Targeted Nanoparticles for Selective Marking of Neuromuscular Junctions and ex Vivo Monitoring of Endogenous Acetylcholine Hydrolysis.

The present work for the first time introduces nanosensors for luminescent monitoring of acetylcholinesterase (AChE)-catalyzed hydrolysis of endogenous acetylcholine (ACh) released in neuromuscular junctions of isolated muscles. The sensing function results from the quenching of Tb(III)-centered luminescence due to proton-induced degradation of luminescent Tb(III) complexes doped into silica nanoparticles (SNs, 23 nm), when acetic acid is produced from the enzymatic hydrolysis of ACh. The targeting of the silica nanoparticles by α-bungarotoxin was used for selective staining of the synaptic space in the isolated muscles by the nanosensors. The targeting procedure was optimized for the high sensing sensitivity. The measuring of the Tb(III)-centered luminescence intensity of the targeted SNs by fluorescent microscopy enables us to sense a release of endogenous ACh in neuromuscular junctions of the isolated muscles under their stimulation by a high-frequency train (20 Hz, for 3 min). The ability of the targeted SNs to sense an inhibiting effect of paraoxon on enzymatic activity of AChE in ex vivo conditions provides a way of mimicking external stimuli effects on enzymatic processes in the isolated muscles.

Silica nanoparticles with Tb(III)-centered luminescence decorated by Ag0 as efficient cellular contrast agent with anticancer effect.

The present work introduces composite luminescent nanoparticles (Ag0-Tb3+-SNs), where ultra-small nanosilver (4 ±â€¯2 nm) is deposited onto amino-modified silica nanoparticles (35±6 nm) doped by green luminescent Tb(III) complexes. Ag0-Tb3+-SNs are able to image cancer (Hep-2) cells in confocal microscopy measurements due to efficient cell internalization, which is confirmed by TEM images of the Hep-2 cells exposed by Ag0-Tb3+-SNs. Comparative analysis of the cytotoxicity of normal fibroblasts (DK-4) and cancer cells (Hep-2) incubated with various concentrations of Ag0-Tb3+-SNs revealed the concentration range where the toxic effect on the cancer cells is significant, while it is insignificant towards the nonmalignant fibroblasts cells. The obtained results reveal Ag0-Tb3+-SNs as good cellular contrast agent able to induce the cancer cells death, which makes them promising theranostic in cancer diagnostics and therapy.

Cellular imaging by green luminescence of Tb(III)-doped aminomodified silica nanoparticles.

The work introduces Tb(III)-centered luminescence of amino-modified silica nanoparticles doped with Tb(III) complexes for cellular imaging. For these purposes water-in-oil procedure was optimized for synthesis of 20 and 35nm luminescent nanoparticles with amino-groups embedded on the surface. The obtained results indicate an impact of the nanoparticle size in decoration, aggregation behavior and luminescent properties of the nanoparticles in protein-based buffer solutions. Formation of a protein-based corona on the nanoparticles surface was revealed through the effect of the nanoparticles on helical superstructure of BSA. This effect is evident from CD spectral data, while no any size impact on the adsorption of BSA onto aminomodified silica surface was observed. Cellular uptake of the nanoparticles studied by confocal and TEM microscopy methods indicates greater cellular uptake for the smaller nanoparticles. Cytotoxicity of the nanoparticles was found to agree well with their cellular uptake behavior, which in turn was found to be greater for the smaller nanoparticles.

Luminescent silica nanoparticles for sensing acetylcholinesterase-catalyzed hydrolysis of acetylcholine.

This work highlights the H-function of Tb(III)-doped silica nanoparticles in aqueous solutions of acetic acid as a route to sense acetylcholinesterase-catalyzed hydrolysis of acetylcholine (ACh). The H-function results from H(+)-induced quenching of Tb(III)-centered luminescence due to protonation of Tb(III) complexes located close to silica/water interface. The H-function can be turned on/switched off by the concentration of complexes within core or nanoparticle shell zones, by the silica surface decoration and adsorption of both organic and inorganic cations on silica surface. Results indicate the optimal synthetic procedure for making nanoparticles capable of sensing acetic acid produced by enzymatic hydrolysis of acetylcholine. The H-function of nanoparticles was determined at various concentrations of ACh and AChE. The measurements show experimental conditions for fitting the H-function to Michaelis-Menten kinetics. Results confirm that reliable fluorescent monitoring AChE-catalyzed hydrolysis of ACh is possible through the H-function properties of Tb(III)-doped silica nanoparticles.

Tb(III)-doped silica nanoparticles for sensing: effect of interfacial interactions on substrate-induced luminescent response.

The present work introduces the easy modification of the water-in-oil microemulsion procedure aimed at the doping of the Tb(III) complexes within core or shell zones of the silica nanoparticles (SNs), which are designated as "core-shell", "shell", and "core". The dye molecules, chelating ligands, and copper ions were applied as the quenchers of Tb(III)-centered luminescence through dynamic or/and static mechanisms. The binding of the quenchers at the silica/water interface results in the quenching of the Tb(III) complexes within SNs, which, in turn, is greatly dependent on the synthetic procedure. The luminescence of "core" SNs remains unchanged under the binding of the quenchers at the silica/water interface. The quenching through dynamic mechanism is more significant for "core-shell" and "shell" than for "core" SNs. Thus, both "core-shell" and "shell" SNs have enough percentage of the Tb(III) complexes located close to the interface for efficient quenching through the energy transfer. The quenching through the ion or ligand exchange is most efficient for "core-shell" SNs due to the greatest percentage of the Tb(III) complexes at the silica/water interface, which correlates with the used synthetic procedure. The highlighted regularities introduce the applicability of "core-shell" SNs used as silica beads for phosphatidylcholine bilayers in sensing their permeability toward the quenching ions.

The energy transfer based fluorescent approach to detect the formation of silica supported phosphatidylcholine and phosphatidylserine containing bilayers.

The work introduces the quenching of silica coated Tb(III) complexes by merocyanine 540 (MC540) and copper ions as a tool to reveal the adsorption of phosphatidylcholine (PC) and phosphatidylserine (PS) at various PS:PC ratio onto silica nanoparticles doped with Tb(III) complex. The binding of MC540 with PC-based bilayers and copper ions with PS-based ones are the basis of their use as organic and inorganic probes to sense PS:PC ratio in silica supported mixed bilayers. The enrichment of mixed bilayers by PS results in the displacement of MC540 anions, while it enhances the binding with copper ions. The displacement or binding of probe ions results in the diverse luminescence response of Tb(III)-centred luminescence. The reestablishment of the steady state and time resolved luminescence is observed, when MC540 anions are applied as probes. The use of copper ions as probes results in the opposite quenching effect. The developed route enables to discriminate the formation of the phospholipids bilayers onto silica surface from those in the bulk of solution under various concentration conditions.

The discrimination between phospholipids of diverse structure and phosphacoumarins of various hydrophobicity through fluorescent response of Tb-doped silica nanoparticles decorated by cationic surfactant.

The work represents colloids of silica nanoparticles displaying fluorescent response on biorelevant compounds exemplified by phosphacoumarins and phospholipids. The luminescent properties of the colloids arise from Tb(III) complexes doped into silica nanoparticles (SNs). The noncovalent decoration of SNs by dicationic surfactant with further interfacial binding of dye anions enables to develop colloids programmed to display a substrate induced fluorescent response. The latter results from the quenching of Tb(III) centered luminescence by dye anions through dynamic mechanism and subsequent displacement of quenching anions by the non-quenching substrates from the interface of SNs. Both negative charge and hydrophobicity of substrates are the key factors affecting the selectivity of the substrate induced fluorescent response. The peculiar effects of zwitter-ionic and anionic phospholipids on the fluorescent response have been revealed. The applicability of the fluorescent procedure in the sensing of impurities in commercial phosphatidylcholine is also introduced.

The interfacial interactions of Tb-doped silica nanoparticles with surfactants and phospholipids revealed through the fluorescent response.

The quenching effect of dyes (phenol red and bromothymol blue) on Tb(III)-centered luminescence enables to sense the aggregation of cationic and anionic surfactants near the silica surface of Tb-doped silica nanoparticles (SN) in aqueous solutions. The Tb-centered luminescence of non-decorated SNs is diminished by the inner filter effect of both dyes. The decoration of the silica surface by cationic surfactants induces the quenching through the energy transfer between silica coated Tb(III) complexes and dye anions inserted into surfactant aggregates. Thus the distribution of surfactants aggregates at the silica/water interface and in the bulk of solution greatly affects dynamic quenching efficiency. The displacement of dye anions from the interfacial surfactant adlayer by anionic surfactants and phospholipids is accompanied by the "off-on" switching of Tb(III)-centered luminescence.