BACKGROUND: Since 2004, we have promoted alcohol-based hand rubbing (HR) with an e-learning program (ELP) among hospital staff. This study sought to determine whether an ELP improves adherence to correct HR. METHODS: This was a cohort study of staff members at Aarhus University Hospital, Skejby, Denmark who completed the ELP and were repeatedly observed for correct HR before and after clinical procedures in 2006 and/or 2007. RESULTS: Of the 496 participants, 13% completed the ELP in both 2006 and 2007, 29% completed the ELP only in 2006, 15% completed the ELP only in 2007, and 43% never completed the ELP. Compared with noncompleters, completers of the 2006 and 2007 ELP had a significantly higher adherence to correct HR both before clinical procedures (odds ratio [OR] = 1.54; 95% confidence interval [CI] = 1.11 to 2.13) and after clinical procedures (OR = 1.40; 95% CI = 1.03 to 1.89). Time since completing the ELP seemed to be inversely associated with adherence to correct HR. CONCLUSION: Completion of an ELP may have a positive impact on the performance of correct HR. The demands of lifelong education and training of hospital staff may call for the use of an ELP as a supplement to existing efforts aimed at improving HR to help prevent health care-related infections.