Failure of cannulation of the right adrenal vein is frequent during AVS for investigation of primary aldosteronism (PA). The aldosterone:cortisol ratio of either adrenal vein compared with the inferior vena cava (AV/IVC index) has been proposed to differentiate between unilateral and bilateral disease, and aid in lateralization of unilateral disease.Sixty-two patients with unilateral or bilateral PA identified by either successful bilateral (45 patients) or unilateral (17 patients) adrenal vein cannulation, and with biochemical remission following surgery were enrolled into the analysis. The diagnostic performances of the previously identified AV/IVC index cutoffs of ≥5.5 to predict ipsilateral disease and ≤0.5 to predict contralateral disease were validated using data from the entire cohort.Fifty-three patients had unilateral PA and 9 patients bilateral PA. The area under ROC curve (AUROC) of the AV/IVC cutoff ≤0.5 for identifying unilateral aldosterone secretion from the contralateral adrenal was 0.95 (95% CI; 0.88-0.99), whereas the AUROC of the AV/IVC cutoff ≥5.5 for identifying unilateral aldosterone secretion from ipsilateral adrenal was 0.96 (95% CI; 0.92-0.99). The AV/IVC index cut-off value of 0.5 had 93 % sensitivity and 91% specificity; and the AV/IVC index cut-off value of 5.5 had 21% sensitivity and 100% specificity. The optimal AV/IVC cutoffs to achieve 100% specificity for our cohort were >2.4 and <0.1 to predict ipsilateral and contralateral disease.Our data confirm that the AV/IVC index is a potential tool for subtype classification and lateralization in patients with PA in the setting of failed bilateral, but successful unilateral, adrenal vein cannulation during AVS.