Quantitative analysis of endosomal escape and intracellular delivery via bioorth...
Quantitative analysis of endosomal escape and intracellular delivery via bioorthogonal luminescent reaction
Intracellular delivery of membrane-impermeable biomacromolecules is essential for a broad spectrum of life sciences, ranging from fundamental biological studies to applied biomedical and pharmaceutical sciences. Despite the great...
Intracellular delivery of membrane-impermeable biomacromolecules is essential for a broad spectrum of life sciences, ranging from fundamental biological studies to applied biomedical and pharmaceutical sciences. Despite the great efforts in developing new intracellular delivery nanocarriers in the recent 20 years, simple, high-throughput, and accurate intracellular delivery quantification in live cells is still technically challenging. In BioLure, I propose an unconventional approach to quantify intracellular delivery and endosomal escape by a bioorthogonal luminescent reaction in live cells. Instead of a bulky fluorophore, I will label the molecule of interest (MOI) to be delivered with a single amino acid tag, which causes minimal changes in MOI’s physicochemical properties and functions. The tag will generate luciferase substrate inside live cells upon successful translocation into the cytoplasm by bioorthogonal reactions with chemoselectivity, rapid kinetics, biocompatibility, and high efficiency. The quantification strategy will initially be applied to the intracellular delivery of proteins via physical membrane disruption and validated by complementary methods. It will then be expanded to nanocarrier-mediated endosomal escape with different MOIs, including therapeutic siRNAs. The successful endosomal escape quantification will allow further nanoparticle screening for siRNA delivery.
I envision that BioLure will lead to a paradigm shift in the intracellular delivery field, facilitating the transformation from qualitative routine fluorescence imaging to high-throughput real-time quantification. The high sensitivity and low background make it an appealing tool for biologists to study endosomal escape and for material scientists to develop potent next-generation non-viral intracellular nanocarriers. Eventually, it will facilitate the design and screening of endosomal escape carriers and future nanomedicine formulations.ver más
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