After transconjunctival enucleation of the rabbit eye, immediately place in 50 milliliters of 4%paraformaldehyde and PBS on ice. After 15 minutes, transfer the eye to a 100 millimeter dissection dish and fill the dish with PBS to prevent the eye from drying out. Under a dissection microscope, Using a surgical blade, start making an incision one millimeter anterior to the limbus, keeping it parallel to the iris plane.
Insert curved scissors into the initial incision and continue cutting circumferentially, maintaining a one millimeter distance from the limbus. Then remove the cornea with forceps and gently wipe away PBS With a laboratory wipe. Place the cornea in 30%sucrose solution at room temperature for cryoprotection.
Make an initial cut with curved scissors through the iris. Then cut circumferentially around the eye, similar to the cornea removal, to separate the iris from the rest of the tissue. Remove the iris with forceps and gently wipe away excess PBS with the laboratory wipe.
Place the iris in 30%sucrose solution at room temperature for cryoprotection. After removing the cornea and iris, place the eye in 50 milliliters of 4%paraformaldehyde and put it on a shaker for gentle agitation. Leave the eye in 4%paraformaldehyde at four degrees Celsius overnight.
The next day, place the eye in a dissection dish and fill the dish with PBS. Under the dissection microscope, use forceps to carefully grasp the anterior lens capsule. Then carefully insert the scissors into the posterior chamber, orienting the blades posteriorly along the lens.
Make an initial cut through the lens zonules with curved scissors. Continue making small cuts in a circumferential pattern through the lens zonules. To confirm lens removal, gently grasp the anterior lens capsule with forceps and attempt to lift it.
Upon separation, place the lens in 30%sucrose solution at room temperature for cryoprotection. Place the eye in 50 milliliters of 30%sucrose and PBS at four degrees Celsius for 24 to 48 hours for cryoprotection. To expedite cryoprotection, replace the initial sucrose solution with a fresh one after eight to 12 hours.
