BackgroundRecent studies have shown that near-infrared (NIR) fluorescence imaging using Indocyanine green (ICG) may improve the efficiency of sentinel lymph node biopsy (SLNB). This study aimed to assess the effectiveness of the combination of ICG and methylene blue (MB) in breast cancer patients undergoing SLNB.Patients and methodWe evaluated ICG plus MB (ICG + MB) identification effectiveness with MB alone using retrospective analysis. From 2016 to 2020, we collected data on 300 eligible breast cancer patients who got SLNB treatment in our institution by ICG + MB or MB alone. By comparing the distribution of clinicopathological characteristics, the detection rate of sentinel lymph nodes (SLNs) and metastatic SLNs, as well as the total number of SLNs in the two groups, we were able to assess the imaging efficiency.ResultsFluorescence imaging allowed 131 out of 136 patients in the ICG + MB group to find SLNs. ICG + MB group and MB group had detection rates of 98.5% and 91.5% (P= 0.007, χ2= 7.352), respectively. Besides, the ICG + MB approach was able to produce improved recognition outcomes. What’s more, compared with the MB group, the ICG + MB group can identify more lymph nodes (LNs) (3.1 to 2.6,P= 0.000, t = 4.447). Additionally, in the ICG + MB group, ICG could identify more LNs than MB (3.1 vs 2.6,P= 0.004, t = 2.884).ConclusionICG has high detection effectiveness for SLNs, and when paired with MB, the detection efficiency can be increased even further. Furthermore, the ICG + MB tracing mode does not involve radioisotopes, which has a lot of promise for clinical use and can take the place of conventional standard detection methods.