Optimization of transferrin receptor engagement is critical for achieving efficient receptor-mediated transport across the blood-brain barrier. We investigated anti-TfR1 VHH shuttle variants spanning a range of binding affinities in bispecific antibody formats and evaluated their transport characteristics in a human BBB model. Comparative analysis revealed affinity-dependent differences in transcytosis performance, providing mechanistic insight into BBB shuttle design and supporting the development of next-generation CNS delivery strategies for biologics and bioconjugates.
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