We have directly compared atrial natriuretic peptide (ANP), B-type natriuretic peptide (BNP), and N-terminal pro-ANP (N-ANP) as markers of patients with left ventricular ejection fraction (LVEF) ≤35%, as measured by radionuclide ventriculography. Venous blood samples were obtained from an unselected group of 87 patients who had been referred for assessment of ventricular function. ANP, BNP, and N-ANP were measured by radioimmunoassay using commercial kits. Receiver-operating characteristic analysis was used for the objective assessment of the diagnostic performance of each assay. There was a weak negative correlation between LVEF and plasma levels of ANP-li (r = −0.50,), BNP-li (r =−0.57), and N-ANP-li (r = −0.49) (p <5.01 for each peptide). Areas under the receiver-operating characteristic curves for BNP (0.880) and N-ANP (0.832) were not significantly different from each other, but were both significantly greater than the value for ANP (0.761): BNP versus ANP, p <0.01; and N-ANP versus ANP, p <0.05. The optimal sensitivity and specificity of each assay for the detection of patients with LVEF ≤35% were: BNP >4 pmol/L—sensitivity 1.0, specificity 0.58; N-ANP >200 pmol/L—sensitivity 0.95, specificity 0.35; and ANP >10 pmol/L—sensitivity 0.90, specificity 0.30. Plasma concentrations of BNP and N-ANP provide sensitive indicators of moderate to severe LV dysfunction; both peptides are objectively superior to ANP for identifying patients with LVEF ≤35%. These simple tests could be used to screen patients with suspected ventricular dysfunction to reduce the demand for further cardiac investigations.