Purified 1D8, B5, and B5×1D8 were labeled with the VivoTag 680XL Protein Labeling Kit (PerkinElmer, NEV11118), according to the manufacturer’s protocol. MC38hB7-H3 cells were subcutaneously implanted into the 6-week-old female C57BL/6 mice. When tumor volume reached about 150 mm3, mice were randomly allocated to different treatment groups (n = 3 to 4 per group) and intravenously injected with 680XL-labeled antibody solution in PBS. Twenty-four hours after intravenous injection, tumor tissue and spleen were prepared and imaged using a red channel of a Fluorescence In Vivo Imaging (FOBI) system (NeoScience), which were then analyzed using NEOimage software (NeoScience). The tumor/spleen ratio was calculated by dividing the mean values of the identified region of interests.

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