We developed radiopharmaceuticals for positron emission tomography (PET tracers) such as [¹⁸F]THK-5351 for imaging tau tangles, which is one of key neuropathological hallmarks in Alzheimer‘s disease (AD). Clinical PET studies revealed that [¹⁸F]THK-5351 possessed off-target binding. From validation studies such as imaging-autopsy correlations, we identified monoamine oxidase-B (MAO-B) as a dominant off-target binding substrate of [¹⁸F]THK-5351. MAO-B was predominantly expressed in astrocytes and elevated in various neurological conditions. Therefore, MAO-B would be an attractive target for imaging reactive astrogliosis. Though compound optimization from [¹⁸F]THK-5351, recently we developed a selective and reversible MAO-B PET tracer, [¹⁸F]SMBT-1. These translational and reverse-translational research from the development to validations of PET tracers lead to the generation of a new biomarker of reactive astrogliosis.