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O. Delgado, L. Burgess, S. Poor, W. Breegi; Novel Vitreous Collection Method in the Rat . Invest. Ophthalmol. Vis. Sci. 2005;46(13):5388.
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Purpose: Most of the experimental protocols designed to study pathological conditions in the posterior segment of the eye require the in situ injection of the therapeutic agent, either in the vitreous or in the subretinal space. An intravitreal injection provides the most direct approach to delivering drugs to the tissues of the posterior segment, and therapeutic tissue drug levels can be achieved. The volume and reproducibility of a vitreal collection method could influence the results of studies evaluating the biodistribution of drugs in the eye. To date, no satisfactory vitreal collection method has been developed. Several vitreal extraction approaches have been evaluated, but are suboptimal as they are inconsistent (operator dependent), time consuming and labor intensive (i.e., freeze–dissections, temperature–dependent enzymes). We have developed and validated a vitreal collection technique that is rapid, quantitative and reproducible. Methods: Eyes from anesthetized adult Long Evans rats of different ages (i.e., 1 months, 2.5 months, 4 months) were enucleated using Westcott scissors and the cornea, iris and lens removed. Residual aqueous humor was removed from the posterior cup before placement up side down in a modified Microcon YM–30 centrifugal device. The anterior capsule was ruptured using two fine forceps and centrifuged to separate the vitreous from the remaining tissue. Results:A consistent vitreal volume in three separate experiments was obtained. The average vitreal volume collected from the three experiments was 4 months = 23.25 µl (±2.08); 2.5 months = 20.68 (±1.43) and 1 months = 16.7 (±1.05). To assess the potential extraction of liquid from the retina during the procedure, 10 retinas were centrifuged three times under the same conditions with no considerable change in the wet weight of the retinas. Conclusions: Several methods (e.g., intravitreal and subconjuntival injections) are currently used to deliver therapeutic agents into the eye. We have developed a reproducible and quantifiable method of vitreal collection in the rat. This method is instrumental in the evaluation of compound biodistribution in the eye.
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