The posterior surface of the corneo-scleral rim was gently touched with a cellulose sponge (Figure 6.1a), to damage or remove the donor endothelium. Trypan blue 0.06% in phosphate buffered sodium chloride (VisionBlue, D.O.R.C. International, The Netherlands) was applied onto the rim to stain Descemets membrane (Figure 6.1b), and the posterior surface of the rim was gently swabbed to completely remove the blue stained Descemets membrane and the endothelium (Figure 6.1c). Corneo-scleral rims were mounted epithelial side down onto a punch block, and a 0.25 mm oversized donor button was punched out with a punch trephine (Ophtec, Groningen, The Netherlands).

Figure 6.1. Removal of the donor Descemets membrane as viewed from the endothelial side of a corneo-scleral rim. (a) The posterior surface of the rim is gently touched with a sponge, to damage the endothelium. (b) Trypan blue is applied to stain the damaged endothelial cells and Descemets membrane. (c) The blue stained Descemets membrane is stripped from the posterior stroma by gently swabbing the posterior surface with the sponge. A flap of Descemets membrane (asterisks) is visible; the arrowheads point to the edge of the flap that is still attached to the posterior stroma. (Source: Cornea 2000;19:427-432).