BACKGROUND: Hand-washing is regarded as a potentially important behavior for preventing transmission of respiratory infection, particularly during a pandemic. OBJECTIVE: The objective of our study was to evaluate whether a Web-based intervention can encourage more frequent hand-washing in the home, and to examine potential mediators and moderators of outcomes, as a necessary first step before testing effects of the intervention on infection rates in the PRIMIT trial (PRimary care trial of a website based Infection control intervention to Modify Influenza-like illness and respiratory infection Transmission). METHODS: In a parallel-group pragmatic exploratory trial design, 517 nonblinded adults recruited through primary care were automatically randomly assigned to a fully automated intervention comprising 4 sessions of tailored motivational messages and self-regulation support (n = 324) or to a no-intervention control group (n = 179; ratio 2:1). Hand-washing frequency and theory of planned behavior cognitions relating to hand-washing were assessed by online questionnaires at baseline (in only half of the control participants, to permit evaluation of effects of baseline assessment on effect sizes), at 4 weeks (postintervention; all participants), and at 12 weeks. RESULTS: Hand-washing rates in the intervention group were higher at 4 weeks than in the control group (mean 4.40, n = 285 and mean 4.04, n = 157, respectively; P