Abstract Background Cryoballloon pulmonary vein isolation (CryoPVI) is well established for atrial fibrillation (AF) ablation. Nevertheless, it is still important to identify possible periprocedural factors influencing its efficacy. With difficult anatomical conditions it is currently unclear whether the number of cryoballoon applications needed to achieve complete vein isolation has an impact on the long-term effectiveness of CryoPVI. Methods We prospectively analyzed AF patients who underwent first-time CryoPVI in a single center cohort between 2018-2023. The group allocation depended on the number of cryo applications until complete isolation was achieved. CryoPVI was performed in a standardized procedure. Follow-up was performed after 3, 6, 12, 18, 24 and 36 months post CryoPVI. Efficacy endpoint was the occurrence of symptomatic atrial arrhythmia. Results A total of 1170 patients were included in the study. Of these, in 610 patients received only one cryoballoon application per pulmonary vein to achieve complete isolation. Gender distribution (56.7% male vs. 43.3% male p=0.09) and AF phenotype (38.5% persistent AF vs. 61.5% paroxysmal AF, p=0.23) did not differ significantly between the two groups, patients who received only cryoballoon application were older (70.1 vs. 68.5, p=0.006) and accordingly had higher CHADS-VASc (2.86 vs. 2.62, p=0.006). After 36 months no difference in the efficacy endpoint was observed between the two groups (47.6% vs. 43.2%, p=0.53). Complications included three major access complications (0.5% vs. 0.5%) and one stroke within the first 30 days after the procedure (0.2% vs. 0.2%) in each group. No pericardial tamponade occurred. Conclusion Multiple- freezes during CryoPVI have no influence on atrial arrhythmia recurrence.