Dialkylated Si-Fluoresceins for Fluorogenic Labeling of Bovine Serum Albumin In Vivo

Fluorogenic labeling of biomolecules/biostructures in vivo is essential in bioimaging. Especially for targets with complex structures, such as bovine serum albumin, reliable labeling depends on the distinct turn-on fluorescence observed immediately upon recognition. Fluorescein derivatives, characterized by a xanthene scaffold, represent a class of bright organic fluorophores. However, strategies for fluorogenic fluorescein remain rare, mainly because of its pH sensitivity and difficulty in shifting spirolactone to the closed form due to the ionic nature of hydroxyl groups. Herein, we designed a series of fluorogenic SiFs (SiFCn) for BSA with fine-tuned structures by dialkylation. Through the collaborative manipulation of intramolecular ring-opening/closing and monomer/aggregation equilibria and the sensitivity to charged environments of the designed probes, SiFC4 and SiFC6 can selectively sense BSA with strong fluorescence, offering a novel strategy for sensing proteins in general. Using the designed molecular probes, BSA can be feasibly analyzed in commercial FBS in situ and in vivo in HeLa cells.