Objective To explore the clinical value of only using blue dye as tracer in the sentinel lymph node biopsy (SLNB) of breast cancer. Methods SLNB was performed with the guidance of the combination of blue dye and isotope in all patients enrolled. SLNB data of blue dye only, and the combination method was recorded respectively for analyses. Results Three hundred and eight patients were enrolled in this prospective study. Significant differences were found in the identification rate (IDR, 93.5% vs. 99.4%, P=0.000), the false negative rate (FNR, 14.8% vs. 3.3%, P=0.007), the accuracy rate (AR, 89.6% vs. 97.8%, P=0.006), and the negative predictive value (NPV, 74.0% vs. 93.3%, P=0.012) between the blue dye alone and the combination method. The IDR and the FNR of the two methods were not significantly associated with the patient age, tumor size and location, histopathological type, type of biopsy and breast surgery, or ER, PR, and HER-2 status (all Pgt;0.05). The FNR of blue dye alone was significantly associated with clinically suspicious turgescence of axillary lymph nodes (P=0.042) and decreased followed by the increased number of sentinel lymph nodes obtained (P=0.000). Conclusions Compared with the combination method, SLNB guided with blue dye alone had significantly poorer IDR, AR, and NPV, and higher FNR. It is recommended that the combination of dye and isotope should be adopted for the guidance of SLNB in clinical practice rather than the use of blue dye alone.