The 500 kHz VisuMax FSL (Carl Zeiss Meditec, Jena, Germany) was used to perform laser trephination. The VisuMax has a curved treatment cone, which conforms to the curvature of the human cornea and is likely to result in minimal corneal distortion on applanation. This contrasts with the Intralase FSL (Abbott Medical Optics Inc., Abbott Park, IL), which has a flat treatment cone that compresses the cornea on applanation.Briefly, each animal was prepared as described above. A small-sized curved docking cone was used in the right eye of each rabbit (N = 8). Following application of suction, the penetrating keratoplasty firing sequence was initiated using the following parameters: diameter 8 mm, energy 180 nJ, 2-μm spot and line separation, angle 90°. In the case of the FSL, the trephination incision advances from the posterior to the anterior cornea. Full-thickness trephination was possible with the FSL without collapse of the anterior chamber as small tissue bridges prevented immediate separation of the incision plane. No attempt was made to manually separate these bridges. Following completion of the laser sequence, each rabbit had subconjunctival cefuroxime, a bandage contact lens, and tarsorraphy performed as described above.
The FSL trephination group was subdivided into two groups, with animals for sacrifice at 4 hours (N = 4) and 3 days (N = 4). Endothelial cell damage was assessed in the left eyes of rabbits in the 3-day group by repeating the FSL trephination procedure as described above, before sacrifice of the animals. Viscoelastic was not used as there was no chamber collapse. Corneoscleral rims were excised for immunohistochemistry (right eyes, N = 4) and electron microscopy (left eyes, N = 4).