Careful evaluation, including provocation tests, is needed to specify an indication for septal reduction therapy in patients with drug-refractory hypertrophic obstructive cardiomyopathy. This study aimed to evaluate the outcome of alcohol septal ablation (ASA) using an intravenous nitroglycerin test (IV-NTG). Of consecutive 156 patients, after excluding cases of severe valvular disease and repeat septal reduction therapy, we investigated the clinical characteristics of patients with labile obstruction (n = 32) and the outcomes after ASA using the IV-NTG test; comparisons were made with those exhibiting basal obstruction (a resting gradient of ≥30 mm Hg). The patients with labile obstruction had less left ventricular mass (141 ± 47 vs 182 ± 59 g, p = 0.003) and less brain natriuretic peptide values (414 ± 576 vs 744 ± 625 pg/ml, p <0.001) than those with basal obstruction. Immediately after ASA, the gradients improved from 15 ± 7 to 5 ± 5 mm Hg and the IV-NTG-provoked gradients improved from 74 ± 25 to 13 ± 9 mm Hg, respectively. At 1-year follow-up, the New York Heart Association functional class had improved from 2.7 ± 0.5 to 1.3 ± 0.5. There was no sudden cardiac death during the follow-up period (5.1 ± 3.0 years), and 8-year survival free from cardiovascular death was 94%. In conclusion, patients with labile obstruction had less-severe left ventricular hypertrophy but exhibited symptoms comparable to those with basal obstruction. The IV-NTG test is a useful method for rapidly confirming acute reduction of the latent gradient after the ASA procedure, and the outcome of ASA for labile obstruction was favorable.