Far-Red Turn-On Fluorogenic Probe for Versatile Detection of Inorganic Polyphosphates

Abstract

Inorganic polyphosphate (polyP) is a ubiquitous phosphate biopolymer involved in diverse cellular processes. Despite its significance, selective detection of polyP remains challenging because of its simple and highly charged structure. Here, we report a far-red fluorogenic turn-on chemosensor for selective polyP detection and imaging, SiX-DPA-Zn. It integrates a silicon-xanthene fluorophore with a Zn(II)-coordinated 2,2’-dipicolylamine recognition unit. Replacing the bridging oxygen in the xanthene scaffold with silicon shifts fluorescence into the far-red region and confers up to 100-fold selectivity for inorganic polyP over other phosphate-containing biomolecules, including nucleic acids and nucleoside di(tri)phosphates. SiX-DPA-Zn enables quantitative detection of polyP at micromolar concentrations in microplate assays and stains a broad range of polyP species, starting from tripolyphosphate, in polyacrylamide gels. In HEK293 cells expressing Escherichia coli polyphosphate kinase 1, the probe visualizes intracellular polyP and enables quantitative analysis of its distribution relative to nuclear proteins─fibrillarin and nucleolin. Stimulated emission depletion (STED) microscopy further resolves subdiffraction-sized polyP granules within nuclear polyP aggregates. Together, these results establish SiX-DPA-Zn as a versatile chemical tool for quantitative polyP detection across biochemical assays and advanced fluorescence imaging.