Abstract: Objective To study the biodistribution of ¹²⁵I-Flt4 PcAb in rat and to establish the foundation of sentinel lymph node (SLN) exceptional orientation. Methods Flt4 PcAb was labeled with ¹²⁵I via chloramine T method. ¹²⁵I-Flt4 PcAb was subcutaneous injected in dorsum of foot in rats. The rats were killed at different stages after injection and the popliteal lymph nodes and major organs were taken out, then the radioactivity count of those organs and lymph nodes were measured. Results The labeling efficiency was 46.5%. The specific activity and the radiochemical purity of ¹²⁵I-Flt4 PcAb were 1.72×10¹²Bq/g and 96.7% respectively. It showed higher distribution in popliteal lymph nodes and blood than the other organs. The radioactivity ratio of popliteal lymph node to blood was more than 2.5 in 16 hours after injection. Conclusions:(1)It was successful to label Flt4 PcAb with ¹²⁵I via chloramine T method.(2)Flt4 PcAb would concentrate in lymph node after subcutaneous injection. (3)This study provided experimental basis for SLN exceptional orientation with Flt4 PcAb.