Objective: The purpose of the study was to investigate the oncological sufficiency of level I axillary dissection for adequate histological nodal staging (ypN) in patients with breast cancer and tumor-involved sentinel lymph node (SLN) after neoadjuvant chemotherapy (NAC). Material and Methods: A prospective multicentre pilot study took place from 01.01.2018 to 30.11.2020 in three mammary centres in the Czech Republic in patients with breast cancer after NAC (NCT03556397). Patients in the cohort with positive histological frozen section of SLN were indicated to separate axillary dissection of levels I and II. Results: Sixty-one patients with breast cancer after NAC were included in the study according to inclusion and exclusion criteria. Twelve patients with breast cancer and tumour involved SLN after NAC were further included in the analysis. Two (16.7%) patients had positive non-sentinel lymph nodes in level I only, one (8.3%) patient had positive lymph nodes in level II only, and seven (58.3%) patients had positive lymph nodes in both levels. Level I axillary dissection in a patient with tumour involved SLN after NAC would have resulted in understaging in five (41.7%) patients, mostly ypN1 instead of ypN2. Conclusion: According to our pilot result, level I axillary dissection is not sufficient in terms of adequate histological nodal staging in breast cancer patients after NAC, and level II axillary dissection should not be omitted.