Facile synthesis of multifunctional nanoparticles encoded with quantum dots and magnetic nanoparticles: cell tagging and MRI.

In this study, fluorescence-encoded magnetic biocompatible nanoparticles (NPs) were constructed from CdSe@ZnS quantum dots (QDs) and Fe3O4 nanoparticles with a one-step reprecipitation-encapsulation method. The resultant hybrid NPs exhibit small size (∼130 nm in diameter), highly bright QDs, two-color emissions (green and red) under single-wavelength excitation, easy separation with a magnet and efficient cellular internalization. Energy transfer between the incorporated QDs was studied to better tailor the encoded fluorescence, and 11 barcodes were obtained by adjusting the ratio of green and red QDs. We used four sets of the barcodes to tag specific cancer cells (HepG2) as a proof-of-concept, and distinguished each set according to respective overlayed fluorescence images using laser confocal microscopy. Moreover, the incorporated Fe3O4 NPs endowed as-constructed optical barcode superparamagnetic property by T 2-enhanced magnetic resonance effect with an r 2 value of 145.25 s-1 mM-1 at 3 T. These results suggest that the multifunctional NPs are very promising for discriminating different cells and dual-modality imaging.

Luminescent ruthenium(II)-containing metallopolymers with different ligands: synthesis and application as oxygen nanosensor for hypoxia imaging.

A series of Ru(II)-containing metallopolymers with different polypyridyl complexes, namely [Ru(N^N)2(L)](PF6)2 (L = bipyridine-branched polymer; N^N = bpy: 2,2'-bipyridine (Ru 1); phen: 1,10-phenanthroline (Ru 2); dpp: 4,7-diphenyl-1,10-phenanthroline (Ru 3)), were synthesized with the motive that adjusting π-conjugation length of ligands might produce competent luminescent oxygen probes. The three hydrophobic metallopolymers were studied with 1H NMR, UV-Vis absorption, and emission spectroscopy, and then were utilized to prepare biocompatible nanoparticles (NPs) via a nanoprecipitation method. Luminescent properties of the NPs were investigated against dissolved oxygen by steady-state and time-resolved spectroscopy respectively. Luminescence quenching of the three NPs all followed a linear behavior in the range of 0-43 ppm (oxygen concentration), but Ru 3-NPs exhibited the highest oxygen sensitivity (82%) and longest emission wavelength (λex = 460 nm; λem = 617 nm). In addition, external interferons from cellular environments (e.g., pH, temperature, and proteins) had been studied on Ru 3-NPs. Finally, dissolved oxygen in monolayer cells under normoxic/hypoxic conditions was clearly differentiated by using Ru 3-NPs as the luminescent sensor, and, more importantly, hypoxia within multicellular tumor spheroids was vividly imaged. These results suggest that such Ru(II)-containing metallopolymers are strong candidates for luminescent nanosensors towards hypoxia. Graphical abstract.

Facile synthesis of fluorinated nanophotosensitizers with self-supplied oxygen for efficient photodynamic therapy.

Tumor hypoxia severely reduces the efficiency of photodynamic therapy (PDT) through the insufficient supply of oxygen. In this work, we reported on a design of fluorinated nanophotosensitizers (NPSs) prepared by a facile reprecipitation-encapsulation method, with the aim of addressing the issue of hypoxia. The fluorinated NPSs consisted of a hybrid particle core of perfluorosiloxane-polystyrene, doped with a fluorinated photosensitizer, and a biocompatible poly-l-lysine shell. Compared with non-fluorinated counterpart NPSs that are similarly prepared except for the replacement of perfluorosiloxane with alkoxysilane, the fluorinated NPSs saturated with O2 exhibit approximately 3.5 fold higher singlet oxygen production yield and higher in vitro PDT efficiency due to the O2-carrying capability of intra-particle 'F-C' bonds.

Polylysine modified conjugated polymer nanoparticles loaded with the singlet oxygen probe 1,3-diphenylisobenzofuran and the photosensitizer indocyanine green for use in fluorometric sensing and in photodynamic therapy.

Conjugated polymer hybrid nanoparticles (NPs) loaded with both indocyanine green (ICG) and 1,3-diphenylisobenzofuran (DPBF) are described. The NPs are dually functional in that ICG acts as the photosensitizer, and DPBF as a probe for singlet oxygen (1O2 probe). The nanoparticle core consists of the energy donating host poly(9,9-dioctylfluorenyl-2,7-diyl)-co-(2,5-p-xylene) (PFP). The polymer is doped with the energy acceptor DPBF. Ratiometric fluorometric detection of singlet oxygen is accomplished by measurement of fluorescence at wavelengths of 415 and 458 nm. In addition, the shell of the positively charged polymeric nanoparticles was modified, via electrostatic interaction, with negatively charged PDT drugs ICG. The integrated nanoparticles of type ICG-DPBF-PFP display effective photodynamic performance under 808-nm laser irradiation. The 1O2 sensing behaviors of samples are evaluated based on the ratiometric fluorescent responses produced by DPBF and PFP. 1O2 can be fluorimetically sensed with a detection limit of 28 µM. The multifunctional nanoprobes exhibit effortless cellular uptake, superior photodynamic activity and a rapid ratiometric response to 1O2. Graphical abstractSchematic of a dual-functional nanoplatform for photodynamic therapy (PDT) and singlet oxygen (1O2) feedback. It offers a new strategy for self-monitoring photodynamic ablation. FRET: fluorescence resonance energy transfer. Indocyanine green is attached in the shell of nanoparticles, and 1,3-diphenylisobenzofuran is doped into the energy donating host conjugated polymer.

A fluorescent nanoprobe for real-time monitoring of intracellular singlet oxygen during photodynamic therapy.

Sensing of intracellular singlet oxygen (1O2) is required in order to optimize photodynamic therapy (PDT). An optical nanoprobe is reported here for the optical determination of intracellular 1O2. The probe consists of a porous particle core doped with the commercial 1O2 probe 1,3-diphenylisobenzofuran (DPBF) and a layer of poly-L-lysine. The nanoparticle probes have a particle size of ~80 nm in diameter, exhibit good biocompatibility, improved photostability and high sensitivity for 1O2 in both absorbance (peak at 420 nm) and fluorescence (with excitation/emission peaks at 405/458 nm). Nanoprobes doped with 20% of DPBF are best suited even though they suffer from concentration quenching of fluorescence. In comparison with the commercial fluorescent 1O2 probe SOSG, 20%-doped DPBF-NPs (aged) shows higher sensitivity for 1O2 generated at an early stage. The best nanoprobes were used to real-time monitor the PDT-triggered generation of 1O2 inside live cells, and the generation rate is found to depend on the supply of intracellular oxygen. Graphical abstract A fluorescent nanoprobe featured with refined selectivity and improved sensitivity towards 1O2 was prepared from the absorption-based probe DBPF and used to real-time monitoring of the generation of intracellular 1O2 produced during PDT.

Optically Encoded Semiconducting Polymer Dots with Single-Wavelength Excitation for Barcoding and Tracking of Single Cells.

Multiplexed optical encoding is emerging as a powerful technique for high-throughput cellular analysis and molecular assays. Most of the developed optical barcodes, however, either suffer from large particle size or are incompatible with most commercial optical instruments. Here, a new type of nanoscale fluorescent barcode (Pdot barcodes) was prepared from semiconducting polymers. The Pdot barcodes possess the merits of small size (∼20 nm in diameter), narrow emission bands (full-width-at-half-maximum (fwhm) of 30-40 nm), three-color emissions (blue, green, and red) under single-wavelength excitation, a high brightness, good pH and thermal stability, and efficient cellular uptake. The Pdot barcodes were prepared using a three-color and six-intensity encoding strategy; for ratiometric readout of the barcodes, one of the colors might be used as an internal reference. We used the Pdot barcodes to label 20 sets of cancer cells and then distinguished and identified each set based on the Pdot barcodes using flow cytometry. We also monitored and tracked single cells labeled with different Pdot barcodes, even through rounds of cell division. These results suggest Pdot barcodes are strong candidates for discriminating different labeled cell and for long-term cell tracking.

Preparation of Fluorescent Dye-Doped Biocompatible Nanoparticles for Cell Labeling.

In this paper, we report a series of fluorescent biocompatible nanoparticles (NPs), prepared by a facile reprecipitation-encapsulation method, for cellular labeling. The as-prepared NPs exhibit a narrow size distribution of 70-110 nm, and a core-shell structure comprised of a hybrid core doped with different dyes and a poly-L-lysine (PLL) shell. With coumarin 6, nile red, and meso- tetraphenylporphyrin as the imaging agents, the fluorescent NPs gave green, orange, and red emissions respectively. Due to the positively charged PLL shell, the fluorescent NPs exhibit neglected cytotoxicity and efficient cellular uptake. After incubation with living cells, the results obtained by laser confocal microscope from green, orange, and red channels all clearly show that the fluores- cent NPs are inhomogenously localized inside the cytoplasm without penetrating into the nucleus. Since such PLL-modified NPs can encapsulate other hydrophobic dyes, a wide spectrum of nanoimaging agents is thus expected. Furthermore, the surface amino groups on the PLL shell afford an anchoring site for further bioconjugation, and targeted imaging is also very promising.

Soft fluorescent nanomaterials for biological and biomedical imaging.

Soft fluorescent nanomaterials have attracted recent attention as imaging agents for biological applications, because they provide the advantages of good biocompatibility, high brightness, and easy biofunctionalization. Here, we provide a survey of recent developments in fluorescent soft nano-sized biological imaging agents. Various soft fluorescent nanoparticles (NPs) (including dye-doped polymer NPs, semiconducting polymer NPs, small-molecule organic NPs, nanogels, micelles, vesicles, and biomaterial-based NPs) are summarized from the perspectives of preparation methods, structure, optical properties, and surface functionalization. Based on both optical and functional properties of the nano-sized imaging agents, their applications are then reviewed in terms of in vitro imaging, in vivo imaging, and cellular-process imaging, by means of specific or nonspecific targeting.

Development of Microfluidic Systems Enabling High-Throughput Single-Cell Protein Characterization.

This article reviews recent developments in microfluidic systems enabling high-throughput characterization of single-cell proteins. Four key perspectives of microfluidic platforms are included in this review: (1) microfluidic fluorescent flow cytometry; (2) droplet based microfluidic flow cytometry; (3) large-array micro wells (microengraving); and (4) large-array micro chambers (barcode microchips). We examine the advantages and limitations of each technique and discuss future research opportunities by focusing on three key performance parameters (absolute quantification, sensitivity, and throughput).

One-Step Nanoengineering of Hydrophobic Photosensitive Drugs for the Photodynamic Therapy.

Nanoengineering of anticancer therapeutic drugs including photosensitizers is highly desired and extremely required for improved therapeutic efficacy. It remains a formidable challenge to achieve nanostructured colloidal particles directly starting from hydrophobic drugs due to their hydrophobic nature and ready aggregation in aqueous ambient. In this work, we report a facile method for a one-pot preparation of hydrophobic photosensitizer nanoparticles by coating with different types of polyelectrolyte as stabilizing agents. Regardless of negatively or positively charged polyelectrolyte used, including Poly-L-lysine (PLL, MW = 15 k-30 k), PLL (MW = 30 k-70 k), heparin, and hyaluronic acid (HA), the hydrophobic photosensitizer BDEA (2,5-Bis(4-(diethylamino)benzylidene)cyclopentanone) as a model drug can be readily manipulated into stable and well-dispersed nanoparticles with size of average 120 nm. Stabilization presumably contributes to electrostatic repulsion of the adsorbed polyelectrolyte layer onto nanoparticles. Their anticancer activity against the HeLa cell line shows that the endocytic internalization of these nanosystems is associated with antiproliferative effects after irradiation with visible light. The one-step preparation strategy may be an alternative approach for the design of nano-formulations of hydrophobic photosensitive drugs, presenting a potential for photodynamic antitumor therapy.

Targetable phosphorescent oxygen nanosensors for the assessment of tumor mitochondrial dysfunction by monitoring the respiratory activity.

Cellular respiration is a worthwhile criterion to evaluate mitochondrial dysfunction by measuring the dissolved oxygen. However, most of the existing sensing strategies merely report extracellular (ec-) or intracellular (ic-) O2 rather than intramitochondrial (im-) O2 . Herein we present a method to assess tumor mitochondrial dysfunction with three phosphorescent nanosensors, which respond to ec-, ic-, and im-O2 . Time-resolved luminescence is applied to determine the respective oxygen consumption rates (OCRs) under varying respiratory conditions. Data obtained for the OCRs and on (intra)cellular O2 gradients demonstrate that mitochondria in tumor cells are distinctly less active than those of healthy cells, resulting from restrained glucose utilization of and physical injury to the mitochondria. We believe that such a site-resolved sensing strategy can be applied to numerous other situations, for example to evaluate the adverse effects of drug candidates.

Organic ultraviolet photodetector based on phosphorescent material.

A new type of organic visible-blind UV-PDs is demonstrated by utilizing phosphorescent material Ir(III)bis[(4,6-difluorophenyl)-pyridinato-N, C(2)'] picolinate (FIrpic) as the electron donor and [6,6]-phenyl-C-61-butyricacidmethylester (PCBM) as the electron accepter, respectively. The peak responsivity of the organic UV-PDs is as high as 140 mA/W, corresponding to an external quantum efficiency of about 48%, under 365 nm UV light with an intensity of 0.018 mW/cm(2). The distinct photoluminescence quenching of FIrpic is obtained by doping PCBM. The organic UV-PDs provide visible-blind performance with the strong photocurrent response in the UV-A region, the rise and fall times of less than one second, and linear response within the incident light-intensity range from 0.018 to 20 mW/cm(2).

Facile one-step synthesis and transformation of Cu(I)-doped zinc sulfide nanocrystals to Cu(1.94)S-ZnS heterostructured nanocrystals.

A facile one-pot heating process without any injection has been developed to synthesize different Cu-Zn-S-based nanocrystals. The composition of the products evolves from Cu(I)-doped ZnS (ZnS:Cu(I)) nanocrystals into heterostructured nanocrystals consisting of monoclinic Cu1.94S and wurtzite ZnS just by controlling the molar ratios of zinc acetylacetonate (Zn(acac)2) to copper acetylacetonate (Cu(acac)2) in the mixture of n-dodecanethiol (DDT) and 1-octadecene (ODE). Accompanying the composition transformation, the crystal phase of ZnS is changed from cubic zinc blende to hexagonal wurtzite. Depending on the synthetic parameters including the reaction time, temperature, and the feeding ratios of Zn/Cu precursors, the morphology of the as-obtained heterostructured nanocrystals can be controlled in the forms of taper-like, matchstick-like, tadpole-like, or rod-like. Interestingly, when the molar ratio of Cu(acac)2 to Zn(acac)2 is increased to 9:1, the crystal phase of the products is transformed from monoclinic Cu1.94S to the mixed phase composed of cubic Cu1.8S and tetragonal Cu1.81S as the reaction time is further prolonged. The crystal-phase transformation results in the morphological change from quasi-spherical to rice shape due to the incorporation of Zn ions into the Cu1.94S matrix. This method provides a simple but highly reproducible approach for synthesis of Cu(I)-doped nanocrystals and heterostructured nanocrystals, which are potentially useful in the fabrication of optoelectronic devices.

Controllable synthesis of silver and silver sulfide nanocrystals via selective cleavage of chemical bonds.

A one-step colloidal process has been adopted to prepare silver (Ag) and silver sulfide (Ag2S) nanocrystals, thus avoiding presynthesis of an organometallic precursor and the injection of a toxic phosphine agent. During the reaction, a layered intermediate compound is first formed, which then acts as a precursor, decomposing into the nanocrystals. The composition of the as-obtained products can be controlled by selective cleavage of S-C bonds or Ag-S bonds. Pure Ag2S nanocrystals can be obtained by directly heating silver acetate (Ag(OAc)) and n-dodecanethiol (DDT) at 200 ° C without any surfactant, and pure Ag nanocrystals can be synthesized successfully if the reaction temperature is reduced to 190 ° C and the amount of DDT is decreased to 1 ml in the presence of a non-coordinating organic solvent (1-octadecene, ODE). Otherwise, the mixture of Ag and Ag2S is obtained by directly heating Ag(OAc) in DDT by increasing the reaction temperature or in a mixture of DDT and ODE at 200 ° C. The formation mechanism has been discussed in detail in terms of selective S-C and Ag-S bond dissociation due to the nucleophilic attack of DDT and the lower bonding energy of Ag-S. Interestingly, some products can easily self-assemble into two- or three-dimensional (2D or 3D) highly ordered superlattice structures on a copper grid without any additional steps. The excess DDT plays a key role in the superlattice structure due to the bundling and interdigitation of the thiolate molecules adsorbed on the as-obtained nanocrystals.

Real-time drug release monitoring from pH-responsive CuS-encapsulated metal-organic frameworks.

Real-time monitoring of drug release behaviors over extended periods of time is critical in understanding the dynamics of drug progression for personalized chemotherapeutic treatment. In this work, we report a metal-organic framework (MOF)-based nanotheranostic system encapsulated with photothermal agents (CuS) and therapeutic drug (DOX) to achieve the capabilities of real-time drug release monitoring and combined chemo-photothermal therapy. Meanwhile, folic acid-conjugated polyethylene glycol (FA-PEG) antennas were connected to the MOF through coordination interactions, endowing the MOF with an enhanced active targeting effect toward cancer cells. It is anticipated that such a theranostic agent, simultaneously possessing tumor-targeting, real-time drug monitoring and effective treatment, will potentially enhance the performance in cancer therapy.

Fluorescein isothiocyanate-doped conjugated polymer nanoparticles for two-photon ratiometric fluorescent imaging of intracellular pH fluctuations.

Herein, we report a two-photon ratiometric fluorescent pH nanosensor based on conjugated polymer poly(9,9-di-n-octylfluorenyl-2,7-diyl) (PFO) nanoparticles loaded with pH-sensitive fluorescein isothiocyanate (FITC) for intracellular pH monitoring. The obtained nanosensor (FITC-PFO NPs) possesses high sensitivity, excellent stability, good reversibility, favorable two-photon excitability and low cytotoxicity. The ratiometric fluorescence of FITC and PFO (F517/F417) in FITC-PFO NPs solution shows an efficient pH-sensitive response over the pH range from 3 to 10 (pKa = 6.43) under two-photon excitation. Additionally, the FITC-PFO NPs is successfully applied for ratiometric imaging of intracellular pH and its fluctuation in both one-photon and two-photon excitation modes. Overall, the two-photon pH nanosensor based on FITC-PFO NPs exhibits great potential in crucial physiological and biological processes related to intracellular pH fluctuations.

Skin-safe nanophotosensitizers with highly-controlled synthesized polydopamine shell for synergetic chemo-photodynamic therapy.

Although photodynamic therapy (PDT) has been extensively studied as an established modality of cancer treatment, it still suffers from a few clinical limitations, such as skin phototoxicity and tumor hypoxia. To circumvent these hurdles, hollow silica mesoporous nanoparticles (HMSNs) loaded with photosensitizers were employed as the nanoplatform to construct multifunctional nanoparticles (NPs). Specifically, an ultra-uniform polydopamine (PDA) shell was highly controlled grown around HMSNs by photogenerated outwards-diffused 1O2, followed by conjugation of folic acid-poly(ethylene glycol) and chelation of Fe2+ ions. Thanks to the optimal thickness of light-absorbing PDA shell, the multifunctional NPs exhibited not only negligible skin phototoxicity but also efficient 1O2 generation and photothermal (PT)-enhanced •OH generation upon respective photoirradiation. Anti-tumor therapy was then performed on both 4 T1 tumor cells and tumor-bearing mice by the combination of 638 nm PDT and 808 nm PT-enhanced chemodynamic therapy (CDT). As a result, high therapeutic efficacy was achieved compared to single-modality therapy, with a cell inhibitory rate of 86% and tumor growth inhibition of 70.4% respectively. More interestingly, tumor metastasis was effectively inhibited by the synergetic treatment. These results convincingly demonstrate that our multifunctional NPs are very promising skin-safe PDT agents combined with CDT for efficient tumor therapy.

A Comprehensive Study of Drug Loading in Hollow Mesoporous Silica Nanoparticles: Impacting Factors and Loading Efficiency.

Although hollow mesoporous silica nanoparticles (HMSNs) have been intensively studied as nanocarriers, selecting the right HMSNs for specific drugs still remains challenging due to the enormous diversity in so far reported HMSNs and drugs. To this end, we herein made a comprehensive study on drug loading in HMSNs from the viewpoint of impacting factors and loading efficiency. Specifically, two types of HMSNs with negative and positive zeta potential were delicately constructed, and three categories of drugs were selected as delivery targets: highly hydrophobic and lipophobic (oily), hydrophobic, and hydrophilic. The results indicated that (i) oily drugs could be efficiently loaded into both of the two HMSNs, (ii) HMSNs were not good carriers for hydrophobic drugs, especially for planar drugs, (iii) HMSNs had high loading efficiency towards oppositely charged hydrophilic drugs, i.e., negatively charged HMSNs for cationic molecules and vice versa, (iv) entrapped drugs would alter zeta potential of drug-loaded HMSNs. This work may provide general guidelines about designing high-payload HMSNs by reference to the physicochemical property of drugs.

Facile synthesis of polypyrrole-rhodamine B nanoparticles for self-monitored photothermal therapy of cancer cells.

Photothermal therapy following microscopic temperature detection can avoid overheating effects or insufficient heating, and thus improve therapeutic efficacy. In this study, biocompatible dual-functional nanoparticles (NPs) are constructed from polypyrrole (PPy) and rhodamine B (RB) by a one-step modified polymerization method. The polypyrrole serves as a photothemal agent, and rhodamine B acts as a temperature-sensing probe. The polypyrrole-rhodamine B (PPy-RB) NPs possess a high photothermal effect on irradiation by 808 nm laser, and a competent temperature sensitivity for the real-time temperature monitoring based on the emission intensity response of rhodamine B. After acting on HepG2 cells, the PPy-RB NPs can effectively induce cancer cell death, and the microscopic temperature is monitored by fluorescence feedback from rhodamine B during PTT by laser confocal microscopy. Hence, the proposed approach can supply a facile and promising way for the fabrication of effective theranostic nanoplatforms assisted by self-monitoring of cancer therapeutic processes.

Plasmon-Enhanced Blue-Light Emission of Stable Perovskite Quantum Dot Membranes.

A series of stable and color-tunable MAPbBr3-xClx quantum dot membranes were fabricated via a cost-efficient high-throughput technology. MAPbBr3-xClx quantum dots grown in-situ in polyvinylidene fluoride electrospun nanofibers exhibit extraordinary stability. As polyvinylidene fluoride can prevent the molecular group MA+ from aggregating, MAPbBr3-xClx quantum dots are several nanometers and monodisperse in polyvinylidene fluoride fiber. As-prepared MAPbBr3-xClx quantum dot membranes exhibit the variable luminous color by controlling the Cl- content of MAPbBr3-xClx quantum dots. To improve blue-light emission efficiency, we successfully introduced Ag nanoparticle nanofibers into MAPbBr1.2Cl1.8 quantum dot membranes via layer-by-layer electrospinning and obtained ~4.8 folds fluorescence enhancement for one unit. Furthermore, the originality explanation for the fluorescence enhancement of MAPbBr3-xClx quantum dots is proposed based on simulating optical field distribution of the research system.