Little is known about the effect of various animal's signalment variables on echocardiographic reference values in the equine species. This study was performed to evaluate the effect of sex, breed, age and body weight (BW) on echocardiographic measurements in the equine species. Echocardiography was performed on 212 ponies or horses of various breeds, aged from 1 day to 37 years old (mean±SD: 7.8 ± 5.8 years), BW 38-890 kg (mean ± SD: 421 ± 133 kg), and free of cardiac disease. Fifty of those animals aged from 2 months to 35 years old (mean ± SD: 11.6 ± 6.4 years old); BW 77-662 kg (mean ± SD: 436 ± 135 kg) were also examined using the pulsed-wave Doppler mode. Standard two-dimensional and M-mode echocardiography were performed on all animals. Standard pulsed-wave Doppler examination of each cardiac valve was performed on the 50 first examined animals. Data were analysed using a general linear model including the effect of sex, age, breed and BW after logarithmic transformation of the data. Therefore, the same analysis was performed separately on animals aged ≤ 2 years-old and on older animals. All dimensional echocardiographic measurements were significantly affected by BW and most of them were significantly affected by breed, but not by sex. Only the aortic and the pulmonary artery internal diameter were significantly affected by age. None of the Doppler measurements were significantly affected by the tested variables. In conclusion, in the equine species, dimensional echocardiographic reference values should be established using regression equations as a function of BW, which could increase the diagnostic value of this leading technique in equine cardiology. Breed could also have an effect on those measurements.