Limbal Approach-Subretinal Injection of Viral Vectors for Gene Therapy in Mice Retinal Pigment Epithelium.
The eye is a small and enclosed organ which makes it an ideal target for gene therapy. Recently various strategies have been applied to gene therapy in retinopathies using non-viral and viral gene delivery to the retina and retinal pigment epithelium (RPE). Subretinal injection is the best approach to deliver viral vectors directly to RPE cells. Before the clinical trial of a gene therapy, it is inevitable to validate the efficacy of the therapy in animal models of various retinopathies. Thus, subretinal injection in mice becomes a fundamental technique for an ocular gene therapy. In this protocol, we provide the easy and replicable technique for subretinal injection of viral vectors to experimental mice. This technique is modified from the intravitreal injection, which is widely used technique in ophthalmology clinics. The representative results of RPE/choroid/scleral complex flat-mount will help to understand the efficacy of this technique and adjust the volume and titer of viral vectors for the extent of gene transduction.
Department of Biomedical Sciences, Seoul National University College of Medicine; FARB Laboratory, Biomedical Research Institute, Seoul National University Hospital.
Sung Wook
SW
Park
FARB Laboratory, Biomedical Research Institute, Seoul National University Hospital.
Jin Hyoung
JH
Kim
College of Life Sciences, Gwangju Institute of Science and Technology.
Woo Jin
WJ
Park
Department of Biomedical Sciences, Seoul National University College of Medicine; FARB Laboratory, Biomedical Research Institute, Seoul National University Hospital; Department of Ophthalmology, Seoul National University College of Medicine; steph25@snu.ac.kr.
Jeong Hun
JH
Kim
D000256:Adenoviridae / Q000235:genetics*; D000818:Animals; D000229:Dependovirus / Q000235:genetics*; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000008:administration & dosage*; D049452:Green Fluorescent Proteins / Q000008:administration & dosage / Q000096:biosynthesis / Q000235:genetics; D016086:Lentivirus / Q000235:genetics*; D016850:Limbus Corneae; D008164:Luminescent Proteins / Q000008:administration & dosage / Q000096:biosynthesis / Q000235:genetics; D051379:Mice; D012164:Retinal Diseases / Q000235:genetics / Q000628:therapy*; D055213:Retinal Pigment Epithelium; D014161:Transduction, Genetic / Q000379:methods; D000097573:Red Fluorescent Protein
D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins; D000097573:Red Fluorescent Protein