Click-Chemistry-Mediated Rapid Microbubble Capture for Acute Thrombus Ultrasound Molecular Imaging

Bioorthogonal coupling chemistry has been studied as a potentially advantageous approach for molecular imaging because it offers rapid, efficient, and strong binding, which might also benefit stability, production, and chemical conjugation. The inverse-electron-demand Diels-Alder reaction between a 1,2,4,5-tetrazine and trans-cyclooctene (TCO) is an example of a highly selective and rapid bioorthogonal coupling reaction that has been used successfully to prepare targeted molecular imaging probes. Here we report a fast, reliable, and highly sensitive approach, based on a two-step pretargeting bioorthogonal approach, to achieving activated-platelet-specific CD62p-targeted thrombus ultrasound molecular imaging. Tetrazine-modified microbubbles (tetra-MBs) could be uniquely and rapidly captured by subsequent click chemistry of thrombus tagged with a trans-cyclooctene-pretreated CD62p antibody. Moreover, such tetra-MBs showed great long-term stability under physiological conditions, thus offering the ability to monitor thrombus changes in real time. We demonstrated for the first time that a bioorthogonal targeting molecular ultrasound imaging strategy based on tetra-MBs could be a simple but powerful tool for rapid diagnosis of acute thrombosis.