The dissection technique illustrates evisceration of the vitreous, retina, and lens from the mouse eye, separation by centrifugation, and characterization with protein assays.
While the mouse retina has emerged as an important genetic model for inherited retinal disease, the mouse vitreous remains to be explored. The vitreous is a highly aqueous extracellular matrix overlying the retina where intraocular as well as extraocular proteins accumulate during disease.1-3 Abnormal interactions between vitreous and retina underlie several diseases such as retinal detachment, proliferative diabetic retinopathy, uveitis, and proliferative vitreoretinopathy.1,4 The relative mouse vitreous volume is significantly smaller than the human vitreous (Figure 1), since the mouse lens occupies nearly 75% of its eye.5 This has made biochemical studies of mouse vitreous challenging. In this video article, we present a technique to dissect and isolate the mouse vitreous from the retina, which will allow use of transgenic mouse models to more clearly define the role of this extracellular matrix in the development of vitreoretinal diseases.
Transgenic mice are an important model for investigating retinal and vitreoretinal disease.6-10 The mouse vitreous body, however, comprises a significantly smaller proportion of the eye in comparison to the human vitreous due to the large lens in the mouse eye.5 This makes it difficult to isolate and purify the mouse vitreous. Understanding the protein changes associated with the vitreous during diseases such as proliferative diabetic retinopathy, age-related macular degeneration, uveitis, and retinal detachment will give insight into the mechanisms by which they progress. This visualized experimental protocol provides a means to obtain and purify the mouse vitreous body, while maximizing the protein yield for enzymatic and proteomic analyses.
The authors have nothing to disclose.
Funding was provided by Fight for Sight and Research to Prevent Blindness.
Name | Company | Catalog Number | Comments |
---|---|---|---|
15°, BD Beaver Microsurgical Blade | Becton-Dickinson | 374881 | |
PBS, pH 7.4 | Invitrogen | 70011-044 | |
0.22 Fine-Castroviejo Suturing Forceps | Storz Ophthalmics | E1805 | |
0.12 Colibri forceps | Storz Ophthalmics | 2/132 | |
Microcon Centrifugal Filter Ultracel YM-100 or YM-50 | Millipore | 42412, 42415 | |
SOD Activity Colorimetric Assay Kit | Abcam | Ab65354 | |
Weck-Cel surgical spears | Medtronic | 0008680 | |
Protease inhibitor cocktail | Roche | 11 836 170 001 | |
Flamingo fluorescent gel stain | Bio-Rad | 161-04910 |