PurposeThis study aimed to compare the prognosis in terms of disease-free survival (DFS) in three populations of women with breast cancer (BC) treated with neoadjuvant systemic treatment (NAST) in which axillary lymph node dissection (ALND) was performed based on different total tumor load (TTL) thresholds in the sentinel nodes.MethodsThis was an observational, retrospective study carried out in three Spanish centers. Data from patients with infiltrating BC who underwent BC surgery after NAST and intraoperative sentinel lymph node biopsy (SLNB) performed by One Step Nucleic acid Amplification (OSNA) technique during 2017 and 2018 were analyzed. ALND was performed according to the protocol of each center, based on three different TTL cut-offs (TTL > 250, TTL > 5000, and TTL > 15,000 CK19-mRNA copies/μL for centers 1, 2, and 3, respectively).ResultsA total of 157 BC patients were included in the study. No significant differences in DFS were observed between centers (Hazard ratio [HR] center 2 vs 1: 0.77;p= 0.707; HR center 3 vs 1: 0.83;p= 0.799). Patients with ALND had a shorter DFS (HR 2.43;p= 0.136), albeit not statistically significant. Patients with a triple negative subtype had a worse prognosis than those with other molecular subtypes (HR 2.82;p= 0.056).ConclusionNo significant differences in DFS were observed between three centers with different surgical approaches to ALND based on different TTL cut-offs in patients with BC after NAST. These results suggest that restricting ALND to those patients with TTL ≥ 15,000 copies/μL is a reliable approximation, avoiding unnecessary morbidities caused by ALND.Supplementary InformationThe online version contains supplementary material available at 10.1007/s10549-023-06954-8.