fang
authors.initials: F
authors.lastname: Chen
authors.forename: Xiao-Yan
authors.initials: XY
authors.lastname: Ding
authors.forename: Xiu-Xia
authors.initials: XX
authors.lastname: Yang
authors.forename: Liao-Xu
authors.initials: LX
authors.lastname: Long