Our objective was to assess the importance of monitoring hand hygiene compliance (HHC) during series of successive contacts with patients or surroundings for measurement and interpretation of the compliance rates. A direct observational study of HHC was performed in four intensive care units (ICUs) and four healthcare settings with non-intensive care wards (NICWs). Hand hygiene (HH) opportunities were differentiated into two categories: extra-series opportunities (ESOs) (before or after a single contact, and before the first contact or after the last contact of a series of successive contacts) or as intra-series opportunities (ISOs) (from the opportunity following the first contact to the opportunity preceding the last in the same series). In all, 903 opportunities of HH were performed in ICUs and 760 in NICWs. The proportion of ISOs was 46.0% in ICUs and 22.9% in NICWs. The overall HHC was significantly higher in NICWs than in ICUs (61.2% vs 47.5%, P<0.00001). The HHC was significantly higher for ESOs than for ISOs (67.7% vs 28.5%, P<0.00001). The HHC for ISOs was significantly higher in ICUs (32.2% vs 19.0%, P<0.005). If the distribution of categories of HH opportunities observed in NICWs had been the same as in ICUs, the overall HHC would have been similar in NICWs (46.4%) and in ICUs (47.5%). Monitoring HHC during entire care episodes in series of successive contacts is necessary to avoid a strong overestimation of the overall compliance rates. Concurrently, comparison of compliance data should take into account the proportion of ISOs included in the evaluation study.