Gnotobiotic Operations and Assembly for Development of Germ-Free Animal Model of Laser-Induced Choroidal Neovascularization

Asadolah Movahedan, Hugo Barba, Melanie Spedale, Nini Deng, Donna Arvans, Urooba Nadeem, Vanessa Leone, Eugene B.chang, Betty Theriault, Dimitra Skondra

Research output: Contribution to journalArticlepeer-review

3 Scopus citations


Purpose: Compelling new evidence reveals a close link between the gut microbiome and the pathogenesis of neovascular age-related macular degeneration (nAMD). Germfree (GF) animal models are the current gold standard for studying host the microbe interactionsinvivo;yet,noGFanimalmodelsofnAMDareavailabletoday.Thisprotocol describes gnotobiotic operations and assembly for a laser-induced choroidal neovascularization (CNV) model in GF mice to study the gut microbiome in neovascular AMD. Methods: We developed a step-wise approach to performing retinal laser photocoagulation in GF C57BL/6J mice that were bred and maintained at the gnotobiotic facility. Following a strict sterility protocol, we administered laser photocoagulation via an Argon 532-nm laser attached to a customized slit-lamp delivery system. Sterility was confirmed by weekly fecal cultures and reverse transcriptase–polymerase chain reaction. Results: The experiment was repeated twice at different time points using seven mice (14 eyes). Stool cultures and RT-PCR remained negative for 14 days post-procedure in all mice. Lectin immunostaining performed on choroidal flatmounts confirmed the presence of CNV lesions 2 weeks after laser treatment. Conclusions: We established a GF mouse model of nAMD with detailed guidelines to deliver retinal laser in GF mice maintaining sterility after the laser procedure. Translational Relevance: To our knowledge, this is the first protocol that describes a GF murine model of laser-induced CNV. In addition to nAMD, this animal model can be used to investigate host–microbial interactions in other eye diseases with laser-induced mouse models such as glaucoma and retinal vein occlusion.

Original languageEnglish (US)
Article number14
JournalTranslational Vision Science and Technology
Issue number9
StatePublished - Aug 2021
Externally publishedYes


  • age-related macular degeneration
  • animal model
  • choroidal neovascularization
  • germ-free
  • gnotobiotic
  • microbiome

ASJC Scopus subject areas

  • Biomedical Engineering
  • Ophthalmology


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