Strong aggregation-induced electrochemiluminescence of pyrene-coordination metal-organic frameworks coupled with zero-valent iron as novel accelerator for ultrasensitive immunoassay.

Polycyclic aromatic hydrocarbons (PAHs) are excellent alternative luminophores for electrochemiluminescence (ECL) immunoassays. However, they are inevitably limited by the aggregation-caused quenching effect. In this study, aimed at eliminating the aggregation quenching of PAHs, luminescent metal-organic frameworks (MOFs) with 1,3,6,8-tetra(4-carboxybenzene)pyrene (H4TBAPy) as the ligand were exploited as a novel nano-emitter for the construction of ECL immunoassays. The luminophore exhibits efficient aggregation-induced emission enhancement, good acid-base resistance property and unusual ECL reactivity. In addition, the simultaneous use of potassium persulfate and hydrogen peroxide as dual co-reactants resulted in a synergistic enhancement of the cathodic ECL efficiency. The use of magnetic iron-nickel alloys as the multifunctional sensing platform can further enhance the ECL activity, and its enriched zero-valent iron as a co-reactant accelerator effectively drives ECL analytical performance. Profiting from the excellent characteristics, signal-on ECL immunoassays have been constructed. With carcinoembryonic antigen as the model analysis target, a detection limit of 0.63 pg/mL was obtained within the linear range of 1 pg/mL to 50 ng/mL, accompanied by excellent analytical performance. This report opens a new window for the rational design of efficient ECL illuminators, and the proposed ECL immunoassays may find promising applications in the detection of disease markers.

Preparation of microbial agent immobilized composites for Cr(VI) removal from wastewater.

Because of its extreme toxicity and health risks, hexavalent chromium [Cr(VI)] has been identified as a major environmental contaminant. Bioreduction is considered as one of effective techniques for cleaning up Cr(VI)-contaminated sites, but the remediation efficiency needs to be enhanced. Here, a novel immobilized microbial agent was produced by immobilizing Bacillus cereus ZY-2009 with sodium alginate (SA) using polyvinyl alcohol (PVA) and activated carbon (AC). To evaluate the decrease of Cr(VI) by immobilized bacterial agents, batch tests were conducted with varying immobilization conditions, immobilization carriers, and dosages of medication. The removal of Cr(VI) by the agent prepared by the composite immobilization method was better than that by the adsorption and encapsulation methods. The optimal preparation conditions were the fraction of magnetic PVA was 5.00%, the fraction of SA was 4.00%, the fraction of CaCl2 was 4.00%, and the calcification time was 12 h. The experimental results indicated that PVA/SA/AC agents accelerated the reduction rate of Cr(VI). The removal rate of Cr(VI) by immobilized cells (90.5%) under ideal conditions was substantially higher than that of free cells (11.0%). This novel agent had a large specific surface area and a rich pore structure, accounting for its high reduction rate. The results suggest that the PVA/SA/AC immobilized Bacillus cereus ZY-2009 agent has great potential to remove Cr(VI) from wastewater treatment systems.

A signal polarity conversion photoelectrochemical immunosensor for neuron-specific enolase detection based on MgIn2S4-sensitized CsPbBr3.

A photoelectrochemical (PEC) immunosensor was designed based on MgIn2S4-decorated inorganic halide perovskite CsPbBr3 combined with the signal polarity conversion strategy for neuron-specific enolase (NSE) detection. CsPbBr3 was applied as the basic photoactive material owing to its excellent optical and electronic properties, which provide a good PEC performance for sensor construction. In order to improve the stability of this perovskite, the three-dimensional flower-like MgIn2S4 with a desirable direct band gap was applied to enhance the PEC response. Also, the excellent structure of MgIn2S4 provides large surface-active sites for CsPbBr3 loaded. For enhancing the detection sensitivity of PEC immunosensor, p-type CuInS2 was used as a signal probe which fixed on detection antibody (Ab2). When the target NSE was present, the photogenerated electrons produced by CuInS2 were transferred to the test solution, and the polarity of PEC signal changes. Based on the above photosensitive materials and signal conversion strategy, the proposed PEC immunosensor showed favorable detection performance, and the linear detection range is 0.0001 ~ 100 ng/mL with a 38 fg/mL of detection limit. The proposed strategy improved the adhibition of CsPbBr3 in the analytical chemistry field as well as provided a reference method for other protein detections.

Antioxidant Cascade Modulated Electrochemiluminescence by a Biomimetic Metal-Organic Framework with Dual Enzymatic Activity for Disease Marker Immunoassays.

High-performance electrochemiluminescence is a significant approach for the examination of disease biomarkers, and the utilization of innovative electrochemiluminescence detection systems represents a viable strategy to enhance the efficacy of ECL analysis. In this work, the biomimetic engineering metal-organic framework (MOF-818) has realized the ultrasensitive ECL immunoassay of disease markers based on the guidance of the free radical scavenging strategy provided by the antioxidant cascade. Initially, we synthesized a hydrogen-bonded organic framework (HOF) consisting of luminol and three active ligands based on simple room-temperature self-assembly. The luminol-HOF (L-HOF) showed more stable and brighter ECL luminescence activity than the monomer due to the nano-confinement enhancement of the coordinated luminol units. Subsequently, MOF-818 with biomimetic superoxide dismutase (SOD) and catalase (CAT) activities were recruited for the first time as quenching agents for sandwich immunoassay mode. The enzyme activity leads to the reverse transformation of superoxide anion radicals (O2-) and further antioxidant decomposition, decreasing in the responsiveness of luminol ECL signals. Using carcinoembryonic antigen (CEA) as an analytical model, a detection limit of 0.457 pg/mL was obtained within a detection range of 0.001-50 ng/mL. We believe that this novel sandwich sensing model based on enzyme activity provides a meaningful potential tool for precise detection, expanding the broader application of nanoenzymes in analysis.

Lifestyle risks associated with brain functional connectivity and structure.

Some lifestyle factors are related to health and brain function and structure, but the brain systems involved are incompletely understood. A general linear model was used to test the associations of the combined and separate lifestyle risk measures of alcohol use, smoking, diet, amounts of physical activity, leisure activity, and mobile phone use, with brain functional connectivity with the high resolution Human Connectome Project (HCP) atlas in 19,415 participants aged 45-78 from the UK Biobank, with replication with HCP data. Higher combined lifestyle risk scores were associated with lower functional connectivity across the whole brain, but especially of three brain systems. Low physical, and leisure and social, activity were associated with low connectivities of the somatosensory/motor cortical regions and of hippocampal memory-related regions. Low mobile phone use, perhaps indicative of poor social communication channels, was associated with low functional connectivity of brain regions in and related to the superior temporal sulcus that are involved in social behavior and face processing. Smoking was associated with lower functional connectivity of especially frontal regions involved in attention. Lower cortical thickness in some of these regions, and also lower subcortical volume of the hippocampus, amygdala, and globus pallidus, were also associated with the sum of the poor lifestyle scores. This very large scale analysis emphasizes how the lifestyle of humans relates to their brain structure and function, and provides a foundation for understanding the causalities that relate to the differences found here in the brains of different individuals.

Orbitofrontal cortex connectivity is associated with food reward and body weight in humans.

The aim was to investigate with very large-scale analyses whether there are underlying functional connectivity differences between humans that relate to food reward and whether these in turn are associated with being overweight. In 37 286 humans from the UK Biobank, resting-state functional connectivities of the orbitofrontal cortex (OFC), especially with the anterior cingulate cortex, were positively correlated with the liking for sweet foods (False Discovery Rate (FDR) P < 0.05). They were also positively correlated with the body mass index (BMI) (FDR P < 0.05). Moreover, in a sample of 502 492 people, the 'liking for sweet foods' was correlated with their BMI (r = 0.06, P < 10-125). In a cross-validation with 545 participants from the Human Connectome Project, a higher functional connectivity involving the OFC relative to other brain areas was associated with a high BMI (≥30) compared to a mid-BMI group (22-25; P = 6 × 10-5), and low OFC functional connectivity was associated with a low BMI (≤20.5; P < 0.024). It is proposed that a high BMI relates to increased efficacy of OFC food reward systems and a low BMI to decreased efficacy. This was found with no stimulation by food, so may be an underlying individual difference in brain connectivity that is related to food reward and BMI.

High-Performance Electrochemiluminescence of a Coordination-Driven J-Aggregate K-PTC MOF Regulated by Metal-Phenolic Nanoparticles for Biomarker Analysis.

The elimination of aggregation-caused quenching of polycyclic aromatic hydrocarbons by metal-ligand coordination is of immense scientific interest in solid-state electrochemiluminescence (ECL) sensing. Herein potassium ion (K+)-mediated J-aggregate K-PTC MOF (PTCA, perylene-3,4,9,10-tetracarboxylic) was synthesized and employed to formulate an ECL immunosensor for biomarker detection. The coordination-driven aggregates are arranged in an end-to-end side mode, which overcomes the aggregation-caused quenching related to PTCA concentration. Compared with PTCA, K-PTC MOF shows a more stable ECL emission with an unprecedented red shift to 718 nm and is equipped with ECL activity for analytical applications at a voltage of -1.1 V. Considering the requirements of accurate detection, metal-phenolic bioactive nanoparticles (MPNPs) were synthesized for the construction of a sandwich sensing platform to realize the steady-state regulation of ECL. As proof of applicability, a constructive experiment was carried out with neuron-specific enolase (NSE), a marker of small cell lung cancer (SCLC), as a targeted analyte. With optimal requirements, the configuration can provide a detection range of 10 pg/mL to 50 ng/mL and a detection limit of 7.4 pg/mL, accompanied by sufficient practical analytical performance. Collectively, this paradigm provides a deeper understanding of the ECL characteristics of coordination-driven J-aggregation and provides more possibilities for the development of ECL patterns based on luminescent metal-organic frameworks.

A novel targeted iron oxide nanocarrier for inhibiting M2-type macrophages in the tumor microenvironment.

Background: Tumor-associated macrophages (TAMs) are vital to the tumor microenvironment. They are classified as antitumor M1-type or protumor M2-type macrophages. M2-type macrophages accumulate in the tumor stroma and are related to poor prognosis. Iron oxide nanoparticles are used as drug delivery vehicles because of the structure of carboxyl groups on their surface and their ability to be easily phagocytosed by macrophages. Aim: The signal transducer and activator of transcription 6 (STAT6) signaling pathway controls M2 macrophage polarization, but the STAT6 signaling pathway inhibitor AS1517499 lacks efficient targeting in vivo. Thus, our study aimed to block the polarization of TAMs to M2-type macrophages. Methods and Material: We used ultrasmall superparamagnetic iron oxide nanoparticles (USPIONs) as drug carriers coated with the STAT6 signaling pathway inhibitors AS1517499 and CD163 monoclonal antibodies to synthesize the targeted nanocomplex AS1517499-USPION-CD163 utilizing the carbodiimide method. Then, we determined its physicochemical properties, including hydrodynamic size distribution, ultrastructure, iron concentration, protein content and activity of the CD163 monoclonal antibody, AS1517499 content, and selectivity for M2-type macrophages, and its biological applications. Results: The hydrodynamic size distribution was stable (average size = 95.37 nm). Regarding biological applications, the targeted nanocomplex selectively inhibited M2-type macrophages. Conclusions: The targeted nanocomplex AS1517499-USPION-CD163 showed high selectivity for M2-type macrophages. Therefore, iron oxide nanoparticles targeting TAMs may be an effective approach to TAM therapy.

Dumbbell Plate-Shaped AIEgen-Based Luminescent MOF with High Quantum Yield as Self-Enhanced ECL Tags: Mechanism Insights and Biosensing Application.

It is widely known that high-performance electrochemiluminescence (ECL) emitters play a crucial part in improving the detection sensitivity of the ECL strategy. Through the combination of aggregation-induced emission luminogens (AIEgens), 1,1,2,2-tetra(4-carboxylbiphenyl)ethylene (H4 TCBPE) with Zr(IV) cations, a dumbbell plate-shaped metal-organic framework (MOF) with high luminous efficiency is synthesized as ECL tags. The resultant MOF exhibits stronger ECL activity than those of H4 TCBPE monomers and aggregates. Herein, this phenomenon is defined as the coordination-triggered electrochemiluminescence (CT-ECL) enhancement effect. Furthermore, the nearly matched ECL and photoluminescence (PL) spectra imply the bandgap emission mechanism. Remarkably, polyethyleneimine (PEI) as the coreactant is covalently connected with MOF to form the uniquely self-enhanced ECL complex of Zr-TCBPE-PEI, where the robust ECL signal is captured owing to the intramolecular-like coreaction acceleration. Based on the resonance energy transfer (RET) behavior, the AuPd@SiO2 composite is designed as the high-efficiency quencher. In this manner, an innovative and ultrasensitive ECL sensor is constructed for neuron-specific enolase (NSE) detection through sandwich-type immunoreaction, with the detection limit down to 52 fg ml-1 . The present study has gone some way toward designing MOF-based self-luminescent ECL materials, thus paving a new avenue to expand the late-model ECL emitters for immunoassay.

PEGylation Improved Electrochemiluminescence Supramolecular Assembly of Iridium(III) Complexes in Apoferritin for Immunoassays Using 2D/2D MXene/TiO2 Hybrids as Signal Amplifiers.

Dynamic self-assembly of iridium complexes in water-soluble nanocontainers is an important bottom-up process for fabricating electrochemiluminescence (ECL) bioprobes. PEGylated apoferritin (PEG-apoHSF) as the host offers a confined space to alter and modify the self-assembly of trans-bis(2-phenylpyridine)(acetylacetonate)iridium(III) [Ir(ppy)2(acac)] based on a pH-dependent depolymerization/reassembly pathway, allowing the formation of ECL-active iridium cores in PEG-apoHSF cavities (Ir@PEG-apoHSF). With an improved encapsulation ratio in PEG-apoHSF, the coreactant ECL behavior of the fabricated Ir@PEG-apoHSF nanodots with tri-n-propylamine (TPrA) was further demonstrated, exhibiting maximum ECL emission at 530 nm that was theoretically dominated by the band gap transition. The application of Ir@PEG-apoHSF as a bioprobe in a "signal-on" ECL immunosensing system was developed based on electroactive Ti3C2Tx MXenes/TiO2 nanosheet (Ti3C2Tx/TiO2) hybrids. Combining with the efficiently catalyzed electro-oxidation of TPrA and Ir(ppy)2(acac) by Ti3C2Tx/TiO2 hybrids, the developed immunosensor showed dramatically amplified ECL responses toward the target analyte of neuron-specific enolase (NSE). Under experimental conditions, linear quantification of NSE from 100 fg/mL to 50 ng/mL was well established by this assay, achieving a limit of detection (LOD) of 35 fg/mL. The results showcased the capability of PEGylated apoHSF to host and stabilize water-insoluble iridium complexes as ECL emitters for aqueous biosensing and immunoassays.

Hypertension is associated with reduced hippocampal connectivity and impaired memory.

BACKGROUND: The objective was a large-scale analysis of the relation between hypertension, memory problems, and brain function. METHODS: The study design was to measure the association between a history of hypertension, and the functional connectivity between 94 brain regions, and prospective and numeric memory, in 19,507 participants from the UK Biobank, with cross-validation in 1,002 participants in the Human Connectome Project, and 13,441 individuals in the second release of the UK Biobank. A history of hypertension was measured by whether individuals were admitted to hospital for the treatment of hypertension, with the control group admissions for other reasons. FINDINGS: A history of hypertension was associated with reduced functional connectivity of the hippocampus, and with reduced prospective memory score (FDR correction p<0.01). The reduced functional connectivity mediated the association between the hypertension history and the prospective memory score. A graded linear relation between both the hippocampal functional connectivity and memory impairment, was found across a wide range of blood pressure (r=-0.04). In 502,537 participants from the UK Biobank, a history of hypertension was associated with impaired prospective memory (p = 9.1 × 10-41, Cohen's d=-0.08) and numeric memory (p = 4.7 × 10-24, Cohen's d=-0.10). The association between hypertension, functional connectivity, and impaired memory was cross-validated with 1,002 participants from the Human Connectome Project; and for functional connectivity in 13,441 individuals in the second release of the UK Biobank imaging dataset. INTERPRETATION: The reduced functional connectivity of the hippocampus, and the memory impairments, both related to hypertension across a wide range of blood pressure, are important for clinical practice.

Recognition of M2 type tumor-associated macrophages with ultrasensitive and biocompatible photoelectrochemical cytosensor based on Ce doped SnO2/SnS2 nano heterostructure.

Tumor-associated macrophages (TAMs) play central roles in the regulation of tumor growth. TAMs can be differentiated into M1 and M2 types, which are responsible for the inhibition and growth of tumor tissues, respectively. Recognition of M2-TAMs is significant for the diagnosis and therapy of cancer, which is however severely limited due to the deficiency of selective and sensitive photoelectrochemical sensors. In this work, using Ce doped SnO2/SnS2 nano heterostructure as the highly sensitive platform, a photoelectrochemical sensor enabling the recognition of M2-TAMs was fabricated for the first time. By the decoration of CD163 antibody on the platform, the ultrasensitive photoelectrochemical sensor can selectively detect the CD163 protein on the surface of M2-TAMs. To our best knowledge, this is the first demonstration for recognition of M2-TAMs using photoelectrochemical method. The fabricated cytosensor has ultra-sensitive photocurrent response, applicable biological compatibility, high selectivity and relatively wide linear sensing range (5 × 101 to 1 × 105 cells/ml) with a low detection limit (50 cells/ml) for the detection of M2-TAMS. This kind of PEC cytosensor would provide a novel analysis and detection strategy for M2-TAMs.

Novel Electron Donor Encapsulation Assay Based on the Split-type Photoelectrochemical Interface.

In this research, a controlled-release photoelectrochemical (PEC) immunosensor is proposed on the basis of a novel encapsulation strategy by all-inorganic semiconductor materials. The controlled-release transmit system has been prepared and represented on account of a group-functional mesoporous silica nanosphere (MSN), utilizing surface-functionalized cadmium sulfide (CdS) nanoparticles as mobilizable caps to encapsulate a PEC electron donor ascorbic acid (AA) within the MSN mesoporous structure. This encapsulation strategy proceeds without any enzyme and acid/alkali to achieve the release of an electron donor. The complex is formed by encapsulating AA within MSN with CdS (CdS@MSN-AA) as a signal amplifier labeled on the secondary antibody. In addition, the immunological recognition process was performed in a 96-well plate, and the reciprocal interference between biorecognition and PEC analysis could be eliminated through a split-type framework. Bi2S3-sensitized porous In2O3 nanoparticles as a substrate matrix provide basic PEC response. The developed sensor exhibited a mensurable output of procalcitonin (PCT) concentration (as an example) in the detection range of 0.001-200 ng/mL along with a limit of detection of 0.31 pg/mL. Featuring the novel method for electron release, this sensitive PEC strategy provides an innovative way for the potential application for other targets.

Electrochemiluminescence behaviour of silver/silver orthophosphate/graphene oxide quenched by Pd@Au core-shell nanoflowers for ultrasensitive detection of insulin.

Herein, a highly efficient electrochemiluminescence resonance energy transfer (ECL-RET) immunosensor was established for ultrasensitive insulin detection. Silver/silver orthophosphate/graphene oxide composites (Ag/Ag3PO4/GO) were prepared as sensing platform for capture-antibody (Ab1) incubation. Ag3PO4 is a novel ECL donor whose emission could be remarkably enhanced by the synergetic assistance of GO with Ag NPs. Notably, GO presented excellent electrical conductivity and ultrahigh specific surface area to improve the loading capacity Ab1 and Ag3PO4, and Ag NPs with fine biocompatibility and catalytic property could immobilize Ab1 via Ag-N bond and further hasten the electron transfer to catalyze the generation of SO4•- radicals for boosting the ECL emission of donor. To establish a new ECL-RET system, Pd@Au core-shell nanoflower was prepared as a suitable ECL acceptor which could immobilize the detection-antibody (Ab2). Due to the fine spectral overlap, Pd@Au nanoflower could significantly quench the ECL emission of Ag3PO4, causing distinct decreases in ECL intensity. The proposed ECL-RET immunosensor exhibited sensitive response to insulin in a linear range of 0.0001-80 ng/mL with a low detection limit of 0.02 pg/mL (S/N = 3), it not only provides a reliable tool for insulin detection in diagnostics of diabetes, but also lights up a new avenue for designing effective ECL-RET pairs in bioanalysis.

Construction of the FRET Pairs for the Visualization of Mitochondria Membrane Potential in Dual Emission Colors.

Mitochondria membrane potential (MMP) play significant roles during metabolism, signaling, and other important bioevents. Visualization of MMP levels is essential for many biological researches. However, fluorescent probes for monitoring MMP levels in dual emission colors are still deficient, which greatly limited the development of relative research areas. In this work, a pair of fluorescent probes have been designed and synthesized to monitor the MMP levels in dual emission colors based on Forster resonance energy transfer (FRET) mechanism. The FRET donor (FixD) is constructed by linking a benzyl chloride group to a fluorophore with bright-green emission. The FixD could target mitochondria and be immobilized in mitochondria by linking to the thiol group of mitochondrial proteins. The FRET acceptor (LA) is designed with green absorption and deep-red emission. In live cells with high MMP levels, FixD and LA both target mitochondria, and deep-red (DR) emission could be detected with the excitation of 405 nm. Particularly, the spectral shift of fluorescence upon the decrease of MMP is up to 110 nm, which is greatly favorable for the clear observation of MMP levels. With the decrease of MMP, LA would be released from mitochondria while FixD would still be immobilized in mitochondria, and decreased DR emission and increased green fluorescence could be detected due to the absence of FRET. In this manner, the MMP levels could be monitored in dual emission colors.

MnCO3 as a New Electrochemiluminescence Emitter for Ultrasensitive Bioanalysis of ß-Amyloid1-42 Oligomers Based on Site-Directed Immobilization of Antibody.

In this work, an electrochemiluminescence (ECL) immunosensor utilizing MnCO3 nanospheres as a novel ECL luminophor and the HWRGWVC (HC-7) heptapeptide as an efficient antibody capturer for site-directed immobilization with high affinity was proposed. MnCO3 nanospheres prepared by a homogeneous precipitation method exhibited high ECL efficiency, low toxicity, favorable biocompatibility, and excellent stability. After the functionalization of polydimethyldiallylammonium chloride (PDDA), the obtained MnCO3/PDDA could combine with gold nanoparticles (Au NPs) via electrostatic interaction (MnCO3/PDDA/Au). Besides, HC-7 as a small peptide ligand has demonstrated an ability to bind the Fc portion of an antibody with high affinity. Because the end of HC-7 is a cysteine, it can connect to MnCO3/PDDA/Au via a Au-S bond. Then, the antibody could be effectively captured by HC-7 through specific interaction with a better maintained activity than traditional coupling reaction. To verify the practicability of the constructed immunosensor, ß-amyloid1-42 oligomers (Aß) were employed as an analyte. On the basis of the above points, the immunosensor performed favorable ECL property to Aß concentrations in a wide linear range (0.1 pg/mL to 10 ng/mL) with a low detection limit (19.95 fg/mL). With excellent repeatability, selectivity, and stability, this method opened up a new avenue for realizing the ultrasensitive detection of Aß and other biomarkers in a real sample analysis.

Interface-Targeting Strategy Enables Two-Photon Fluorescent Lipid Droplet Probes for High-Fidelity Imaging of Turbid Tissues and Detecting Fatty Liver.

Lipid droplets (LDs) with unique interfacial architecture not only play crucial roles in protecting a cell from lipotoxicity and lipoapoptosis but also closely relate with many diseases such as fatty liver and diabetes. Thus, as one of the important applied biomaterials, fluorescent probes with ultrahigh selectivity for in situ and high-fidelity imaging of LDs in living cells and tissues are critical to elucidate relevant physiological and pathological events as well as detect related diseases. However, available probes only utilizing LDs' waterless neutral cores but ignoring the unique phospholipid monolayer interfaces exhibit low selectivity. They cannot differentiate neutral cores of LDs from intracellular other lipophilic microenvironments, which results in extensively cloud-like background noise and severely limited their bioapplications. Herein, to design LD probes with ultrahigh selectivity, the exceptional interfacial architecture of LDs is considered adequately and thus an interface-targeting strategy is proposed for the first time. According to the novel strategy, we have developed two amphipathic fluorescent probes (N-Cy and N-Py) by introducing different cations into a lipophilic fluorophore (nitrobenzoxadiazole (NBD)). Consequently, their cationic moiety precisely locates the interfaces through electrostatic interaction and simultaneously NBD entirely embeds into the waterless core via hydrophobic interaction. Thus, high-fidelity and background-free fluorescence imaging of LDs are expectably realized in living cells in situ. Moreover, LDs in turbid tissues like skeletal muscle slices have been clearly imaged (up to 82 µm depth) by a two-photon microscope. Importantly, using N-Cy, we not only intuitively monitored the variations of LDs in number, size, and morphology but also clearly revealed their abnormity in hepatic tissues resulting from fatty liver. Therefore, these unique probes provide excellent imaging tools for elucidating LD-related physiological and pathological processes and the interface-targeting strategy possesses universal significance for designing probes with ultrahigh selectivity.

Phospholipid-Biomimetic Fluorescent Mitochondrial Probe with Ultrahigh Selectivity Enables In Situ and High-Fidelity Tissue Imaging.

In situ and directly imaging mitochondria in tissues instead of isolated cells can offer more native and accurate information. Particularly, in the clinical diagnose of mitochondrial diseases such as mitochondrial myopathy, it is a routine examination item to directly observe mitochondrial morphology and number in muscle tissues from patients. However, it is still a challenging task because the selectivity of available probes is inadequate for exclusively tissue imaging. Inspired by the chemical structure of amphiphilic phospholipids in mitochondrial inner membrane, we synthesized a phospholipid-biomimetic amphiphilic fluorescent probe (Mito-MOI) by modifying a C18-alkyl chain to the lipophilic side of carbazole-indolenine cation. Thus, the phospholipid-like Mito-MOI locates at mitochondrial inner membrane through electrostatic interaction between its cation and inner membrane negative charge. Simultaneously, the C18-alkyl chain, as the second targeting group, is deeply embedded into the hydrophobic region of inner membrane through hydrophobic interaction. Therefore, the dual targeting groups (cation and C18-alkyl chain) actually endow Mito-MOI with ultrahigh selectivity. As expected, high-resolution microscopic photos showed that Mito-MOI indeed stained mitochondrial inner membrane. Moreover, in situ and high-fidelity tissue imaging has been achieved, and particularly, four kinds of mitochondria and their crystal-like structure in muscle tissues were visualized clearly. Finally, the dynamic process of mitochondrial fission in living cells has been shown. The strategy employing dual targeting groups should have reference value for designing fluorescent probes with ultrahigh selectivity to various intracellular membranous components.

Spatially Dependent Fluorescent Probe for Detecting Different Situations of Mitochondrial Membrane Potential Conveniently and Efficiently.

The feedback from mitochondrial membrane potential (MMP) in different situations (normal, decreasing, and vanishing) can reflect different cellular status, which can be applied in biomedical research and diagnosis of the related diseases. Thus, the efficient and convenient detection for MMP in different situations is particularly important, yet the operations of current fluorescent probes are complex. In order to address this concern, we presented herein a spatially dependent fluorescent probe composed of organic cationic salt. The experimental results from normal and immortalized cells showed that it could accumulate in mitochondria selectively when MMP was normal. Also, it would move into the nucleus from mitochondria gradually with the decrease of MMP, and finally it targeted the nucleus exclusively when MMP vanished. According to the cell morphology, there is a straightforward spatial boundary between the nucleus and cytoplasm where mitochondria locate; thus, the three situations of MMP can be point-to-point indicated just by fluorescence images of the probe: that all probes accumulate in mitochondria corresponds to normal MMP; that probes locate both in the mitochondria and nucleus corresponds to decreasing MMP; that probes only target the nucleus corresponds to vanishing MMP. It is worth noting that counterstaining results with S-11348 indicated that the spatially dependent probe could be applied to distinguishing dead from viable cells in the same cell population. Compared with the commercial Cellstain-Double staining kit containing calcein-AM and propidium iodide (PI), this probe can address this concern by itself and shorten the testing time, which brings enormous convenience for relevant researches.

A single fluorescent probe enables clearly discriminating and simultaneously imaging liquid-ordered and liquid-disordered microdomains in plasma membrane of living cells.

Liquid-ordered (Lo) and liquid-disordered (Ld) microdomains in plasma membrane play different yet essential roles in various bioactivities. However, discrimination of the two microdomains in living cells is difficult, due to the similarity in their constituents and structures. Up to now, polarity sensitive probes are the only tool for imaging the two microdomains, but their small difference between emission spectra in the two microdomains (less than 50 nm) limited their application in living cells. In this work, we first presented an aggregation/monomer type of fluorescent probe (2,7-9E-BHVC12) with much larger separation in emission wavelength (up to 100 nm), for dual-color visualizing the two membrane microdomains in living cells. The probe can form red-emissive aggregates and yellow-emissive monomers when induced by Lo and Ld microdomains, respectively, and thus enables clear visualization of the two membrane microdomains in living cells with dual colors, and thus high-fidelity images of substructures of plasma membrane have been obtained. According to the images of three kinds of normal cells and three kinds of cancer cells stained with 2,7-9E-BHVC12, significant difference in plasma membrane microstructure of cancer cells was found. In terms of 2,7-9E-BHVC12, normal cells were mainly consisted of either Lo or Ld microdomains all over their membranes, while cancer cells all clearly display coexistence of Lo and Ld membrane microdomains. Therefore, 2,7-9E-BHVC12 can serve as a powerful tool for studies of membrane microdomains, and the different results of normal and cancer cells would also deepen our understanding in cancer science.