Abstract: CLDN18.2 exhibits specific high expression in various solid tumors, such as gastric and pancreatic cancers, and has emerged as a widely investigated anti-tumor drug target in recent years. In this study, a ¹⁷⁷Lu-labeled radioimmunotherapy agent targeting CLDN18.2, ¹⁷⁷Lu-DOTA-h1D6, was constructed, and its in vitro stability, in vivo biodistribution in tumor-bearing mice, Micro-SPECT/CT imaging performance, and anti-tumor efficacy were preliminarily evaluated. The fully humanized monoclonal antibody h1D6 with high affinity for CLDN18.2 was modified with the bifunctional chelator (p-SCN-Bn-DOTA) and labeled with the radionuclide ¹⁷⁷Lu. Gentisic acid was introduced into the reaction system as a radiostabilizer, and the ¹⁷⁷Lu-DOTA-h1D6 conjugate was obtained after purification. The results show that the radiochemical purity (RCP) of ¹⁷⁷Lu-DOTA-h1D6 is greater than 95%, and the partition coefficient (lg D) is −3.16±0.09, indicating good hydrophilicity. The in vitro stability results indicate that the RCP remains above 95% after 168 h of incubation at 37 ℃, demonstrating good in vitro stability. The biodistribution results in tumor-bearing mice show that ¹⁷⁷Lu-DOTA-h1D6 is rapidly and specifically taken up in BGC823^CLDN18.2 xenografts. The tumor uptake reaches (14.59±2.23)%ID/g at 4 h, peaks at (47.73±14.90)%ID/g at 72 h, and exhibits continuous specific retention within 240 h ((13.65±3.62)%ID/g). The clearance from blood is relatively fast, and the target-to-non-target ratio increases significantly over time. Micro-SPECT/CT imaging and in vivo pharmacodynamics results demonstrate distinct radioactive accumulation in the tumor lesions starting from 24 h and continuous visualization up to 240 h. A single intravenous injection (18.5 MBq) significantly inhibits tumor growth in tumor-bearing mice without obvious systemic toxicity, further confirming the in vivo targeting specificity and therapeutic efficacy of ¹⁷⁷Lu-DOTA-h1D6. The ¹⁷⁷Lu-DOTA-h1D6 prepared in this study exhibits good in vitro stability, high in vivo targeting specificity, and significant anti-tumor effects, showing potential application value in the radioimmunotherapy of CLDN18.2-positive tumors.