Background and aim The diagnosis and therapeutic management of large single pancreatic cystic lesions (PCLs) represent major issues for clinicians and essentially rely on endoscopic ultrasound fine-needle aspiration (EUS-FNA) findings. Needlebased confocal laser endomicroscopy (nCLE) has high diagnostic performance for PCLs. This study aimed to evaluate the impact of nCLE on the therapeutic management of patients with single PCLs. Methods Retrospective and comparative study. Five independent pancreatic disease experts from tertiary hospitals independently reviewed data from a prospective database of 206 patients with single PCL, larger than 2 cm and who underwent EUS-FNA and nCLE. Two evaluations were performed. The first one included the sequential review of clinical information, EUS report and FNA results. The second one included the same data + nCLE report. Participants had to propose a therapeutic management for each case. Results The addition of nCLE to EUS-FNA led to significant changes in therapeutic management for 28% of the patients (p < 0.001). nCLE significantly increased the interobserver agreement of 0.28 (p < 0.0001), from 0.36 (CI 95% 0.33-0.49) to 0.64 (CI 95% 0.61-0.67). nCLE improved the rates of full agreement among the five experts of 24% (p < 0.0001), from 30 to 54%. With nCLE, the surveillance rate of benign SCAs fell by 35%, from 40 (28/70) to 5% (4/76). Conclusion The addition of nCLE to EUS-FNA significantly improves reliability of PCL diagnosis and could impact the therapeutic management of patients with single PCLs. ClinicalTrials.gov number, NCT01563133.Keywords EUS-FNA · Pancreatic cysts · Needle-based confocal laser endomicroscopy Abbreviations BD-IPMN Branch duct-intraductal papillary mucinous neoplasm CT Computed tomography CEA Carcinoembryonic antigen EUS-FNA Endoscopic ultrasound fine-needle aspiration IML Indeterminate mucinous lesion MCN Mucinous cystic neoplasm MRI Magnetic resonance imaging nCLE Needle-based confocal laser endomicroscopy NEN Neuroendocrine neoplasm PC Pseudocyst PCLs Pancreatic cystic lesions SCA Serous cystadenoma The therapeutic management of pancreatic cystic lesions (PCL) depends on the cyst type. Surveillance or surgical and Other Interventional TechniquesPublisher's Note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.