Cardiac Autoantibodies and AF Recurrence Introduction Recent evidence has suggested that autoantibodies may play an important role in the development of atrial fibrillation (AF). The predictive value of preprocedural autoantibodies against beta-1 adrenergic receptor (anti-β1-R) and M2-muscarinic acetylcholine receptor (anti-M2-R) for AF recurrence following cryoballoon-based pulmonary vein isolation (PVI) is still unclear. We aimed to determine the predictive value of preprocedural anti-β1-R and anti-M2-R levels for AF recurrence. Methods Eighty patients (mean age 54.25 ± 7.70 years; 40% female) with paroxysmal AF and preserved left ventricular function who underwent cryoballoon-based PVI were included in the study. Preprocedural anti-M2-R and anti-β1-R levels were measured with ELISA. Results At 1-year follow-up after ablation, late AF recurrence was observed in 17 (21.25%) patients. In the Cox regression model, including number of antiarrhythmic drugs, early AF recurrence, anti-β1-R levels >159.88 ng/mL, anti-M2-R levels >277.65 ng/mL, AF duration, and left atrial volume index, only anti-β1-R levels >159.88 ng/mL (HR: 4.281, P = 0.039) and anti-M2-R levels >277.65 ng/mL (HR: 4.313, P = 0.030) were found to be independent predictors of late AF recurrence. Anti-β1-R level >159.88 ng/mL was shown to predict late AF recurrence with a sensitivity of 70.59% and specificity of 90.48%. A cut-off value of 277.65 ng/mL for anti-M2-R level predicted AF recurrence with a sensitivity of 70.59% and specificity of 95.24%. Conclusion Preprocedural serum anti-β1-R and anti-M2-R levels are independent predictors of late AF recurrence following cryoballoon-based PVI in paroxysmal AF patients. Detection of preprocedural anti-β1-R and anti-M2-R levels may serve as a novel method for determination of paroxysmal AF patients who may not benefit from cryoballoon-based PVI.