The use of heart rate measures in research requires accurate detection and timing of beat-to-beat values. Numerous technologies are available to researchers; however, benchmarking of a specific apparatus is seldom conducted. Since heart rate variability provides a portal to the neural regulation of the heart, accurate detection and timing of beat-to-beat values is essential to both basic physiological research and the clinical application of heart rate variability measures. The current study evaluated the accuracy of an ambulatory system, the LifeShirt (Vivometrics), relative to a standard laboratory-based heart rate monitoring equipment (Biopac), during baseline and exercise conditions. LifeShirt performed equivalently to the Biopac during both conditions, experienced few errors of detection, generated similar times between sequential heart periods, and produced similar summary indices of heart rate and heart rate variability.