PURPOSE: To describe a novel method for analyzing Descemet membrane endothelial keratoplasty (DMEK) graft damage after implantation into human cadaveric donor eyes and to compare results achieved by performing DMEK with a surgeon's long-established technique compared with those of an unfamiliar technique. METHODS: Eight DMEK grafts were implanted into previously frozen human cadaveric eyes. Four grafts were implanted using a Straiko injector and tap technique familiar to the surgeon (C.S.S., 3-yr experience), and 4 grafts were implanted using the Tan EndoGlide and "donor mat device" pull-through technique new to the surgeon. After implanting a DMEK graft and attaching it to the recipient stroma with an air bubble tamponade, the corneoscleral cap was "recovered" from the cadaveric globe using standard techniques. The DMEK graft was stained with Calcein-AM. After staining, a 9.5-mm stromal "carrier button" was punched, and the carrier and graft were transferred to a microscope slide. Grafts were imaged and analyzed using FIJI trainable segmentation. RESULTS: Donor graft characteristics were similar between both groups. Grafts implanted using the surgeon's routine technique showed an average endothelial cell loss (ECL) of 31% ± 4% (n = 3). Grafts implanted using the technique unfamiliar to the surgeon showed an average ECL of 47% ± 24%, but with a trend toward improvement (1 = 76%, 2 = 65%, 3 = 32%, 4 = 17% ECL). CONCLUSIONS: Our proof-of-principle experiment shows that this imaging approach enables quantification of ECL caused by different instruments and surgical techniques after graft implantation. We have used this method to visualize the learning curve of 1 surgeon when learning a new surgical technique.