Radioisotopes and blue dyes are used as dual tracers in the current gold standard procedure of sentinel lymph node (SLN) biopsy (SLNB) performed for breast cancer. However, the blue dye or the radioisotope as a single tracer is also being applied in some institutes. We aimed to explore the risk factors for the miss-detection of SLNs with the radioisotope and the blue dye and to describe the distribution of SLNs missed by each tracer. Patients undergoing SLNB with radioisotope and blue dye as dual mapping agents were enrolled between August 2010 and August 2018. Radioactivity count, blue dye staining status, and size and location of each SLN were prospectively documented. In total, 2382 SLNs from 1010 patients were included for statistical analyses. The sentinel node identification rate was 100% for dual tracers, 99.4% for radioisotope, and 89.1% for blue dye. SLN identification using the blue dye was more likely to fail in patients undergoing breast-conserving surgery (p < 0.001) and mastectomy with reconstruction (p = 0.005). Furthermore, miss-detection was significantly more frequent in smaller and uninvolved nodes. Among all SLNs, 8.2% were located in level II and one was in level III. Notably, single tracer of blue dye tended to fail in the detection of lymph nodes in higher levels (p < 0.001). This study explored the association between features and the incidence of the failure to detect SLNs using radioisotope and blue dye. The locations of the miss-detected SLNs are demonstrated to provide a reference for SLNBs conducted using blue dye or radioisotope as a single tracer. © 2020 Yang et al.