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Sachin Parikh, Andrew Le, Julian Davenport, Michael B Gorin, Anna Matynia, Steven Nusinowitz; An alternative and validated method for injection into the subretinal space via a transcleral posterior approach. Invest. Ophthalmol. Vis. Sci. 2017;58(8):4108.
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© ARVO (1962-2015); The Authors (2016-present)
Subretinal injections are the most common technique for delivering therapeutic agents to photoreceptors and the retinal pigment epithelium. We describe an alternative injection method in mice that successfully targets the subretinal space with minimal collateral damage and fast recovery times.
A lateral conjunctival incision was made in anesthetized male and female adult mice. The surrounding connective tissue was resected 0.5mm temporal to the optic nerve, exposing the injection site and avoiding the orbital blood sac. Fluorescein (0.3µl, n=18; 0.5µl, n=8; or 1.0µl, n=5 at 0.01%) was injected through a sclerotomy into the subretinal space (n=31 eyes in total). The location and topographic distribution of fluorescein was visualized by fundus imaging. Retinal thickness (Bruch’s membrane to the inner limiting membrane) was measured prior to injection, immediately after, and 2- and 4- weeks post injection using spectral domain optical coherence tomography. Retinal thickness measures were obtained at the injection site at adjacent areas. Rod-mediated retinal function was evaluated concurrently using electroretinography (ERG).
No unintended retinal detachments, punctures of the neurosensory retina, leakage into the vitreous cavity, or lens nicking was observed. In addition, no evidence of an inflammatory response, uveitis, or post-surgical infection was present. All detachments were completely resolved 2-weeks post-injection. Total retinal thickness showed an average 6.5 ± 1.9% thinning at the injection site (pre=196µm ± 1, n=31; 4-weeks=183µm ± 4, n=31, p=0.001) with no significant thinning distal to the injection site at 4-weeks post injection. Rod-mediated ERGs showed an average 9.6 ± 3.5% decrease in a-wave amplitude (pre=-356 ± 9 µV; 4-weeks=-324 ± 9 µV; p=0.013) and a trend towards reduced b-wave amplitude (pre=600 ± 17 µV; 4-weeks=552 ± 18 µV; p=0.055) at 4-weeks post injection.
Due to the high efficacy, minimal damage and fast recovery, this method is recommended for targeting viral vectors, pharmacological agents, or induced pluripotent stem cells to the subretinal space.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
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