Despite its extensive use in physiological and clinical research, the analysis of HRV (heart rate variability) is still poorly supported by rigorous reliability studies. The main aim of the present study was to perform an in-depth assessment of absolute and relative reliability of standard indexes of HRV from short-term laboratory recordings. In 39 healthy subjects [mean age (min–max): 38 (26–56) years; 18 men and 21 women] we recorded 5 min of supine ECG during spontaneous and paced (15 breaths/min) breathing. The test was repeated on the next day under the same conditions. From the RR intervals we computed standard indexes of HRV: SDNN (S.D. of RR interval values), RMSSD (root-mean-square of successive RR interval differences), LF (low frequency) and HF (high frequency) power (absolute and normalized units) and LF/HF. Absolute reliability was assessed by 95% limits of random variation; relative reliability was assessed by the ICC (intraclass correlation coefficient). The sample size needed to detect a mean difference ≥30% of the between-subject S.D. was also estimated. Although there was no significant mean change between the two tests, we found that in individual subjects the second measurement was as high/low as 1.9/0.5 times (SDNN, best case) and 3.5/0.3 times (LF/HF, worst case) the first measurement, due to pure random variation. For most parameters the ICC was >0.8 (range 0.65–0.88). The estimated sample size ranged from 24–98 subjects. Reliability indexes tended to improve during paced breathing. We conclude that short-term HRV parameters are subject to large day-to-day random variations. Random error, however, represents a limited part of the between-subject variability; therefore observed differences between individuals mostly reflect differences in the subjects' error-free value rather than random error. Overall, paced breathing improves reliability.

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