To assess 5-year Descemet's stripping endothelial keratoplasty (DSEK) graft survival and endothelial cell loss in the surviving grafts. Retrospective, interventional case series. One hundred sixty-five eyes of 149 patients treated with primary DSEK. Donor corneal-scleral rims were dissected manually or with a microkeratome and were cut with a trephine. The graft was folded endothelial side inward and was inserted with forceps through a 5-mm incision. The cumulative probability of secondary graft failure was calculated using Kaplan-Meier survival analysis and the log-rank test. Endothelial cell density (ECD) was determined from baseline preoperative donor and 1-year, 3-year, and 5-year postoperative central endothelial images. Graft survival and ECD at 5 years. The median recipient age was 71 years (range, 22-90 years) and 62% were female. Eighteen eyes (11%) were treated for pseudophakic or aphakic corneal edema and 147 eyes (89%) were treated for Fuchs' dystrophy. The cumulative 5-year survival rate was significantly lower in pseudophakic or aphakic corneal edema eyes (76%) versus Fuchs' eyes (95%; P = 0.0087). In particular, the 5-year survival rate was reduced significantly in eyes with prior glaucoma shunt or trabeculectomy surgery vs. those without (40% vs. 95%; P<0.0001). The causes of secondary graft failure were endothelial decompensation in 6 eyes (3.6%) and unsatisfactory corrected distance acuity (20/60 to 20/100) in 4 eyes (2.4%). No grafts experienced traumatic wound rupture or failed as a result of ocular surface complications. The median 5-year endothelial cell loss was 53% (range, 7.5%-89%). The 5-year graft ECD was correlated weakly with the baseline donor ECD (r = 0.22 and P = 0.04) and was not significantly correlated with recipient gender (P = 0.075), age (P = 0.85), or diagnosis (P = 0.78). The 5-year graft survival rates for DSEK were similar to those reported for penetrating keratoplasty in the multicenter Cornea Donor Study (95% vs. 93% for Fuchs' dystrophy and 76% vs. 73% for pseudophakic or aphakic corneal edema). Furthermore, the 5-year endothelial cell loss after DSEK compared favorably with that measured after penetrating keratoplasty in the Cornea Donor Study (53% vs. 70%).