Novel turn-on fluorescent biosensors for selective detection of cellular Fe³⁺ in lysosomes: Thiophene as a selectivity-tuning handle for Fe³⁺ sensors

Iron-selective turn-on sensors are indispensable tools for understanding iron-related cell death processes and human diseases. In this study, we report a novel class of fluorescent sensors derived from an indolizino[3,2-c]quinoline scaffold that exhibit high selectivity for Fe³⁺ over other biologically abundant cations in cells, including Fe²⁺, Al³⁺, Zn²⁺, and Mn²⁺. IQ18 works as a ratiometric sensor with a K_d value of 7.1 × 10 ⁻⁷ M and a detection limit of 5.2 nM in ethanol, whereas IQ44 displays fluorescence enhancement upon binding with Fe³⁺ in both ethanol and water. In aqueous solution, IQ44 exists as 150-nm nanoparticles. The suppressed fluorescent emission of IQ44 nanoparticles in water is switched on in response to Fe³⁺, working as a turn-on nanoparticle sensor. Structure-property relationship analysis with IQ derivatives revealed that the thiophene ring confers selectivity for Fe³⁺. By installing thiophene in IQ44 as a selectivity-tuning handle, fluorescence in the presence of Fe³⁺ resulting from restriction of intramolecular rotation (RIR)and increased torsion angle induced by iron demonstrated that IQ44 is specifically localized in lysosomes, where it recognizes cellular Fe³⁺ in live cells, as determined using confocal microscopy. In addition, the increased fluorescent puncta of IQ44 in the presence of Fe³⁺ colocalized well with the RFP-tagged LC3 proteins (pmRFP-LC3), enabling the detection of the autophagy process.