Amino-terminal pro-B-type natriuretic peptide (NT-proBNP) is a convenient molecule to work with in clinical laboratories, with preanalytic and analytic advantages, such as excellent stability at different temperatures, flexibility in sample type, and strong harmony across all commercially available NT-proBNP assays (including recently released point-of-care methods). Another major advantage of NT-proBNP assays is that they show excellent analytic precision. Reference values for NT-proBNP testing are strongly affected by the population tested. Among nondiseased populations, lower values are expected, whereas in diseased populations, such as in patients with acute dyspnea, higher reference values are more useful. Also, the biologic variability of NT-proBNP should be taken into account to evaluate the significance of any change in its values. When analyzed in patients with stable heart failure, biologic variability was 25%-40%. This article reviews the laboratory aspects of NT-proBNP testing from the perspective of the clinical laboratorian.