scleral tissue posterior to the limbus is grasped with a 0.22 forcep and the globe ( eye ball ) is stabilized . a microsurgical blade is used to make a linear incision in the cornea from limbus to limbus . a 0.12 colibri forcep the anterior chamber fluid is then absorbed with a weck - cel surgical spear . a 0.12 colibri forcep grasping the cornea is used to stabilize the globe . a fine curved needle holder ( or curved dressing forceps ) is inserted behind the lens toward the posterior aspect of the globe . pressure is applied using the forceps on the external surface of the sclera , which pushes the lens forward through the corneal incision while leaving the eye wall intact . the lens - vitreous tissue is then placed into a filtered centrifugation tube containing 20 microliters of protease inhibitor cocktail ( roche ) dissolved in pbs .   continue to stabilize the globe with 0.12 colibri forceps grasping the corneal incision . a fine curved needle holder ( or curved dressing forceps ) is placed as far posterior to the globe as possible , near the optic nerve . pressure is applied using the forceps on the external surface of the sclera , which pushes the retina forward through the corneal incision . the retina - vitreous tissue is then placed into the filtered centrifuge tube containing the lens - vitreous tissue .   the filtered centrifuge tube is placed in a benchtop centrifuge . cross - sectional schematic of the human and mouse eye shows the relative smaller vitreous volume in the mouse eye due to its larger lens . protein profiles of mouse vitreous , 13.6 mg / ml ( lane 1 ) , and retina , 11.35 mg / ml ( lane 2 ) . gel electrophoresis was performed at 150 kv for 45 minutes , stained with flamingo fluorescent gel stain and visualized using a versadoc imaging system ( biorad , hercules , ca ) . measurement of total sod activity ( sod activity colorimetric aasay kit , # ab65354 , abcam , inc , cambridge , ma ) was performed on vitreous and retina collected by evisceration from dba and albino mice . this was compared to sod activity in human vitreous core and human retina collected from post - mortem human donor eyes to determine relative activity . undiluted vitreous core samples were manually aspirated using a 23-gauge needle ,   and retina was collected by flowering the eye and cutting the tissue away from the rpe / choroid complex . although the assay does not distinguish between sod isoforms , the vitreous sod activity is likely to reflect sod3 , since it is the only extracellular isoform . the retina sod activity is likely to reflect all three sod isoforms : intracellular sod1 , mitochondrial sod2 , and extracellular sod3 .   for highly sensitive enzymatic assays , including this sod assay , the possibility of cross - contamination of proteins is possible and requires further study . scleral tissue posterior to the limbus is grasped with a 0.22 forcep and the globe ( eye ball ) is stabilized . a microsurgical blade is used to make a linear incision in the cornea from limbus to limbus . a 0.12 colibri forcep a 0.12 colibri forcep grasping the cornea is used to stabilize the globe . a fine curved needle holder ( or curved dressing forceps ) pressure is applied using the forceps on the external surface of the sclera , which pushes the lens forward through the corneal incision while leaving the eye wall intact . the vitreous appears as a translucent gel and is partially adherent to the lens . the lens - vitreous tissue is then placed into a filtered centrifugation tube containing 20 microliters of protease inhibitor cocktail ( roche ) dissolved in pbs . continue to stabilize the globe with 0.12 colibri forceps grasping the corneal incision . a fine curved needle holder ( or curved dressing forceps ) is placed as far posterior to the globe as possible , near the optic nerve . pressure is applied using the forceps on the external surface of the sclera , which pushes the retina forward through the corneal incision . the retina - vitreous tissue is then placed into the filtered centrifuge tube containing the lens - vitreous tissue . cross - sectional schematic of the human and mouse eye shows the relative smaller vitreous volume in the mouse eye due to its larger lens . protein profiles of mouse vitreous , 13.6 mg / ml ( lane 1 ) , and retina , 11.35 mg / ml ( lane 2 ) . gel electrophoresis was performed at 150 kv for 45 minutes , stained with flamingo fluorescent gel stain and visualized using a versadoc imaging system ( biorad , hercules , ca ) . measurement of total sod activity ( sod activity colorimetric aasay kit , # ab65354 , abcam , inc , cambridge , ma ) was performed on vitreous and retina collected by evisceration from dba and albino mice . this was compared to sod activity in human vitreous core and human retina collected from post - mortem human donor eyes to determine relative activity . undiluted vitreous core samples were manually aspirated using a 23-gauge needle ,   and retina was collected by flowering the eye and cutting the tissue away from the rpe / choroid complex . although the assay does not distinguish between sod isoforms , the vitreous sod activity is likely to reflect sod3 , since it is the only extracellular isoform . the retina sod activity is likely to reflect all three sod isoforms : intracellular sod1 , mitochondrial sod2 , and extracellular sod3 .   for highly sensitive enzymatic assays , including this sod assay , the possibility of cross - contamination of proteins is possible and requires further study . 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 . 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 . experiments on animals were performed in accordance with the association for research and vision in ophthalmology statement for the use of animals in ophthalmic and vision research .
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 .