Fluorescent probes provide essential tools for studying biological systems. For the prolonged imaging of cellular analytes, the fast clearance of small-molecular probes and products is a matter of concern in the quantitative analysis. The activatable probes that produce insoluble products inside cell can be used for the prolonged imaging, but those with ratiometric imaging capability are rare. We disclose the novel sensing platform that is capable of the prolonged imaging, in addition to ratiometric signaling for the reliable quantitative analysis. Specifically, 3-(pyridin-4-yl)-8-(pyrrolidin-1-yl)-2 H benzo[ g ]chromen-2-one and its pyridinium salt as a dye couple constitute the ratiometric sensing platform. As the former dye produces highly emissive insoluble nanoaggregates inside cells, a fluorescent probe in the latter form, enables prolonged imaging of the target analyte in cells as well as in tissue by two-photon microscopy.