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A physicochemical study of ophthalmological vital dyes: From dimerization equilibrium in buffer solution to their liposomal dispersions
摘要： Vital dyes are used both in intra-operative and diagnostic ophthalmology as liquid solutions wherein dyes self-associate reducing their effects. Hence, an efficient dyes-delivery system is required to carry the maximum fraction of dye administered to the target site. In this study primarily two aspects of indocyanine green, patent blue V and brilliant blue G have been investigated. First, band profiles of UV–Vis absorption and emission spectra, acquired as a function of dye concentration in buffer solutions(pH = 7.4), have been analyzed to calculate the dimerization constant. Results demonstrate that their self-assembly properties have to be correlated to the molecular polarization. Second, the vital dyes have been loaded into liposomes prepared with the thin-film hydration method followed by extrusion. Encapsulation efficiency has been calculated to be 30%. Scanning electron microscopy and dynamic light scattering measurements have revealed that dye inclusion reduces the liposome diameter. Activation energy of liposome diffusion has been extracted by Arrhenius plots. Zeta potential measurements as a temperature function have been exploited to evaluate a dimensionless surface charge and prove that ratio charge(dye-loaded liposome)-to-charge(blank liposome) is always greater than unity and decreases with temperature.
关键词： Liposomes,Fluorescence,Vital dyes,UV–Visible,ζ-potential
更新于2025-09-19 17:15:36
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Theranostic liposomes as a bimodal carrier for magnetic resonance imaging contrast agent and photosensitizer
摘要： The present study is focused on the development of liposomes bearing gadolinium chelate (GdLip) providing two functionalities for magnetic resonance imaging (MRI) and photodynamic therapy of cancer. A lipid derivative of gadolinium(III) diethylenetriamine pentaacetic acid salt (GdDTPA1) was inserted in the liposomal membrane and served as MRI contrast agent whereas a zinc phthalocyanine (ZnPc) was used as a model photosensitizer. In addition to conventional liposomes, pegylated lipids were used for the preparation of 'stealth' liposomes. The characterization of different GdLip formulations involved evaluation of the liposomes size by nanoparticle tracking analysis, thermal phase behavior by differential scanning calorimetry and ZnPc-mediated singlet oxygen production. Furthermore, relaxivity measurements were performed as well as cytotoxicity and photodynamic activity against cancerous and normal cell lines was studied. Size and thermal behavior were only slightly influenced by GdLip composition, however it distinctly affected singlet oxygen production of ZnPc-loaded GdLip. The quantum yields of singlet oxygen generation by zinc phthalocyanine incorporated in GdLip containing cationic or/and pegylated lipids were smaller than those obtained for non-pegylated carriers with L-α-phosphatidylglycerol. In general, all formulations of GdLip, irrespectively of composition, were characterized by relaxivities higher than those of commercially used contrast agents (e.g. Magnevist?). NMR study has shown that the incorporation of ZnPc into the formulations of GdLip increases the relaxation parameters r1 and r2, compared to the values for the non-loaded vesicles. GdDTPA1 did not influence the photodynamic activity of ZnPc against HeLa cells.
关键词： Liposomes,Phthalocyanines,Theranostic,Photodynamic therapy,Magnetic resonance imaging
更新于2025-09-19 17:15:36
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Synergistical Use of Electrostatic and Hydrophobic Interactions for the Synthesis of a New Class of Multifunctional Nanohybrids: Plasmonic Magneto-Liposomes
摘要： By carefully controlling the electrostatic interactions between cationic liposomes, which already incorporate magnetic nanoparticles in the bilayers, and anionic gold nanoparticles, a new class of versatile multifunctional nanohybrids (plasmonic magneto-liposomes) that could have a major impact in drug delivery and controlled release applications has been synthesized. The experimental results confirmed the successful synthesis of hydrophobic superparamagnetic iron oxide nanoparticles (SPIONs) and polyethylene glycol functionalized (PEGylated) gold nanoparticles (AuNPs). The SPIONs were incorporated in the liposomal lipidic bilayers, thus promoting the formation of cationic magnetoliposomes. Different concentrations of SPIONs were loaded in the membrane. The cationic magnetoliposomes were decorated with anionic PEGylated gold nanoparticles using electrostatic interactions. The successful incorporation of SPIONs together with the modifications they generate in the bilayer were analyzed using Raman spectroscopy. The plasmonic properties of the multifunctional nanohybrids were investigated using UV-Vis absorption and (surface-enhanced) Raman spectroscopy. Their hyperthermic properties were recorded at different frequencies and magnetic field intensities. After the synthesis, the nanosystems were extensively characterized in order to properly evaluate their potential use in drug delivery applications and controlled release as a result of the interaction with an external stimulus, such as an NIR laser or alternating magnetic field.
关键词： superparamagnetic nanoparticles,hyperthermia,multifunctional nanohybrids,gold nanoparticles,magneto-liposomes
更新于2025-09-16 10:30:52
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Augmented COlorimetric NANoplasmonic (CONAN) Method for Grading Purity and Determine Concentration of EV Microliter Volume Solutions
摘要： This protocol paper describes how to assign a purity grade and to subsequently titrate extracellular vesicle (EV) solutions of a few microliters in volume by microplate COlorimetric NANoplasmonic (CONAN) assay. The CONAN assay consists of a solution of gold nanoparticles (AuNPs) into which the EV preparation is added. The solution turns blue if the EV preparation is pure, whereas it stays red if soluble exogenous single and aggregated proteins (SAPs; often referred to as protein contaminants) are present. The color change is visible by the naked eye or can be quantified by UV-Vis spectroscopy, providing an index of purity (a unique peculiarity to date). The assay specifically targets SAPs, and not the EV-related proteins, with a detection limit <50 ng/μl (an order of magnitude higher resolution than that of the Bradford protein assay). For pure solutions, the assay also allows for determining the EV number, as the color shift is linearly dependent on the AuNP/EV molar ratio. Instead, it automatically reports if the solution bears SAP contaminants, thus avoiding counting artifacts. The CONAN assay proves to be robust and reliable and displays very interesting performances in terms of cost (inexpensive reagents, run by standard microplate readers), working volumes (1–2 μl of sample required), and time (full procedure takes <1 h). The assay is applicable to all classes of natural and artificial lipid microvesicles and nanovesicles.
关键词： extracellular vesicles,particle number,nanoplasmonics,titration,liposomes,synthetic vesicles,nanoparticles,purity
更新于2025-09-16 10:30:52
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Liposome-Templated Indocyanine Green J- Aggregates for <i>In Vivo</i> Near Infrared Imaging and Stable Photothermal Heating
摘要： Indocyanine green (ICG) is an FDA-approved near-infrared fluorescent dye that has been used in optical imaging and photothermal therapy. Its rapid in vivo clearance and photo-degradation have limited its application. ICG pharmacokinetics and biodistribution have been improved via liposomal encapsulation, while its photothermal stability has been enhanced by ICG J-aggregate (IJA) formation. In the present work, we report a simple approach to engineer a nano-sized, highly stable IJA liposomal formulation. Our results showed that lipid film hydration and extrusion method led to efficient IJA formation in rigid DSPC liposomes, as supported by molecular dynamics modeling. The engineered DSPC-IJA formulation was nano-sized, and with spectroscopic and photothermal properties comparable to free IJA. Promisingly, DSPC-IJA exhibited high fluorescence, which enabled its in vivo tracking, showing prolonged blood circulation and significantly higher tumor fluorescence signals, compared to free ICG and IJA. Furthermore, DSPC-IJA demonstrated high photo-stability in vivo after multiple cycles of 808 nm laser irradiation. Finally, doxorubicin was loaded into liposomal IJA to utilize the co-delivery capabilities of liposomes. In conclusion, with both liposomes and ICG being clinically approved, our novel liposomal IJA could offer a clinically relevant theranostic platform enabling multimodal imaging and combinatory chemo- and photothermal cancer therapy.
关键词： liposomes,J-aggregates,indocyanine green,theranostics,photothermal therapy
更新于2025-09-16 10:30:52
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Novel Triamcinolone Acetonide-Loaded Liposomal Topical Formulation Improves Contrast Sensitivity Outcome After Femtosecond Laser-Assisted Cataract Surgery
摘要： Purpose: To assess visual results, macular modi?cations, and the incidence of clinically signi?cant macular edema (CSME) in patients using a topical triamcinolone acetonide-loaded liposomal formulation (TA-LF) after femtosecond laser-assisted cataract surgery (FLACS). Methods: Fifty-six eyes after FLACS were selected. Twenty-eight eyes in the combined therapy group (P + N) were treated with prednisolone 1% and nepafenac 0.1% for 21 days postoperatively, whereas 28 eyes in the TA-LF group received a liposomal formulation containing 2 mg/mL of TA (0.2%) for the same period of time. Follow-up visits at 1 day, 6 weeks, and 12 weeks after surgery consisted of visual acuity, contrast sensitivity (CS), central foveal thickness (CFT), total macular volume (TMV) measurements, and the detection of CSME. Results: CS improved in the TA-LF group (basal value: 1.087 – 0.339 vs. 1.276 – 0.147 at week 12, P = 0.0346), whereas in the P + N group, CS was not different from the baseline (basal value: 1.130 – 0.331 vs. 1.274 – 0.133 at week 12, P = 0.1276). There were similar increases in postoperative CFT and TMV in both groups. CFT and TMV signi?cantly correlate with CS only in the TA-LF group. The r2 for CFT and CS was 0.1963 (P = 0.0206), whereas the r2 for TMV and CS was 0.3615 (P = 0.0007) at 12 weeks. No difference was observed in the incidence of CSME between the groups. Conclusion: TA-LF is associated with better CS outcomes compared to combined therapy after FLACS.
关键词： femtosecond laser-assisted cataract surgery,macular edema,liposomes,visual outcomes,contrast sensitivity
更新于2025-09-11 14:15:04
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Sulfolobus acidocaldarius Microvesicles Exhibit Unusually Tight Packing Properties as Revealed by Optical Spectroscopy
摘要： In this study, we used optical spectroscopy to characterize the physical properties of microvesicles released from the thermoacidophilic archaeon Sulfolobus acidocaldarius (Sa-MVs). The most abundant proteins in Sa-MVs are the S-layer proteins, which self-assemble on the vesicle surface forming an array of crystalline structures. Lipids in Sa-MVs are exclusively bipolar tetraethers. We found that when excited at 275 nm, intrinsic protein fluorescence of Sa-MVs at 23 °C has an emission maximum at 303 nm (or 296 nm measured at 75 °C), which is unusually low for protein samples containing multiple tryptophans and tyrosines. In the presence of 10–11 mM of the surfactant n-tetradecyl-β-d-maltoside (TDM), Sa-MVs were disintegrated, the emission maximum of intrinsic protein fluorescence was shifted to 312 nm, and the excitation maximum was changed from 288 nm to 280.5 nm, in conjunction with a significant decrease (>2 times) in excitation band sharpness. These data suggest that most of the fluorescent amino acid residues in native Sa-MVs are in a tightly packed protein matrix and that the S-layer proteins may form J-aggregates. The membranes in Sa-MVs, as well as those of unilamellar vesicles (LUVs) made of the polar lipid fraction E (PLFE) tetraether lipids isolated from S. acidocaldarius (LUVPLFE), LUVs reconstituted from the tetraether lipids extracted from Sa-MVs (LUVMV) and LUVs made of the diester lipids, were investigated using the probe 6-dodecanoyl-2-dimethylaminonaphthalene (Laurdan). The generalized polarization (GP) values of Laurdan in tightly packed Sa-MVs, LUVMV, and LUVPLFE were found to be much lower than those obtained from less tightly packed DPPC gel state, which echoes the previous finding that the GP values from tetraether lipid membranes cannot be directly compared with the GP values from diester lipid membranes, due to differences in probe disposition. Laurdan’s GP and red-edge excitation shift (REES) values in Sa-MVs and LUVMV decrease with increasing temperature monotonically with no sign for lipid phase transition. Laurdan’s REES values are high (9.3–18.9 nm) in the tetraether lipid membrane systems (i.e., Sa-MVs, LUVMV and LUVPLFE) and low (0.4–5.0 nm) in diester liposomes. The high REES and low GP values suggest that Laurdan in tetraether lipid membranes, especially in the membrane of Sa-MVs, is in a very motionally restricted environment, bound water molecules and the polar moieties in the tetraether lipid headgroups strongly interact with Laurdan’s excited state dipole moment, and “solvent” reorientation around Laurdan’s chromophore in tetraether lipid membranes occurs very slowly compared to Laurdan’s lifetime.
关键词： glycerol dialkyl glycerol tetraether (GDGT),liposomes,intrinsic protein fluorescence,microvesicles,red edge excitation shift (REES),glycerol dialkyl calditol tetraether (GDNT),generalized polarization (GP),thermoacidophilic archaea,membrane probe,Laurdan
更新于2025-09-11 14:15:04
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A Quantitative and Reliable Calibration Standard for Dual-Color Fluorescence Cross-Correlation Spectroscopy
摘要： Fluorescence Cross-Correlation Dual-color Spectroscopy (dcFCCS) allows binding analysis of biomolecules. Combining cross- and autocorrelation amplitudes yields binding degrees and concentrations of bound and unbound species. However, non-ideal detection volume overlap reduces the cross-correlation, causing overestimation of the Kd. The overlap quality factor that relates measured and true cross-correlation amplitudes has been difficult to determine, because neither a perfect 1 : 1 labeled sample nor perfectly overlapping volumes are readily accomplished. Here, we describe how a stochastically labeled sample can be used for quantitative calibration. Lipid vesicles doped with green and red fluorescent dyes yield highly reproducible relative cross-correlations and allow determination of the setup-dependent overlap quality factor. This reliable, affordable and quick-to-prepare calibration standard expedites any quantitative co-localization or binding analysis by dcFCCS.
关键词： binding analysis,fluorescence,membranes,spectroscopy,calibration,liposomes
更新于2025-09-10 09:29:36
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[IEEE 2018 40th Annual International Conference of the IEEE Engineering in Medicine and Biology Society (EMBC) - Honolulu, HI, USA (2018.7.18-2018.7.21)] 2018 40th Annual International Conference of the IEEE Engineering in Medicine and Biology Society (EMBC) - In vitro Measurement of Release Kinetics of Temperature Sensitive Liposomes with a Fluorescence Imaging System
摘要： Temperature sensitive liposomes (TSL) are a promising type of nanoparticles for localized drug delivery. TSL typically release the contained drug at mild hyperthermic temperatures (40-42 oC). Combined with localized hyperthermia, this allows for local drug delivery. In vitro characterization of TSL involves measurements of drug release at varying temperatures, but current methods are inadequate due to low temporal resolution of ~8 – 10 seconds. We present a novel method for measuring the drug release with sub-second temporal resolution. In the proposed system, the TSL entrapping the fluorescent drug (Doxorubicin) are pumped through a capillary tube. The tube is rapidly heated to a desired temperature via Peltier element. Since fluorescence increases as drug is released from TSL, drug release kinetics can be measured via fluorescent imaging. By fitting exponential models, we calculated the time constants of drug release at temperatures of 39.5, 40.5 and 41.5?C were about 6.09, 2.06 and 1.03 seconds, respectively. Our initial tests show that the developed system can measure TSL release at subsecond resolution, and thus allow adequate in vitro evaluation of TSL formulations.
关键词： drug delivery,Peltier element,Temperature sensitive liposomes,fluorescence imaging,capillary tube
更新于2025-09-09 09:28:46