Purpose: To compare outcomes of manual Descemet membrane endothelial keratoplasty (M-DMEK) and femtosecond laser–assisted Descemet membrane endothelial keratoplasty (F-DMEK) in treatment of penetrating keratoplasty (PK) graft failure. Methods: SETTING: Retrospective, interventional comparative case series. PATIENT POPULATION: Included were all patients with a failed PK graft who underwent either F-DMEK (10 eyes of 10 patients) or M-DMEK (29 eyes of 29 patients) at Toronto Western Hospital and the Kensington Eye Institute between 2014 and 2019, and had 6 months of postoperative follow-up. OUTCOME MEASURES: Rates of graft detachment, rebubbling, rejection and failure, best spectacle-corrected visual acuity (BSCVA), and endothelial cell (EC) density. Results: Rate of significant graft detachment (detachment requiring either rebubble or repeat keratoplasty) was 10.0% in F-DMEK and 65.5% in M-DMEK (P = .003). Rebubble rate was 10.0% in F-DMEK and 58.6% in M-DMEK (P = .011). Primary failure rate was 0% in F-DMEK and 27.6% in M-DMEK (P = .086). Rates of rejection and secondary failure did not differ between the groups (P = 1.000 for both). In a multivariable analysis, F-DMEK was found to be the only independent factor significantly associated with reduced postoperative detachment. Postoperative BSCVA at 6 months returned to prefailure levels in both groups, with no significant difference between the groups at any time point. EC loss rates were 43.8% in F-DMEK and 38.0% in M-DMEK at 6 months (P = .453). Conclusions: F-DMEK is a safe and effective procedure in failed PK patients, with outcomes comparable to M-DMEK, and with reduced detachment and rebubble rates. A trend towards reduced primary failure should be further studied.