Objective: A number of small portable systems that can measure HRV are available to address questions related to autonomic regulation in free-living subjects. However, ambulatory HRV measurements obtained through use of these systems have not previously been validated against standard clinical measurements such as Holter recordings. The objective of this study was to validate HRV obtained using a commonly used system, Actiheart, during occupational and leisure-time activities.Method: Full-day ambulatory electrocardiography (ECG) signals were recorded from 8 females simultaneously using Actiheart and Holter recorders, and signals were processed to RR-interval time series. Segments of 5-minute duration were sampled every 30 minutes, and spectral components of the heart rate variability were calculated. Actiheart and Holter values were compared using Deming regression analysis and Bland-Altman plots.Results: In total, 489 segments were available with an HRV value from both Actiheart and Holter recordings after filtering out segments with >10% interpolated beats. No systematic differences between Actiheart and Holter HRV were found. The random deviations between Actiheart and Holter were comparable to the repeatability standard deviation between consecutive Holter measurements.Discussion: The results show that Actiheart is suited as a stand-alone ambulatory method for heart rate variability monitoring during occupational and leisure-time activities.