Triplet-Triplet Annihilation Upconversion Based Nanosensors for Fluorescence Detection of Potassium

Abstract

Typical ionophore-based nanosensors use Nile blue derived indicators called chromoionophores, which must contend with strong background absorption, autofluorescence, and scattering in biological samples that limit their usefulness. Here, we demonstrate potassium-selective nanosensors that utilize triplet-triplet annihilation upconversion to minimize potential optical interference in biological media and a pH-sensitive quencher molecule to modulate the upconversion intensity in response to changes in analyte concentration. A triplet-triplet annihilation dye pair (platinum­(II) octaethylporphyrin and 9,10-diphenylanthracene) was integrated into nanosensors containing an analyte binding ligand (ionophore), charge-balancing additive, and a pH indicator quencher. The nanosensor response to potassium was shown to be reversible and stable for 3 days. In addition, the nanosensors are selective against sodium, calcium, and magnesium (selectivity coefficients in log10 units of −2.2 for calcium, −2.0 for sodium, and −2.4 for magnesium), three interfering ions found in biological samples. The lack of signal overlap between the upconversion nanosensors and GFP, a common biological fluorescent indicator, is demonstrated in confocal microscope images of sensors embedded in a bacterial biofilm.

Stimuli‐Responsive Molecular Photon Upconversion

“…Cash and co‐workers combined ionophore‐based optical sensors (IBOS) with TTA‐UC for anti‐Stokes potassium detection . IBOS are typically composed of a pH indicator, an analyte‐binding ligand (ionophore), and a charge‐balancing additive suspended in a highly plasticized hydrophobic polymer matrix which is surrounded by a lipid layer.…”

Stimuli‐Responsive Molecular Photon Upconversion

“…It is straightforward to convert conventional fluorescence sensing to anti‐Stokes sensing through triplet sensitization. As mentioned before, most fluorescent dyes compete with strong background absorption, autofluorescence, and scattering in biological samples, which can be solved with upconverted emission . Han, Ye et al.…”

“…Adapted with permission from ref. and . Copyright 2019 The Royal Society of Chemistry and 2020 American Chemical Society …”

“…To expand the scope of TTA‐UC to sensing applications, it is pivotal to develop a toolbox of mechanisms to make TTA‐UC responsive to a variety of external stimuli. TTA‐UC emission has been successfully regulated by external stimuli including temperature, oxygen, light, mechanical force, electric field, and chemicals . However, there has been no systematic understanding of the underlying mechanisms to switch TTA‐UC.…”

A dual-fluorophore sensor approach for ratiometric fluorescence imaging of potassium in living cells

“…24,25 Notably, recent years have also seen advances in the development of genetically encoded potassium probes and nanosensors. [26][27][28][29][30][31] In this context, small-molecule uorescent sensors offer an attractive approach to meet this goal, as they are valued as non-invasive, high signal-to-noise reagents that can accurately map a variety of biological analytes, particularly metal ions. [32][33][34][35][36][37] Despite notable advances in the development of smallmolecule uorescent sensors for potassium, [38][39][40][41][42][43][44][45] imaging intracellular K + in biological samples with high selectivity over Na + remains a signicant and insufficiently solved challenge, owing in large part to high resting K + concentrations within the cell (150 mM), which requires a low-affinity molecular recognition element that still achieves high K + selectivity.…”