BALB/c mice were injected with 2×10⁵ 4T1-Luc mammary carcinoma cells into the fourth mammary fat pad. When the tumor size reached approximately 50 mm³, micelles incorporating DiR were administered locally. (A) Schematic of local administration. Peritumoral administration was performed by injecting 50 μL of solutions into the peritumoral region approximately 5 mm away from the tumor. IT administration was performed by injecting 10 μL of solutions directly into the center of the tumor. (B and C) Ex vivo fluorescence images of tumor and TDLNs at 96 hours after peritumoral or IT micelle dosing and their quantification (n = 5). (D and E) Either conventional IT dosing of DiR or peritumoral dosing of micelle incorporating DiR was performed in 4T1-Luc tumor-bearing mice. For conventional IT dosing, DiR was first dissolved in Tween 80 (40 mg/mL) and then ethanol (2.5 mg/mL) sequentially, which was diluted with PBS into a final concentration. IT administration was performed by injecting 10 μL of solutions directly into the center of the tumor. Ex vivo fluorescence images of tumor and TDLNs at 96 hours after micelle or conventional IT dosing shown in D and their quantification in E (n = 3). Scale bars: 5 mm. Unpaired 2-tailed Student’s t test. Data are representative of 2 independent experiments. Data presented as mean ± SEM. LN, lymph node; In, inguinal; Ax, axillary; CL, contralateral; %ID/g, percent of injected dose per gram tissue.