<p>Adeno-associated virus (AAV) mediated gene therapy is advancing and needs a noninvasive imaging tool to evaluate its effective targeting, biodistribution and clearance for precise use in humans. In this study, two serotypes of AAVs, AAV9-CMV-fLuc, and a brain targeting variant, AAVBR1-CMV-fLuc, are directly radiolabeled with the positron emission tomography (PET) radioisotope, ⁸⁹Zr. A radiolabeling synthon, [⁸⁹Zr]Zr-DFO-Bn-NCS or [⁸⁹Zr]Zr-DBN, was employed for the direct radiolabeling of AAVs, which enables tracking of AAVs by PET imaging for up to 18 days post-injection. The ⁸⁹Zr radiolabeled AAVs were administered to BALB/c mice via tail vein and assessed for their biodistribution at various time points up to day 18 post-injection. Imaging of AAVs was followed by ex-vivo biodistribution at day 18, or luciferase imaging at 3rd week or &gt; 30 days post-injection. The two serotypes showed differences in their biodistribution and trafficking in mice as early as 10&#xa0;min post-injection. The brain targeting serotype, [⁸⁹Zr]Zr-AAVBR1-CMV-fLuc, showed significantly higher uptake in the brain as compared to [⁸⁹Zr]Zr-AAV9-CMV-fLuc. The luciferase expression-based infection profile correlated with both PET imaging and ex-vivo biodistribution data. The developed methodology provides a noninvasive approach to image the pharmacokinetics of AAVs in a longitudinal manner and renders a selection of specific AAV serotypes for tissue/organ specific targeting.</p>