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To quantify consistency of endothelial cell density (ECD) measurements among technicians in a single US eye bank operating under typical operating conditions.In this retrospective analysis of 51 microscopy technicians using a semiautomated counting method on 35,067 eyes from July 2007 to May 2015, technician- and date-related marginal ECD effects were calculated using linear regression models. ECD variance was correlated with the number of specular microscopy technicians.Technician mean ECDs ranged from 2386 ± 431 to 3005 ± 560 cells/mm2. Nine technicians had statistically and clinically significant marginal effects. Annual mean ECDs adjusted for changes in technicians ranged from 2422 ± 433 to 2644 ± 430 cells/mm2. The period of 2007 to 2009 had statistically and clinically significant marginal effects. There was a nonstatistically significant association between the number of technicians and ECD standard deviation.There was significant ECD variability associated with specular microscopy technicians and with the date of measurement. We recommend that eye banks collect data related to laboratory factors that have been shown to influence ECD variability.