Whole Vitreous Humor Dissection for Vitreodynamic Analysis

The overall goal of this procedure is to show an effective technique for isolating whole intact vitreous cores and cortexes from postmortem and nucleated porcine eyes. This is accomplished by first dissecting away the peri limbal conjunctiva and exposing the sclera. In the second step, a full thickness scleral flap is created and expanded.

The choroid is then separated away from the sclera and the choroid retina tissue is peeled away from the vitreous. Ultimately, the whole intact vitreous core and cortex dissection can be used to study the physical, biomechanical and chemical properties of the vitreous humor. The main advantage of this technique over other existing methods like partial vitreous dissections, is that this technique results in the isolation of the whole Intact vi is core and cortex of maintaining vitreous anatomical structure Begin by pinning the enucleated eye through the surrounding excess tissue onto a styrofoam dissection board.

Once the eye is secure, use fine forceps and micro scissors to incise the conjunctiva at the limbus and bluntly dissect it off the sclera. Cut the conjunctiva along the limbus as the blunt dissection proceeds to allow further dissection. Then remove all but a small amount of conjunctiva all the way around the eye to expose as much sclera as possible.

Next, use the scalpel to gently make a scleral incision about five millimeters parallel and three millimeters posterior to the limbus until the darkly pigmented choroid is visible to create a scleral flap. Then carefully blunt, dissect the tissue along the plane between the sclera and the choroid until it is possible to push the choroid gently inward to enlarge the space between the two tissues. Using sharp micro scissors, make another scleral cut perpendicular to the first to create a T-shaped incision.

Then continue to bluntly dissect circumferentially to remove the scleral tissue away from the underlying choroid, and gently push the choroid away from the sclera to enlarge the flap. Gently blunt, dissect the choroid away from the sclera until the incision made perpendicular to the limbus reaches the optic nerve and the incision made parallel to the limbus is at least 45 degrees of the eye's circumference. Use this flap to repeat the blunt dissection on the other side of the eye.

Then cut away both scleral flaps to expose a large area of the choroid. Now, continue the incision around the eye's circumference blunt. Dissect the choroid away from the sclera, and remove the remaining choroid retina tissue within the exposed region.

Make an incision into the choroid starting from the optic nerve. Then use forceps to gently peel the tissue off of the underlying vitreous body. Continue blunt dissecting and peeling away the choroid until the whole intact vitreous is obtained.

Then use the scleral rim attached to the cornea to position the whole vitreous in the desired location. At this point, the vitreous should still be attached to the anterior sclera and lens. Finally, blunt, dissect the vitreous from the inside of the sclera and around the lens.

To remove all of the sclera, use a blunt tool to scoop the lens away from the vitreous as necessary. Following the protocol as just demonstrated will lead to a successful vitreous dissection with the core and cortex intact as evident from the residual pieces of retina adhered to the vitreous cortex. In this image, the intact whole vitreous humor can then be used in several ways for specific viremic experiments.

For example, here the diffusion rate of oxygen in an intact vitreous humor and its corresponding time constant was studied in the presence of the vitreous cortex. The diffusion of the oxygen occurred over a longer interval. Don't forget that working with sharps can be hazardous, and precautions such as proper handling and disposal of s must always be taken while performing this procedure.