ArticleJournal of the American Chemical Society2025
Design of Antigen-Targeting Fluorogenic Probes Utilizing Intramolecular Addition Reaction of Protein-Dye Hybrids.
Article in Journal of the American Chemical Society, 2025. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Not yet cited in PubMed.
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Abstract
Fluorogenic probes for antigens are useful for various purposes, including medical diagnostics and imaging, but achieving a rapid, large fluorescence increase is difficult. Here, we report a new class of fluorogenic probes for antigens based on a conjugate of an antibody-mimetic DARPin bearing a site-specifically incorporated cysteine and silicon-pyronine (SiP), which reacts reversibly with thiols. By using a library-screening approach, we found that the fluorescence of SiP conjugated to a DARPin is quenched via π-deconjugating addition reaction of the cysteine installed in the DARPin to SiP. Upon antigen binding, the equilibrium of this reaction is shifted to dissociation, restoring π-conjugation in the SiP and resulting in a large increase in fluorescence. As proof of concept of this chemical design principle, we constructed fluorogenic probes targeting GFP and EpCAM, which showed 25- and 12-fold fluorescence increases upon binding, respectively. The latter probe enabled wash-free cancer cell imaging with a low background.
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