Pretargeted brain PET imaging reveals amyloid-β pathology using a TCO-modified antibody and a fluorine-18 labelled tetrazine

Antibody-based positron emission tomography (PET) imaging holds promise for visualising and quantifying disease-related proteins in the brain, yet clinical application has been limited by the poor blood-brain barrier (BBB) penetration of antibodies and the need for long-lived radionuclides. Pretargeted imaging offers a solution by separating the delivery of the antibody from the radiolabelled imaging agent, enabling the use of clinically relevant short-lived radionuclides such as fluorine-18 (¹⁸F). Here, we demonstrate pretargeted PET imaging in the brain using a bispecific antibody modified with trans-cyclooctene (TCO) that targets both amyloid-β (Aβ), the main imaging target, and the transferrin receptor for enhanced BBB transport. Following administration of the TCO-modified antibody, a radiolabelled tetrazine ([¹⁸F]HTzA) was injected and underwent in vivo conjugation via a bioorthogonal click reaction at the brain target site. In a Alzheimer’s disease (AD) mouse model, this approach yielded specific imaging of brain Aβ pathology, with significantly higher uptake in AD mice than in wild-type controls or mice receiving unmodified antibody. Our results establish a generalisable method for high-contrast in vivo imaging of brain protein targets using pretargeted PET, with the potential to expand molecular imaging to currently inaccessible targets in diseases affecting the brain.