Purpose: Mosaicism is a feature of several inherited tumor syndromes. Only few cases of mosaicism in Multiple Endocrine Neoplasia type 1 (MEN1) have been described. Next generation sequencing (NGS) offers new possibilities for detecting mosaicism. Here we report the first study to systematically look for MEN1 mosaicism, using blood DNA, in MEN1-suspected patients but without MEN1 pathogenic variants (PV) in a heterozygous state. Methods: digital targeted NGS, including unique molecular identifiers (UMIs), was performed in routine practice and the analytic performance of this method was verified. Results: Among a cohort of 119 patients harboring from 2 to 5 MEN1 lesions, we identified 3 patients with MEN1 mosaic PVs. The allele frequencies ranged from 2.3 to 9.5%. The detection rate of MEN1 mosaicism in patients bearing at least 3 MEN1 lesions was 17% (3/18). No cases were detected in patients with 2 lesions. Conclusion: We report here 3 new cases with MEN1 mosaicism. This study examined the performance of UMI in the diagnosis of MEN1 mosaicism in routine practice and our results underline that the frequency of mosaicism is probably underestimated in patients with suspected MEN1.