Purpose <p>Carbonic anhydrase IX (CAIX) represents a promising target for molecular imaging of clear cell renal cell carcinoma (ccRCC). We designed three radiotracers targeting CAIX to improve the diagnostic value of imaging by increasing tumor uptake and the liver excretion pathway. We also aimed to demonstrate the feasibility of applying small-molecule radiotracers in VEGFR-TKI efficacy monitoring in renal cancer.</p> Methods <p>This study developed three novel CAIX-targeting radiotracers, which were evaluated in vitro and in vivo; The study also included 8 patients with ccRCC, all of whom had paired [⁶⁸Ga]Ga-NOTA-FZCI-3 PET/CT and [¹⁸F]-FDG PET/CT. The diagnostic efficacies of these two PET tracers were compared. Moreover, VEGFR-TKI efficacy monitoring experiments using [⁶⁸Ga]Ga-NOTA-FZCI-3 was conducted on OS-RC-2 models.</p> Results <p>Three radiotracers showed that different liver/renal accumulations were better correlated with factors including distinct plasma protein binding abilities. [⁶⁸Ga]Ga-NOTA-FZCI-3, primarily cleared through the hepatobiliary system in patients, demonstrated a higher SUV_max of renal lesions (median, 13.10 vs. 3.10, <i>n</i> = 4; *<i>P</i> = 0.04) and superior tumor-to-muscle ratio of metastatic lesions (9.25 ± 5.10 vs.3.64 ± 2.11, <i>n</i> = 16; **<i>P</i> = 0.004) to [¹⁸F]-FDG in a prospective cohort with 8 ccRCC patients. In VEGFR-TKI efficacy monitoring experiments, [⁶⁸Ga]Ga-NOTA-FZCI-3 showed markedly decreased tumor radiotracer uptake throughout treatment. Moreover, VEGFR-AKT-mTOR pathway inhibition and CAIX downregulation in the treatment group confirmed that [⁶⁸Ga]Ga-NOTA-FZCI-3 PET/CT is a viable strategy for monitoring TKI efficacy in renal cell cancer.</p> Conclusion <p>[⁶⁸Ga]Ga-NOTA-FZCI-3 is a promising radiotracer for improving diagnosis, staging, and VEGFR-TKI efficacy monitoring in ccRCC.</p>