BackgroundDifferentiating innocent cardiac murmurs from murmurs caused by congenital cardiac anomalies can be challenging with auscultation alone in asymptomatic puppies.HypothesisPlasma N‐terminal pro‐B‐type natriuretic peptide (NT‐proBNP) concentrations and phonocardiograms recorded by an electronic stethoscope can differentiate innocent from pathologic cardiac murmurs.AnimalsA total of 186 client‐owned asymptomatic dogs: 135 Cairn Terriers (age: 45–124 days), 20 adult Cairn Terriers (age: 7.5 months to 13.5 years), and 31 puppies of various breeds (age: 29–396 days).MethodsStudy design is a cross‐sectional survey. Each dog was auscultated, and when a cardiac murmur was heard, a phonocardiogram was recorded and an echocardiogram was performed. Plasma NT‐proBNP concentrations were measured by a single laboratory by an ELISA.ResultsNo significant (P = .41) difference in plasma NT‐proBNP levels was found between puppies without a murmur and puppies with an innocent murmur (median 300 versus 326 pmol/L), and between clinically healthy adult Cairn Terriers and Cairn Terrier puppies. Plasma NT‐proBNP levels in puppies with a congenital heart disease were significantly (P < .001) higher than those in puppies with innocent murmurs (median 1,102 versus 326 pmol/L). However, some puppies with severe pulmonic stenosis did not have increased plasma NT‐proBNP levels. On phonocardiograms, innocent murmurs had a significantly (P < .001) shorter “murmur‐to‐systole duration ratio” than the abnormal ones (median 66 versus 100%). The “murmur‐to‐S1 (first cardiac sound) amplitude ratio” was significantly (P < .001) lower of the innocent murmurs compared with that of the abnormal ones (median 16 versus 58 %).Conclusions and Clinical ImportancePlasma NT‐proBNP concentrations within the reference range do not rule out a congenital cardiac anomaly. Murmurs longer than 80% of the systole are most likely abnormal, whereas murmurs shorter than that could be either innocent or pathologic.