Nanoparticle‐Based Fluoroionophore for Analysis of Potassium Ion Dynamics in 3D Tissue Models and In Vivo

Abstract

The imaging of real-time fluxes of K+ ions in live cell with high dynamic range (5-150 mM) is of paramount importance for neuroscience and physiology of the gastrointestinal tract, kidney and other tissues. In particular, the research on high-performance deep-red fluorescent nanoparticle-based biosensors is highly anticipated. We found that BODIPY-based FI3 K+-sensitive fluoroionophore encapsulated in cationic polymer RL100 nanoparticles displays unusually strong efficiency in staining of broad spectrum of cell models, such as primary neurons and intestinal organoids. Using comparison of brightness, photostability and fluorescence lifetime imaging microscopy (FLIM) we confirmed that FI3 nanoparticles display distinctively superior intracellular staining compared to the free dye. We evaluated FI3 nanoparticles in real-time live cell imaging and found that it is highly useful for monitoring intra- and extracellular K+ dynamics in cultured neurons. Proof-of-concept in vivo brain imaging confirmed applicability of the biosensor for visualization of epileptic seizures. Collectively, this data makes fluoroionophore FI3 a versatile cross-platform fluorescent biosensor, broadly compatible with diverse experimental models and that crown ether-based polymer nanoparticles can provide a new venue for design of efficient fluorescent probes.

Bright and Photostable TADF‐Emitting Zirconium(IV) Pyridinedipyrrolide Complexes: Efficient Dyes for Decay Time‐Based Temperature Sensing and Imaging

“…In contrast, quaternary ammonium groups harboring Zr( Mes IPDP t ‐BuPh ) 2 ‐PVA‐TMA nanoparticles promoted efficient endo‐ and lysosome‐like staining ( Figure A–C; Figure S12, Supporting Information), similar to other previously described nanosensors. [ 9,12,38,82 ] As with the sulforhodamine‐based temperature probe, [ 38 ] we saw no effect of the positively charged nanoparticles on the cell viability (Figure S13, Supporting Information). We also observed relatively long retention of the nanoparticles inside the cells, lasting longer than 48 h (Figure S14, Supporting Information).…”

Background-Free Fluorescence-Decay-Time Sensing and Imaging of pH with Highly Photostable Diazaoxotriangulenium Dyes

“…Although they showed no evident toxicity after internalization by human colon-cancer HCT116 cells (SI Figure S-32), the staining was very fast (1 h or less) and highly similar to that with the free dye, which may indicate leaching of the dye from the nanoparticles inside the cells, as a result of their small size (SI Figures S-33 and S-34). For comparison, the nanoparticles prepared from the same polymer typically require 3 h or more for cell internalization. , The assumption of leaching was also supported by the FLIM measurements in cells: no reliable pH response or loss of mitochondrial-like localization after dissipation of the mitochondrial pH gradient were observed (SI Figures S-35 and S-36). Therefore, for intracellular-pH FLIM sensing, it appears to be essential to completely eliminate “in-cell” leaching of the dye out of the nanosensors, which can be possible by using longer (or more branched) anchor groups or via covalent grafting of the indicators into the polymer.…”

“…For comparison, the nanoparticles prepared from the same polymer typically require 3 h or more for cell internalization. 58,59 The assumption of leaching was also supported by the FLIM measurements in cells: no reliable pH response or loss of mitochondrial-like localization after dissipation of the mitochondrial pH gradient were observed (SI Figures S-35 and S-36). Therefore, for intracellular-pH FLIM sensing, it appears to be essential to completely eliminate "in-cell" leaching of the dye out of the nanosensors, which can be possible by using longer (or more branched) anchor groups or via covalent grafting of the indicators into the polymer.…”

Lack of biochemical signalling in GelMA leads to polarity reversion in intestinal organoids independent from mechanoreciprocity

“…For live imaging, organoids embedded in 20 μL Matrigel or GelMA hydrogel domes, were grown on microscopy dishes (Ibidi) for 3 days (37°C, 5% CO 2 ). To stain organoids with K + -sensitive fluor-ionophore nanoparticles, FI4 nanoparticles 39 (10 μg/mL) were added to the IntestiCult media for overnight incubation prior to the imaging. To visualise lysosomes, LysoSensor Green (4 μM, Thermo Fisher, Cat.…”