Background: It is established that ethanol infusion in the vein of Marshall (EIVOM) effectively creates a linear ablation lesion in the mitral isthmus (MI). However, data on the long-term success rates of MI ablation remains limited. Methods and Results: Our cohort consisted of 560 patients with non-paroxysmal atrial fibrillation (AF) who underwent an initial MI ablation. Ablations were performed by only radiofrequency (RF) in 384 patients (RF group) or by RF and EIVOM in 176 patients (EIVOM/RF group). Ethanol of 5 mL was used to perform EIVOM in advance of RF. Following EIVOM, RF pulses were delivered to the lateral MI line. Bidirectional MI block was fully achieved in 353 (first 318, re-do 35) patients of the RF group and 171 (first 128, re-do 43) patients of the EIVOM/RF group (p = 0.09 in the first, 0.10 in the re-do ablation cases). In cases with complete MI line block, recurrent AF or atrial tachycardia (AT) was observed in 130 (37%) patients of the RF group and in 64 (37%) patients of the EIVOM/RF group (log-rank p = 0.12 in the first, 0.30 in the re-do ablation cases). Of the total 194 patients, 112 with drug refractory AF or AT proceeded to the subsequent ablation process. Reconnection of MI block line was observed in 39 (49%) patients in the RF group and 25 (58%) patients in the EIVOM/RF group (p = 0.32). Conclusion: EIVOM effectively ensures MI line block; however, the reconnection rate was similar between the two groups.