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Enormous success has been achieved in antitumor therapy with PD-1/PD-L1 inhibitors; however, less than 3 out of 10 patients show clinical benefits. An interesting phenomenon of patients who experienced durable clinical responses to PD-L1 blockade usually being the patients with high PD-L1 expression was noticed. Accordingly, the evaluation of PD-L1 is of great benefit to predict the immunotherapy response against PD-1/PD-L1. A small-molecule single photon emission computed tomography (SPECT) tracer [99mTc]Tc-SG2C-CBM containing a biphenyl moiety was designed and synthesized for the measurement of PD-L1 expression. The radiolabeling yield (RLY) and radiochemical purity (RCP) of [99mTc]Tc-SG2C-CBM were detected to be over 93% under the optimized labeling conditions. [99mTc]Tc-SG2C-CBM was discovered to be equipped with not only good stability but also suitable lipophilicity. In the cellular assays, [99mTc]Tc-SG2C-CBM showed moderate affinity to PD-L1 and 6.87-fold higher uptake in A375-hPD-L1 cells than in A375 cells. Then, the in vivo biodistribution results of the A375-hPD-L1 tumor-bearing mice showed that the radioactivity uptake in tumors was about 2.96 ± 0.76 and 2.31 ± 0.26 fold that in muscles and bones at 2 h post-injection, respectively. SPECT imaging was consistent with the biological distribution of [99mTc]Tc SG2C-CBM, with the uptake being mainly in the liver and less pronounced in tumors. The results of these studies indicate that [99mTc]Tc-SG2C-CBM may be a probe for imaging PD-L1 expression, but further research is needed. [ABSTRACT FROM AUTHOR]

Aggregation-induced emission (AIE)-active micelles are a type of fluorescent functional materials that exhibit enhanced emissions in the aggregated surfactant state. They have received significant interest due to their excellent fluorescence efficiency in the aggregated state, remarkable processability, and solubility. AIE-active micelles can be designed through the self-assembly of amphipathic AIE luminogens (AIEgens) and the encapsulation of non-emissive amphipathic molecules in AIEgens. Currently, a wide range of AIE-active micelles have been constructed, with a significant increase in research interest in this area. A series of advanced techniques has been used to characterize AIE-active micelles, such as cryogenic-electron microscopy (Cryo-EM) and confocal laser scanning microscopy (CLSM). This review provides an overview of the synthesis, characterization, and applications of AIE-active micelles, especially their applications in cell and in vivo imaging, biological and organic compound sensors, anticancer drugs, gene delivery, chemotherapy, photodynamic therapy, and photocatalytic reactions, with a focus on the most recent developments. Based on the synergistic effect of micelles and AIE, it is anticipated that this review will guide the development of innovative and fascinating AIE-active micelle materials with exciting architectures and functions in the future. [ABSTRACT FROM AUTHOR]

Fluorescent hypochlorite probes with ratiometric imaging ability are highly desirable for imaging hypochlorite in biological systems. However, it is still challenging to develop new scaffolds for these probes. In this study, we demonstrate that phenothiazine-fused boron complexes are promising scaffolds for the design of ratiometric fluorescent hypochlorite probes. The synthesized complexes based on the scaffold show ultrafast and ratiometric absorption/fluorescence changes for hypochlorite. We also developed an endoplasmic reticulum-targeting probe and demonstrated that it has excellent real-time imaging ability for both endogenous and exogenous hypochlorite in the endoplasmic reticulum of living cells. [ABSTRACT FROM AUTHOR]

SO2 plays an important role in our life and is also associated with many diseases, so it is a double-edged sword. Therefore, it is necessary to develop a probe for detecting sulfur dioxide derivatives. Herein, on account of the Michael addition reaction mechanism, a near-infrared (NIR) fluorescent probe, HBQI, toward sulfur dioxide derivatives was developed, and the probe has high selectivity and strong anti-interference ability. The application study indicates that probe HBQI can detect sulfur dioxide derivatives in real water samples with good recovery. In addition, HBQI has been applied to live-cell imaging through confocal fluorescence techniques. [ABSTRACT FROM AUTHOR]

We developed a ratiometric oxygen-sensitive nanosensor and demonstrated application in monitoring metabolic oxygen consumption in microbial samples over time. Based on a near-infrared (NIR) emitting oxygen-quenched luminophore, platinum(II) octaethylporphine ketone (PtOEPK), along with a stable dioctadecyl dicarbocyanine reference dye (DiD), this nanosensor system provides an advantageous approach for overcoming imaging issues in biological systems, such as autofluorescence and optical scattering in the visible wavelength region. The dyes are encapsulated within a polymer-based nanoparticle matrix to maintain them at a constant ratio in biological samples, precluding the need for complex synthetic approaches. With this constant ratio of the two dyes, the nanosensor response can be measured as a ratio of their two signals, accounting for nanosensor concentration artifacts in measurements. The nanosensors are reversible, which enabled us to temporally monitor systems in which dissolved oxygen concentrations both increase and decrease. These sensors were applied for the monitoring of oxygen in samples of Saccharomyces cerevisiae (brewing yeast) in a 96-well optical fluorescence plate reader format over 60 h. By mixing the nanosensors directly into the sample well with the yeast, we were able to dynamically track metabolic activity changes over time due to varying cell concentration and exposure to an antimicrobial agent. This system could be a potential platform for high-throughput screening of various species or variants of microbes with unknown metabolic rates in response to external stimuli (antimicrobials, metabolites, etc.). [ABSTRACT FROM AUTHOR]

Nonlinear optical microscopy that leverages an objective based total internal reflection (TIR) excitation scheme is an attractive means for rapid, wide-field imaging with enhanced surface sensitivity. Through select combinations of distinct modalities, one can, in principle, access complementary chemical and structural information for various chemical species near interfaces. Here, we report a successful implementation of such a wide-field nonlinear optical microscope system, which combines coherent anti-Stokes Raman scattering (CARS), two-photon fluorescence (TPF), second harmonic generation (SHG), and sum frequency generation (SFG) modalities on the same platform. The intense optical fields needed to drive these high order nonlinear optical processes are achieved through the use of femtosecond pulsed light in combination with the intrinsic field confinement induced by TIR over a large field of view. The performance of our multimodal microscope was first assessed through the experimental determination of its chemical fidelity, intensity and polarization dependences, and spatial resolution using a set of well-defined model systems. Subsequently, its unique capabilities were validated through imaging complex biological systems, including Hydrangea quercifolia pollen grains and Pantoea sp. YR343 bacterial cells. Specifically, the spatial distribution of different molecular groups in the former was visualized via vibrational contrast mechanisms of CARS, whereas co-registered TPF imaging allowed the identification of spatially localized intrinsic fluorophores. We further demonstrate the feasibility of our microscope for wide-field CARS imaging on live cells through independent characterization of cell viability using spatially co-registered TPF imaging. This approach to TIR enabled wide-field imaging is expected to provide new insights into bacterial strains and their interactions with other species in the rhizosphere in a time-resolved and chemically selective manner. [ABSTRACT FROM AUTHOR]

Formaldehyde (FA), as a reactive carbonyl species, is endogenously generated in various biological processes. Abnormal levels of FA could lead to various cellular dysfunction and pathological conditions. Here, we develop a new activatable fluorescent probe for highly selective visualization of FA in living cells. Our probe (Naph-1) is designed using a naphthalene derivative as the fluorophore and hydrazone as a recognition site for FA. Naph-1 is essentially nonemissive. After reacting with FA, the amine moiety is converted into a Schiff base with electron-withdrawing ability and the fluorescence is simultaneously turned on due to synergetic intramolecular charge transfer and favoured excited state intramolecular proton transfer effects. Naph-1 exhibits a large Stokes shift upon reaction with FA. Furthermore, it possesses high selectivity and superior sensitivity toward FA with an estimated limit of detection of 0.35 μM. Moreover, Naph-1 is also successfully applied to image both endogenous and exogenous formaldehyde in living cells. These features demonstrate that Naph-1 holds great potential in the detection and imaging of formaldehyde in biological systems. [ABSTRACT FROM AUTHOR]

Hypochlorous acid (HClO) is one of the most important reactive oxygen species (ROS) and plays a vital role in many physiological and pathological processes. The comprehensive exploration of mechanistic details and the potential clinical translation necessitate the development of reliable probes for prompt and accurate detection of HClO in complex biological environments. Herein we report a fluorinated bihydrazide conjugate as a 19F NMR/MRI probe with a "turn-on" character for the detection of HClO. This probe could selectively respond to HClO, leading to a significant recovery of 19F signals for 19F NMR/MRI. Activatable sensing and imaging of HClO were achieved with SMMC-7721 cells and nude mice, which demonstrates that this small molecular conjugate could serve as a selective probe for real-time sensing and imaging of HClO in biological systems. [ABSTRACT FROM AUTHOR]

We report a novel fluorescent probe HBN–TCF for the detection of SO2 derivatives. This probe exhibited near-infrared fluorescence emission with an excitation wavelength of 620 nm. After reacting with SO32−, the emission channel at 664 nm decreased, while the new strong emission channel at 482 nm increased (λex = 400 nm), with a large emission distance (Δλ = 182 nm) observed. This probe exhibited the rapid and selective detection of SO2 derivatives compared with other sulfur-containing species and featured a low detection limit (82 nM). This colorimetric and ratiometric fluorescent probe showed high selectivity and sensitivity for detecting SO2 derivatives. The probe was also successfully exploited for the fluorescence imaging of intracellular and exogenous SO2 derivatives in BEL-7402 cells. [ABSTRACT FROM AUTHOR]

A concept for a pure DNA hydrogel has been demonstrated in which triplex structures act as the core components and are assisted by single strands and artfully designed Y-shaped structures. Typically, the triplex structure which is based on protonated cytosine–guanine–cytosine (C–G·C+) is formed at pH 5.0 and disassembles at pH 7.0, whereas the thymine–adenine–thymine (T–A·T)-based triplex structure is formed at pH 7.0 and disassembles at pH 10.0. Therefore, such triplex DNA structure-based pure DNA hydrogels provide pH-controlled reversible self-assembly of DNA structures with a transition between gel and liquid states. More significantly, the introduction of both a fluorophore (FAM) and a quencher (BHQ1) to the hydrogels provides an innovative method for monitoring the self-assembly and disassembly processes through a fluorescence technology. [ABSTRACT FROM AUTHOR]

As a superb tool to visualize and study the spatial-temporal distribution of chemicals, Raman microscopy has made a big impact in many disciplines of science. While label-free imaging has been the prevailing strategy in Raman microscopy, recent development and applications of vibrational/Raman tags, particularly when coupled with stimulated Raman scattering (SRS) microscopy, have generated intense excitement in biomedical imaging. SRS imaging of vibrational tags has enabled researchers to study a wide range of small biomolecules with high specificity, sensitivity and multiplex capability, at a single live cell level, tissue level or even in vivo. As reviewed in this article, this platform has facilitated imaging distribution and dynamics of small molecules such as glucose, lipids, amino acids, nucleic acids, and drugs that are otherwise difficult to monitor with other means. As both the vibrational tags and Raman instrumental development progress rapidly and synergistically, we anticipate that this technique will shed light onto an even broader spectrum of biomedical problems. [ABSTRACT FROM AUTHOR]

A novel laser-ablation sample introduction system has been developed for the Fluidigm® Helios™ mass cytometer to enable highly multiplexed imaging of biological samples. Herein, we report on the analytical figures of merit for this system using a slide, coated uniformly with a metal-doped PMMA film, as a test sample. The system is based on the use of a fifth harmonic Nd–YAG laser focused to a spot size of 1 micrometer or less. A fast wash-out ablation chamber provides resolution of individual plumes when operated at a 100 Hz or 200 Hz laser repetition frequency. The system exhibits a linear response to metal concentration in the sample over 4 orders of magnitude. The variation of signals generated for 1 micrometer diameter pixels is ≤15% RSD. Pixel-to-pixel sample contamination due to redeposition of the ablated material is approximately 0.6%. The variation of the ablated crater diameter with laser energy is also explored, demonstrating a linear behaviour over a limited range of laser energies. [ABSTRACT FROM AUTHOR]

Visualization of dynamic functional and molecular events in an unperturbed in vivo environment is essential for understanding the complex biology of living organisms and of disease state and progression. To this end, optoacoustic (photoacoustic) sensing and imaging have demonstrated the exclusive capacity to maintain excellent optical contrast and high resolution in deep-tissue observations, far beyond the penetration limits of modern microscopy. Yet, the time domain is paramount for the observation and study of complex biological interactions that may be invisible in single snapshots of living systems. This review focuses on the recent advances in optoacoustic imaging assisted by smart molecular labeling and dynamic contrast enhancement approaches that enable new types of multiscale dynamic observations not attainable with other bio-imaging modalities. A wealth of investigated new research topics and clinical applications is further discussed, including imaging of large-scale brain activity patterns, volumetric visualization of moving organs and contrast agent kinetics, molecular imaging using targeted and genetically expressed labels, as well as three-dimensional handheld diagnostics of human subjects. [ABSTRACT FROM AUTHOR]

In the field of organic light-emitting diodes (OLEDs), research interests focus on making the optically dark triplet excitons shine in order to increase the electro-optic conversion efficiency of devices. In this work, two kinds of phenazine compounds, i.e. dibenzo[a,c]phenazine (DBP) and tribenzo[a,c,i]phenazine (TBP), were synthesized and used as model compounds to regulate the emission efficiency of the dark triplet excitons by chemical modification. Charge-transfer induced ultrafast intersystem crossing (CT-ISC) with a time constant of ∼1 ps was observed for these two phenazine derivatives upon photoexcitation with a high triplet yield of 77.1% for DBP and 58.7% for TBP. The triplet excited states of DBP can produce ultra-long phosphorescence with lifetime as long as 318 ms at 77 K. The quantum yield for phosphorescence (ΦP) is determined to be 8.45%. In sharp contrast, the triplet-excited 3TBP* undergoes an efficient reverse intersystem crossing (RISC) process, resulting in bright delayed fluorescence emission with negligible phosphorescence. A controllable luminescence behavior from the triplet states between fluorescence and phosphorescence in phenazine derivatives is demonstrated. Theoretical calculations reveal that the structure-dependent triplet evolution is due to the charge-transfer induced energy level alignment within these compounds. Our results may have potential applications in the design of OLEDs and high triplet yield pure organic materials. [ABSTRACT FROM AUTHOR]

A luminescent probe synthesised via biomimetic crystallization of zinc phosphate in the presence of protein fragment stabilised gold (Au) nanoclusters is reported. The engineered probe – with Au nanoclusters assembled on the crystal – was protease resistant and offered efficient bioimaging and uptake studies. [ABSTRACT FROM AUTHOR]

A novel class of superfluorinated and NIR-luminescent gold nanoclusters were obtained starting from a branched thiol, bearing 27 equivalent 19F atoms per molecule. These unprecedented clusters combine in a unique nanosystem both NIR photoluminescence and 19F NMR properties, thus representing a promising multimodal platform for bioimaging applications. [ABSTRACT FROM AUTHOR]

The cytoskeleton is a key cellular structure that is important in the control of cellular movement, structure, and sensing. To successfully image the individual cytoskeleton components, high resolution and super-resolution fluorescence imaging methods are needed. This review covers the three basic cytoskeletal elements and the relative benefits and drawbacks of fixed versus live cell imaging before moving on to recent studies using high resolution and super-resolution techniques. The techniques covered include the near-diffraction limited imaging methods of confocal microscopy and TIRF microscopy and the super-resolution fluorescence imaging methods of STORM, PALM, and STED. [ABSTRACT FROM AUTHOR]

Janus particles, named after the two-faced Roman god Janus, have different surface makeups, structures or compartments on two sides. This review highlights recent advances in employing Janus particles as novel analytical tools for live cell imaging and biosensing. Unlike conventional particles used in analytical science, two-faced Janus particles provide asymmetry and directionality, and can combine different or even incompatible properties within a single particle. The broken symmetry enables imaging and quantification of rotational dynamics, revealing information beyond what traditional measurements offer. The spatial segregation of molecules on the surface of a single particle also allows analytical functions that would otherwise interfere with each other to be decoupled, opening up opportunities for novel multimodal analytical methods. We summarize here the development of Janus particles, a few general methods for their fabrication and, more importantly, the emerging and novel applications of Janus particles as multi-functional imaging probes and sensors. [ABSTRACT FROM AUTHOR]

A host–guest inclusion complex based on a monofunctionalized pillar[5]arene and a tetraphenylethene derivative was prepared, resulting in an enhanced emission from the tetraphenylethene-based guest, which was applied in cell imaging. [ABSTRACT FROM AUTHOR]

The parameters influencing laser ablation inductively coupled plasma mass spectrometry as a tool for imaging elemental distribution in tissues are critically commented on in this work, and penile cancer tissue was used as a model. General aspects of LA-ICP-MS are discussed, and among them, issues regarding sample preparation and calibration. The optimization process of the following variables is described in detail in a step-by-step tutorial manner: laser intensity, laser frequency, laser resolution, ICP radiofrequency, nebulizer and auxiliary gas flow rates, and C isotopes as internal standards. Once the best condition is found for each variable, MATLAB software is used to generate two-dimensional images. Herein it is also explained how to use MATLAB software to generate tissue images using acquisition, exporting and data treatment parameters. [ABSTRACT FROM AUTHOR]

The advent of laser ablation coupled to inductively coupled plasma-mass spectrometry has allowed for the elemental analysis of solid samples in their native state, or with little sample preparation required. Furthermore, with the addition of a cooled ablation cell spatially resolved elemental imaging of soft and semi-soft samples, specifically biological tissues, is attainable. A new single Peltier element laser ablation cell is described and a demonstration of its applicability to biological sampling is discussed to evaluate its performance. Through the analysis of three different biological tissues the device is shown to provide spatially resolved images of mapped elements while maintaining the structural integrity of the tissues during the hours long sampling times. [ABSTRACT FROM AUTHOR]

A 213 nm Nd:YAG laser ablation system (LA) coupled to a quadrupole ICP-MS was used to ablate thin slices of gelatin and Technovit, both commonly used as standard materials for bioimaging applications. Particle sizes and their distribution within the ablated aerosols were investigated using an optical particle counter inserted in-line between the LA and the ICP-MS, while helium served as carrier gas for material transport. A considerably larger number of particles above 0.3 μm was found for Technovit, while the gelatin aerosol carried mostly particles in the lower nm regime. Increasing the laser fluence during ablation resulted in a larger number of μm sized particles for both materials. Subsequently, fractionation effects during the material transport into the ICP and the ionization within the ICP were analyzed. An increase in the transportation tube length resulted in a signal broadening for shot experiments, but no material loss was observed. A comparison of ionization characteristics of gelatin particles ablated within the LA to wet aerosol particles introduced via nebulization showed similar ionization efficiencies for both aerosols, but the vaporization of gelatin particles required a prolonged time. Additionally, fractionation during the ablation process in gelatin was studied by recording isotope ratios of spiked standards (i.e. Ti and V) for varying laser fluences. The results indicate a strong deviation of up to 20% from the expected 51V/48Ti ratio for laser fluences close to the threshold for ablation. [ABSTRACT FROM AUTHOR]

A laser ablation system (LA) was coupled to an in-house developed low-flow inductively coupled plasma ion source for mass spectrometry (ICP-MS). In this setup, the low-flow torch showed best analytical performance at a total argon gas flow rate of only 1.27 L min−1 and a generator power of 900 W. The optimized system was applied to elemental bioimaging. Two different sections of one kidney sample were analyzed and images of the elements Al and Br from the staining agents hematoxylin and eosin, respectively, and I from a contrast agent iodixanol were recorded selectively and with high sensitivity. For detection of the elements, the mass traces 27Al, 79Br, 81Br, and 127I were selected to demonstrate the system's analytical performance over a wide mass range. In total, the argon consumption could be reduced by ＞90% compared to conventional ICP-MS systems. [ABSTRACT FROM AUTHOR]

Polarization/depolarization levels of different single mitochondria in a cell are inhomogeneous, and always varying. Because depolarization is an indicator of mitochondrial dysfunction, tracing local depolarization is highly desirable. The existing fluorescent probes, however, are not well suited for this task, although they are applicable to assess the average polarization extents of whole cells. A multifunctional and bipolar probe MITFPS is thus developed, which includes a positively charged hydrophilic group and an environment sensitive fluorophore. In the probe design, the hydrophilic anchoring unit is chemically immobilized on a membrane protein, while the lipophilic fluorophore can be inserted deep into the phospholipid layer. The probe exhibits a sensitive response to the local variation in polarization by changing its fluorescence lifetime. MITFPS's applicability is confirmed by real-time in situ imaging of the complete process of an uncoupler-induced depolarization under a two-photon fluorescence lifetime microscope. The imaging result reveals that one mitochondrion could have quite different polarization than the other, even though they are in the same cell. [ABSTRACT FROM AUTHOR]

A Cu2+-specific colorimetric reversible fluorescent receptor HL was designed and synthesized which showed a naked eye observable colour change from colourless to pink on addition of an aqueous buffer (pH 7.4) solution of 30 ppb Cu2+. Short response time (≤5 s) and low detection limit (nearly 3 ppb) make HL suitable as a reliable “dip-in” open eye sensor for Cu2+. Bio-imaging application in live brine shrimp Artemia enabled HL to detect Cu2+ at as low as 10 ppb exposure. [ABSTRACT FROM AUTHOR]

Recently, the strategy of protection–deprotection of functional groups has been widely employed to design fluorescent probes, as the protection–deprotection of functional groups often induces a marked change in electronic properties. Significant advances have been made in the development of analyte-responsive fluorescent probes based on the protection–deprotection strategy. In this tutorial review, we highlight the representative examples of small-molecule based fluorescent probes for bioimaging, which are operated via the protection–deprotection of key functional groups such as aldehyde, hydroxyl, and amino functional groups reported from 2010 to 2014. The discussion includes the general protection–deprotection methods for aldehyde, hydroxyl, or amino groups, as well as the design strategies, sensing mechanisms, and deprotection modes of the representative fluorescent imaging probes applied to bio-imaging. [ABSTRACT FROM AUTHOR]

A turn-on fluorescein-based probe (1) has been successfully developed for the highly sensitive and selective detection of sulfite. The probe exhibits high SO32− selectivity over other common anions and biothiols (Cys, Hcy and GSH), and the detection limit was measured to be 0.55 μM. Probe 1 exhibited high selectivity towards SO32− with a broad pH (4–9). A significant change from colorless to yellow could be directly observed by the naked eye. Furthermore, living cell imaging results demonstrated its practical application value in biological systems. [ABSTRACT FROM AUTHOR]

Synthetic, spectroscopic, computational and biological imaging studies of platinum trimethyl bipyridyl thiolate complexes of the general formula [PtMe3(bpy)SR] reveal these to be easily accessed, tunable bioimaging agents which feature an unusual σ–π* Inter-Ligand Charge Transfer (ILCT) transition, and in some cases emit into the Near infra-red (NIR). [ABSTRACT FROM AUTHOR]

Probe E3 has been developed as the first ratiometric fluorescent cobalt probe with high sensitivity and selectivity based on internal charge transfer (ICT). Most importantly, the probe achieved the imaging and detection of cobalt in cells with ratiometric measurement. [ABSTRACT FROM AUTHOR]

A dye emitted red fluorescence with aggregation-enhanced emission properties was reported here. It can be utilized to specifically recognize the cell membrane of MCF-7 and 293T cell lines during bio-imaging. [ABSTRACT FROM AUTHOR]

Multi-modal imaging in tissue and animals has integrated the advantages of individual molecular imaging modalities and shown great potential in both pre-clinical and clinical fields. In the present work, a high-performance nanoparticulate contrast agent based on PEGylated gadolinium hydroxycarbonate nanoparticles (PEG-GdNPs) was designed and synthesized via a facile approach. Due to the outstanding X-ray absorption coefficient/K-edge value and the presence of unpaired 4f electrons of element Gd, our well-prepared nanoprobes were successfully used as dual-modal contrast agents for in vivo synergistic magnetic resonance and X-ray computed tomography imaging. Bio-distribution of the nanoparticles was further studied via in vivo long-term tracking during the imaging process, revealing that most of the nanoprobes could accumulate in the liver and therefore act as a liver-targeted imaging agent. Moreover, toxicity including cell viability, blood biochemical assay, as well as changes in body weight and histology indicated the low systemic toxicity of our nanoprobes. Taken together, we believe that PEG-GdNPs with their excellent imaging capability and high biocompatibility may find widespread application in the field of biomedicine. [ABSTRACT FROM AUTHOR]

Dual modal nanoprobes are promising tools for accurately detecting target molecules as part of the diagnosis of diseases including cancers. We have explored a new dual modal bioimaging probe that is comprised of a quantum dot (QD)–magnetic nanoparticle (MNP) hybrid. The MNP–QD heterodimers explored are fabricated by using a platinum–guanine coordination bonding guided self-assembly process, employing the metal–DNA conjugation method. Investigations utilizing energy dispersive spectroscopy (EDS) equipped high resolution transmission electron microscopy (HRTEM) demonstrate that the heterodimer contains an iron (Fe) dominant MNP and a cadmium (Cd) dominant QD. Finally, the results of cell studies show that the MNP–QD conjugates display good HeLa cell uptake in the absence of non-specific binding to the cell membrane and, as such, they can be used to label cells in vitro and in vivo as part of a new cell imaging technique. [ABSTRACT FROM AUTHOR]

The first two-photon fluorescent probe for specific detection of nitroxyl is designed and synthesized, and we have further demonstrated that the new two-photon fluorescent probe could be employed to image nitroxyl in living cells and tissues. [ABSTRACT FROM AUTHOR]

Laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) has received significant attention over the last 10 years and has been widely used for the analysis of biological samples. The technique allows the determination of elements and isotopes in biological tissues and related materials with a spatial resolution typically ranging from 10 to 100 µm. When compared to other techniques usually employed to obtain bioimages, the greater advantage of LA-ICP-MS is its higher sensitivity. The literature survey over the last 10 years concerning the use of LA-ICP-MS for biological tissue analysis is reviewed in this article. Instrumentation, strategies of calibration for quantitative analysis, challenges and recent advances in this field are discussed. Applications of the isotope ratio (IR), including tracer experiments, and isotope dilution (ID), are reviewed for biological samples (briefly for proteins, in order to show the utility of LA-ICP-MS). Bioimaging methods, studies and applications for animal and plants tissues are emphasized, demonstrating the importance of bioimaging of metals and metalloids in biomedical research, bioaccumulation and bioavailability studies for ecological and toxicological risk assessment in humans, animals and plants. The usefulness of the IR associated with bioimaging for predicting geographical origin, habitat, movement of subjects, diet and lifestyle are also demonstrated. [ABSTRACT FROM AUTHOR]

The ability to 'see' down to nanoscale has always been one of the most challenging obstacles for researchers to address fundamental questions. For many years, researchers have been developing scanning probe microscopy techniques to improve imaging capability at nanoscale. Among them, atomic force microscopy (AFM) has received considerable attention, which allows probing topography of biological species at real space under physiological environment. Importantly, force measurements in AFM enable researchers to reveal not only the topography but also the relevant physical-chemical properties. AFM-based dynamic nanomechanical mapping (DNM) provides insights into the functions of biological systems by the interpretation of 'force', which are inaccessible by most of the other analytic techniques. This review is aiming to shed light on these recently developed AFM-based DNM techniques for biomolecular imaging, and discuss the relative applications in biological research from the nanomechanical point of view. [ABSTRACT FROM AUTHOR]

A diformyl-p-cresol (DFC)-8-aminoquinoline based dual signaling probe was found to exhibit colorimetric and fluorogenic properties on selective binding towards Mg2+ and Zn2+. Turn-on fluorescent enhancements (FE) as high as 40 fold and 53 fold in 9 : 1 MeCN/water (v/v) at pH 7.2 in HEPES buffer for Mg2+ and Zn2+, respectively, were observed. The binding constants determined from the fluorescence titration data are: K = (1.52 ± 0.21) × 105 M−1 and (9.34 ± 4.0) × 103 M−2 at n = 1 and 0.5, for Mg2+ and Zn2+, respectively. The L : M binding ratios were also determined by Job's method, which support the above findings. This is further substantiated by HRMS analysis. Due to solubility in mixed organo-aqueous solvents as well as cell permeability it could be used for the in vitro/in vivo cell imaging of Mg2+ and Zn2+ ions with no or negligible cytotoxicity. This probe could be made selective towards Mg2+ over Zn2+ in the presence of TPEN, both under intra- and extracellular conditions and is superior to other Mg2+ probes which suffer from selectivity of Mg2+ over Ca2+ or Zn2+. Furthermore the dissociation constant (Kd = 6.60 μM) of the Mg2+-(DFC-8-AQ) complex is far lower than the so far reported Mg2+ probes which fall in the mM range. [ABSTRACT FROM AUTHOR]

The observation of viral plaques is the standard method for determining the viral titer and understanding the behaviors of viruses. Here, we report the application of a wide field-of-view (FOV), time-lapse, on-chip imaging platform, termed the ePetri, for plaque analysis of murine norovirus 1 (MNV-1). The ePetri offers the ability to dynamically track plaques at the individual cell death event level over a wide FOV of 6 mm × 4 mm. As demonstration, we captured high-resolution time-lapse images of MNV-1-infected cells at 30 min intervals. We implemented a customized image-processing program containing a density-based clustering algorithm to analyze the spatial-temporal distribution of cell death events to identify plaques at their earliest stages. By using the results in a viral titer count format, we showed that our approach gives results that are comparable to conventional plaque assays. We further showed that the extra information collected by the ePetri can be used to monitor the dynamics of plaque formation and growth. Finally, we performed a demonstration experiment to show the relevance of such an experimental format for viral inhibitor study. We believe the ePetri is a simple and compact solution for the automation of viral plaque assays, plaque behavior analysis, and antiviral drug discovery and study. [ABSTRACT FROM AUTHOR]

Matrix effects in mass spectrometry imaging (MSI) may affect the observed molecular distribution in chemical and biological systems. In this study, we use mouse brain tissue of a middle cerebral artery occlusion (MCAO) stroke model to examine matrix effects in nanospray desorption electrospray ionization MSI (nano-DESI MSI). This is achieved by normalizing the intensity of the sodium and potassium adducts of endogenous phosphatidylcholine (PC) species to the intensity of the corresponding adduct of the PC standard supplied at a constant rate with the nano-DESI solvent. The use of MCAO model with an ischemic region localized to one hemisphere of the brain enables immediate comparison of matrix effects within one ion image. Furthermore, significant differences in sodium and potassium concentrations in the ischemic region in comparison with the healthy tissue allowed us to distinguish between two types of matrix effects. Specifically, we discuss matrix effects originating from variations in alkali metal concentrations and matrix effects originating from variations in the molecular composition of the tissue. Compensation for both types of matrix effects was achieved by normalizing the signals corresponding to endogenous PC to the signals of the standards. This approach, which does not introduce any complexity in sample preparation, efficiently compensates for signal variations resulting from differences in the local concentrations of sodium and potassium in tissue sections and from the complexity of the extracted analyte mixture derived from local variations in molecular composition. [ABSTRACT FROM AUTHOR]

Environment-related parameters, including viscosity, polarity, temperature, hypoxia, and pH, play pivotal roles in controlling the physical or chemical behaviors of local molecules. In particular, in a biological environment, such factors predominantly determine the biological properties of the local environment or reflect corresponding status alterations. Abnormal changes in these factors would cause cellular malfunction or become a hallmark of the occurrence of severe diseases. Therefore, in recent years, they have increasingly attracted research interest from the fields of chemistry and biological chemistry. With the emergence of fluorescence sensing and imaging technology, several fluorescent chemosensors have been designed to respond to such parameters and to further map their distributions and variations in vitro/in vivo. In this work, we have reviewed a number of various environment-responsive chemosensors related to fluorescent recognition of viscosity, polarity, temperature, hypoxia, and pH that have been reported thus far. [ABSTRACT FROM AUTHOR]

Fluorescence provides a mechanism for achieving contrast in biological imaging that enables investigations of molecular structure, dynamics, and function at high spatial and temporal resolution. Small-molecule organic fluorophores have proven essential for such efforts and are widely used in advanced applications such as single-molecule and super-resolution microscopy. Yet, organic fluorophores, like all fluorescent species, exhibit instabilities in their emission characteristics, including blinking and photobleaching that limit their utility and performance. Here, we review the photophysics and photochemistry of organic fluorophores as they pertain to mitigating such instabilities, with a specific focus on the development of stabilized fluorophores through derivatization. Self-healing organic fluorophores, wherein the triplet state is intramolecularly quenched by a covalently attached protective agent, exhibit markedly improved photostabilities. We discuss the potential for further enhancements towards the goal of developing "ultra-stable" fluorophores spanning the visible spectrum and how such fluorophores are likely to impact the future of single-molecule research. [ABSTRACT FROM AUTHOR]

The use of organic fluorescent probes in biochemical and biophysical applications of single molecule spectroscopy and fluorescence microscopy techniques continues to increase. As single molecule measurements become more quantitative and new developments in super-resolution imaging allow researchers to image biological materials with unprecedented resolution, it is becoming increasingly important to understand how the properties of the probes influence the signals measured in these experiments. In this review, we focus on the photochemical and photophysical processes of organic fluorophores that affect the properties of fluorescence emission. This includes photobleaching, quenching, and the formation of non-emissive (dark) states that result in fluorescence blinking in a variety of timescales. These processes, if overlooked, can result in an erroneous interpretation of the data. Understanding their physical origins, on the other hand, allows researchers to design experiments and interpret results so that the maximum amount of information about the system of interest can be extracted from fluorescence signals. [ABSTRACT FROM AUTHOR]

To explore the photophysical properties of coordination compounds with enhanced two-photon absorption, two novel six-coordinated metal complexes (ML2, M = Cd(II), Zn(II)) from carbazole β-diketone ligand (HL = 4,4,4-trifluoro-1-(9-butylcarbazole-3-yl)-1,3-butanedione) were prepared and fully characterized. Their crystal structures were determined by X-ray diffraction analysis. Both variable temperature ¹H NMR spectra and MALDI-TOF mass spectrometry proved that the coordination compounds exhibit good stability in solution. The results of time-dependent density functional theory (TD-DFT) calculations indicated that the complexation of the ligands with metal ion extends the electronic delocalization in the coordination compounds, leading to enhanced two-photon absorption. The photophysical properties for the coordination compounds were identified relying on both experimentally and theoretically studies. Finally, confocal microscopy and two-photon microscopy fluorescent imaging of HepG2 cells labeled with the Zn(II) complexe revealed its potential applications as a biological fluorescent probe. [ABSTRACT FROM AUTHOR]

A facile hydrothermal method was used to synthesize luminescent carbon dots from single-walled carbon nanotubes (SWNTs). Then 4,7,10-trioxa-1,13-tridecanediamine (TTDDA) was grafted onto the carbon dots to increase their water solubility and luminescent properties. The as-prepared carbon dots were characterized by UV-Vis absorption, infrared (IR), Raman, fluorescence, and transmission electron microscopy (TEM). The results confirmed that these carbon dots were monodispersed in water and emitted bright yellow fluorescence. The surface modified carbon dots were used to label biological cells, and the images obtained by a laser scanning confocal microscope showed that the carbon dots were gradually taken up by HeLa cells. The cytotoxicity of such carbon dots toward HeLa cells was very low, for 24 h the LC50 was over 5 mg mL−1. [ABSTRACT FROM AUTHOR]

As a unique nanomaterial, quantum dots (QDs) are not only applied in fluorescent labeling and biological imaging, but are also utilized in novel sensing systems. Because QDs have attractive optoelectronic characteristics, QD-based sensors present high sensitivity in detecting specific analytes in the chemical and biochemical fields. In this review, we describe the basic principles and different conjugation strategies in QD-based sensors. An overview of recent advances and various models of QD-sensing systems is also provided. Furthermore, perspectives for sensors based on QDs are discussed. [ABSTRACT FROM AUTHOR]

A new self-assembly nanoprobe, mercaptoethylamine-modified-gold nanoparticles–Lysine-bridged-bis(β-cyclodextrins)–fluorescein (MGNPs–Lys-bis(β-CDs)–FL), based on fluorescence resonance energy transfer (FRET) was developed for determination of trypsin firstly in biological systems. With the Lys-bis(β-CDs)–FL complex as an energy donor and mercaptoethylamine (MEA)-modified gold nanoparticles (MGNPs) as an energy acceptor, the two parts assemble an efficient FRET nanoprobe through an amide bond. Trypsin is specific for the hydrolysis of amide linkages of lysine. Therefore, in the presence of trypsin, the nanoprobe is cleaved by trypsin on the binding sites of amide with good specificity and sensitivity, resulting in the fluorescence recovery of the quenched FL. The nanoprobe has good biological applicability and provides a potential assay for further clinical research of trypsin in biosystems. [ABSTRACT FROM AUTHOR]

A novel calibration scheme for elemental analysis and imaging of biological thin sections by laser ablation ICP mass spectrometry is described. The procedure is based on spiking whole blood or blood serum with elemental standards (Sr, Gd, Pt), freezing the spiked aliquots in a customised block of CMC and sectioning by cryomicrotome. The approach is robust, convenient to implement and affords matrix matching of standards to samples in terms of chemistry and physical properties. Linear calibration and good method reproducibility was achieved with limits of detection at the ng g−1level. Application to quantitative analysis and imaging of metallodrugs (Sr, Pt based) and MRI contrast agent (Gd based) was demonstrated. [ABSTRACT FROM AUTHOR]

Quantum dot toxicity has become a hot topic in recent years due to the emergence of semiconductor nanoparticles as highly efficient biological imaging agents. The use of quantum dots in biology is arguably the most successful application of pure nanotechnology in recent times, although unfortunately, the most useful semiconductor particles contain elements that are often thought to be detrimental to health and the environment. In this article, we explore some key reports on this issue. [ABSTRACT FROM AUTHOR]

In this paper, we report new rhodamine hydrazone derivatives bearing thiol and carboxylic acid groups as selective fluorescent and colorimetric chemosensors for Hg2+. The ring-opening process of spirolactam enables the large fluorescent enhancement and colorimetric change upon the addition of Hg2+. The sample containing Hg2+was mixed with one of the chemosensors in a microchannel where the sensor was examined using confocal laser scanning microscopy. A plot of the fluorescent intensities of both chemosensors versusthe log concentration of Hg2+exhibited a linear response (r2= 0.95) in the range of 1 nM–1 μM, and the detection limits were 1 nM and 4.2 nM, respectively. Both chemosensors also enable the visualization of Hg2+accumulated in the nematode Caenorhabditis eleganspreviously exposed to nanomolar concentrations of Hg2+. [ABSTRACT FROM AUTHOR]

Confocal Raman micro-spectroscopy (CRMS) was used to measure spectral images of immunological synapse formation between dendritic and T cells without using molecular labels or other invasive procedures. The purpose-built inverted CRMS instrument integrated an environmental enclosure and a near-infrared laser to allow measurements on live cells maintained under physiological conditions. The integration of the wide-field fluorescence also enabled viability assays and direct comparison between Raman spectral images and gold-standard immuno-fluorescence images for specific molecules. Raman spectral images of nucleus and proteins were built by fuzzy c-mean clustering method. The Raman images were found to be in good correspondence with the immuno-fluorescence images of DNA and actin. These results indicate that actin is a main contributor to the Raman spectrum of the cytoplasm of dendritic and T cells. While for control cells the Raman spectral images of proteins indicated a more homogeneous distribution of proteins in the cytoplasm of dendritic cells, they indicated a higher accumulation of proteins at the immunological synapses when dendritic cells were pre-treated with laminin. These conclusions were also supported by confocal immuno-fluorescence imaging after cell fixation and labelling. This study demonstrates the potential of CRMS for label-free non-invasive imaging of junctions between live cells. Therefore, this technique may become a useful tool for studying cellular processes in live cells and where non-invasive molecular specific imaging is desirable, such as cell–cell interactions. [ABSTRACT FROM AUTHOR]

The emerging field of bionanotechnology aims at revolutionizing biomedical research and clinical practice viaintroduction of nanoparticle-based tools, expanding capabilities of existing investigative, diagnostic, and therapeutic techniques as well as creating novel instruments and approaches for addressing challenges faced by medicine. Quantum dots (QDs), semiconductor nanoparticles with unique photo-physical properties, have become one of the dominant classes of imaging probes as well as universal platforms for engineering of multifunctional nanodevices. Possessing versatile surface chemistry and superior optical features, QDs have found initial use in a variety of in vitroand in vivoapplications. However, careful engineering of QD probes guided by application-specific design criteria is becoming increasingly important for successful transition of this technology from proof-of-concept studies towards real-life clinical applications. This review outlines the major design principles and criteria, from general ones to application-specific, governing the engineering of novel QD probes satisfying the increasing demands and requirements of nanomedicine and discusses the future directions of QD-focused bionanotechnology research (critical review, 201 references). [ABSTRACT FROM AUTHOR]