Fluorescence Resonance Energy Transfer Research Articles

Detection of intracellular microRNA-21 for cancer diagnosis by a nanosystem containing a ZnO@polydopamine and DNAzyme probe.

MicroRNAs (miRNAs) are a series of single-stranded non-coding ribonucleic acid (RNA) molecules which associated closely with various human diseases. Efficient strategies for detecting miRNAs are of great significance to cancer diagnosis and prognosis. Here we provide a novel nanosystem that can be applied for the detection of miRNAs. The nanosystem consists of a single-stranded deoxyribonucleic acid (DNA) probe and a probe carrier. The DNA probe was designed based on a deoxyribozyme (DNAzyme) with several necessary functional sequences and two fluorescent dyes labeled at proper sites. The ZnO@polydopamine (ZnO@PDA) nanomaterial serves not only as a probe carrier, but also as a supplier of Zn2+ that can activate the DNAzyme. The DNA probe will undergo a conformation alteration induced by miRNA-21, which then trigger the DNAzyme catalyzed self-cleavage reaction with the assist of Zn2+ provided by ZnO decomposition under weak acid environment. A change of fluorescent color will occur due to the interruption of fluorescence resonance energy transfer between the two fluorescent dyes, and the dissociated miRNA-21 can repeatedly induce the above responses to amplify the fluorescence signal. The feasibility of the whole procedure was demonstrated by various experiments. This nanosystem showed a good selectivity towards miRNA-21, and under the optimal incubation time of 2 hours, a good linear relationship was obtained in a concentration range of 0.01-2.0 nM with a detection limit of 3.8 pM. In in vivo detection, an obvious fluorescence color change from red to green can be observed in the presence of miRNA-21. The results proved that this miRNA detection strategy has a broad application prospect in tumor diagnosis and miRNA related biological studies.

A point-of-care aptasensor based on the upconversion nanoparticles/MoS2 FRET system for the detection of Pseudomonas aeruginosa infection.

The rapid detection of Pseudomonas aeruginosa (P. aeruginosa) is of great significance for the diagnosis of medical infection. In view of the above, a novel aptasensor based on fluorescence resonance energy transfer (FRET) was developed. It contained aptamer-coupled upconversion nanoparticles (UCNPs-apt) as a donor (excitation 980nm) and molybdenum disulfide (MoS2) nanosheets as an acceptor. The upconversion fluorescence aptamer system was investigated to obtain the optimal parameters of MoS2 concentration, the incubation time of UCNPs-apt/MoS2 and P. aeruginosa, and pH. Based on the optimal parameters, a linear calibrationequation (emission 654nm) with a wide detection range 8.7 × 10 ~ 8.7 × 107cfu/mL, a high coefficient of determination R2 0.9941, and a low limit of determination (LOD) 15.5cfu/mL were established. The method was validated with P. aeruginosa infected foci of mouse wound. The advantage of this aptasensor isthat analysisresults can be obtained within 1.5h, which was much faster than that ofthe standard method (18-24h). Furthermore, combined with a portable instrument, it can be used as a point-of-care testing for the early detection of P. aeruginosa infection, which is useful for selecting the correct antibiotics to achieve good therapeutic effects. Additionally, it also has a broad application prospect in food and environmental areas.

A label-free ratiometric fluorescent aptasensor based on a peroxidase-mimetic multifunctional ZrFe-MOF for the determination of tetrodotoxin.

A Fe/Zr bimetal-organic framework (ZrFe-MOF) is utilized to establish a ratiometric fluorescent aptasensor for the determination of tetrodotoxin (TTX). The multifunctional ZrFe-MOF possesses inherent fluorescence at 445nm wavelength, peroxidase-mimetic activity, and specific recognition and adsorption capabilities for aptamers, owing to its organic ligand, and Fe and Zr nodes. The peroxidation of o-phenylenediamine (OPD) substrate generates fluorescent 2,3-diaminophenazine (OPDox) at 555nm wavelength, thus quenching the inherent fluorescence of ZrFe-MOF because of the fluorescence resonance energy transfer (FRET) effect. TTX aptamers, which are absorbed on the material surface without immobilization or fluorescent labeling, inhibit the peroxidase-mimetic activity of ZrFe-MOF. It causes the decreased OPDox fluorescence at 555nm wavelength and the inverse restoration of ZrFe-MOF fluorescence at 445nm wavelength. With TTX, the aptamers specifically bind to TTX, triggering rigid complex release from ZrFe-MOF surface and reactivating its peroxidase-mimetic activity. Consequently, the two fluorescence signals exhibit opposite changes. Employing this ratiometric strategy, the determination of TTX is achieved with a detection limit of 0.027ng/mL and a linear range of 0.05-500ng/mL. This aptasensor also successfully determines TTX concentrations in puffer fish and clam samples, demonstrating its promising application for monitoring trace TTX in food safety.

Overlapping characteristics of weak interactions of two transcriptional regulators with WDR5

The WD40 repeat protein 5 (WDR5) is a nuclear hub that critically influences gene expression by interacting with transcriptional regulators. Utilizing the WDR5 binding motif (WBM) site, WDR5 interacts with the myelocytomatosis (MYC), an oncoprotein transcription factor, and the retinoblastoma-binding protein 5 (RbBP5), a scaffolding element of an epigenetic complex. Given the clinical significance of these protein-protein interactions (PPIs), there is a pressing necessity for a quantitative assessment of these processes. Here, we use biolayer interferometry (BLI) to examine interactions of WDR5 with consensus peptide ligands of MYC and RbBP5. We found that both interactions exhibit relatively weak affinities arising from a fast dissociation process. Remarkably, live-cell imaging identified distinctive WDR5 localizations in the absence and presence of full-length binding partners. Although WDR5 tends to accumulate within nucleoli, WBM-mediated interactions with MYC and RbBP5 require their localization outside nucleoli. We utilize fluorescence resonance energy transfer (FRET) microscopy to confirm these weak interactions through a low FRET efficiency of the MYC-WDR5 and RbBP5-WDR5 complexes in living cells. In addition, we evaluate the impact of peptide and small-molecule inhibitors on these interactions. These outcomes form a fundamental basis for further developments to clarify the multitasking role of the WBM binding site of WDR5.

Enhanced detection of acrylamide using a versatile solid-state upconversion sensor through spectral and visual analysis

Acrylamide (AM) generally forms in high-temperature processes and has been classified as a potential carcinogen. In this study, we put forward a maneuverable solid-state luminescence sensor using polydimethylsiloxane (PDMS) as the matrix coupled with upconversion nanoparticles as the indicator. The core-shell upconversion nanoparticles emitting cyan light were uniformly encapsulated in PDMS. Then it was further modified with complementary DNA of AM aptamer. The nanocrystalline fluorescein isothiocyanate isomer (FITC), coupled with AM aptamer, was attached to the surface of PDMS. FITC effectively quenched the upconversion luminescence through fluorescence resonance energy transfer (FRET). The introduction of AM resulted in preferentially bound to aptamer caused the separation of the quencher and the donor, and led to luminescence recovery. The developed sensor was applied for both spectral and visual monitoring, demonstrating a detection limit (LOD) of 1.00 nM and 1.07 nM, respectively. Importantly, in the actual foodstuffs detection, there is no obvious difference between the results of this study and the standard method, which indicates the developed method has good accuracy. Therefore, this solid-state sensor has the potential for on-site detection using a smartphone device and an Android application.

Syringa reticulata potently inhibits the activity of SARS-CoV-2 3CL protease

The ongoing coronavirus infectious disease (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) still urgently requires effective treatments. The 3C-like (3CL) protease of SARS-CoV-2 is a highly conserved cysteine protease that plays an important role in the viral life cycle and host inflammation, providing an ideal target for developing broad-spectrum antiviral drugs. Herein, we describe the discovery of a large number of herbs mainly produced in Heilongjiang Province, China, that exhibited different inhibitory activities against SARS-CoV-2 3CL protease. We confirmed that Syringa reticulata, which is used for clinical treatment of chronic bronchitis and asthma, is a specific and potent inhibitor of 3CL protease. A 70 % ethanol extract of S. reticulata dose-dependently inhibited the cleavage activity of 3CL protease in a fluorescence resonance energy transfer assay with an IC50 value of 0.0018 mg/mL, but had minimal effect in pseudovirus-based cell entry and luciferase-based RNA-dependent RNA polymerase assays. These results suggest that S. reticulata will be a potential leading candidate for COVID-19 treatment.

Dynamically photo-tunable multicolor luminescence of supramolecular gel based on fluorescent gelator and used for information encryption

The ingenious collaboration of photo-isomerized molecules and luminescent gelator can create self-supporting materials used for multi-level and multi-dimensional information encryption. Herein, a supramolecular gel (SDTD) was prepared by using photoluminescent gelator 1,3:2,4-di(benzylidene)-D-sorbitol (DBS), lanthanide ions (Tb3+), as well as reversible photo-isomerized diarylethylene (DTE), and spiropyran (SP) derivative. Rheological studies showed that the prepared SDTD gel possessed good self-supporting ability owing to the 3D network structure formed by DBS aggregates. Additionally, SDTD gel displayed dynamic multicolor luminescence due to the presence of three luminescent centers including DBS, Tb3+, and MC-o (the photoisomer of SP). The luminescent color of SDTD gel can be easily tuned by irradiation wavelengths and time. It was found that a photo-tunable fluorescence resonance energy transfer (FRET) occurred between Tb3+ and MC-o, Tb3+ and DTE-c (the photoisomer of DTE), the corresponding FRET efficiency was 47 % and 52 %, respectively. The results also indicated that the photo-tunable inner filter effect (IFE) occurred between DBS and DTE-c, DBS and MC-o, respectively, which endowed SDTD gel with dynamically photo-tunable multicolor luminescence capability. SDTD gel has the advantages of simple preparation, low cost, and flexible luminescence regulation. In addition, based on the multicolor luminescence of SDTD gel, a new method of multi-dimensional information encryption was proposed, which shows a potential application in the field of anti-counterfeiting materials.

3D walkable DNA gears for ultrasensitive detection of multiple microRNAs in lung cancer cell lysates

As a tumor biomarker with therapeutic application potential, microRNA (miRNA) was crucial for the accurate and sensitive detection of early-stage tumors. Herein, a unique three dimensional (3D) DNA nanomachine (DNM) was created, which was capable detecting lung cancer-related biomarkers miRNA-21, miRNA-205 and miRNA-125b in lung cancer cell lysates with extreme sensitivity. The 3D DNM was composed of DNA scissors and three flexible walkable DNA gears modified with various species of silver nanoclusters (AgNCs). Based on the flexibility of DNA scissors and the walkability of DNA gears, neighboring DNA gears closed the distance between different species of AgNCs by walking in the presence of targets, generating fluorescence resonance energy transfer (FRET) effect and emitting different kinds of fluorescence to complete the highly sensitive detection of single targets and multiple targets. The findings demonstrated that a linear model provided an excellent match for the association between fluorescence signal and target miRNAs. For miRNA-21, miRNA-205, and miRNA-125b, the limits of detection (LODs) (signal/noise = 3) were 4.2 pmol/L (pM), 6.3 pM, and 10.2 pM, respectively. Their recoveries in A549 cell lysate samples ranged from 95.3 to 108.8 % with relative standard deviations of 1.26 %–4.88 %. Satisfactorily, the 3D DNM displayed exceptional analytical performance with high sensitivity and stability, strong specificity and reproducibility, which was triumphantly employed to identify miRNAs in tumor cell lysates, providing a workable technique in creating adaptable nanostructure for dependable bioanalysis and clinical diagnosis of cancer biomarkers.

Aptamer-based flares hybridized with single-stranded DNA-conjugated MoS2 nanosheets for ratiometric fluorescence sensing and imaging of potassium ions and adenosine triphosphate in human fluids and living cells

Addressing the limitations observed in previous studies, where the quantitative range of nanoprobes for detecting K+ and adenosine triphosphate (ATP) did not cover concentrations found within living cells, the present study aimed to develop ratiometric nanoprobes that can accurately sense changes in K+ and ATP levels in living cells and quantify them in human fluids. The proposed nanoprobes consisted of recognition flares modified with 6-carboxyfluorescein (FAM) and 5-carboxytetramethylrhodamine (TAMRA), along with thiolate single-stranded DNA (ssDNA) and molybdenum disulfide nanosheets (MoS2 NSs). The thiolate ssDNA acts as a linker between the flares and the MoS2 NSs, directly forming a functional nanostructure at room temperature. The direct conjugation of labeled flares to the MoS2 NSs simplifies the fabrication process. In the absence of K+ and ATP, the hybridization of flares and thiolate ssDNA caused FAM to move away from TAMRA, suppressing the fluorescence resonance energy transfer (FRET) process. However, upon the introduction of K+ and ATP, the flares undergo a structural transformation via the formation of G-quadruplex formation and the generation of hairpin-shaped structures, respectively. This structural change leads to the release of the flares from the ssDNA-conjugated nanosheet surface. The release of the flares brings FAM and TAMRA into close proximity, allowing FRET to occur, leading to FRET and static quenching. By monitoring the ratio between the fluorescence intensities of FAM and TAMRA, the concentration of K+ (5–100 mM) and ATP (0.3–5 mM) can be accurately determined by the proposed nanoprobes. The advantages of these nanoprobes lie in their ability to provide ratiometric measurements, which enhance the accuracy and reliability of the quantification process. The proposed nanoprobes offer potential applications as ratiometric imaging probes for monitoring K+ and ATP-related reactions in living cells, providing valuable insights into cellular processes. Additionally, they can be employed for determining the levels of K+ and ATP in human fluids, offering potential diagnostic applications in various clinical settings.

Applications of Optical Method As an Effective Platform for Progesterone Sensing Using Carbon Dots and Graphene Oxide

Today, ‘Hormonal imbalance’ is a significant concern worldwide that lead to various types of complications in human body. For instance, abnormal levels of steroid hormones, such as progesterone are associated with irregular or absent menstrual periods, reproductive disorders, infertility, cardiovascular diseases, and cancer. Nowadays, progesterone is being excessively consumed as oral contraceptives, menopausal hormone therapy and treating infertility, resulting into the deleterious effects due to its endocrine disrupting activity. Furthermore, its presence as a potent endocrine disruptor in surface or environmental waters and various daily use products is another concern. Therefore, monitoring progesterone hormone is vital in environmental samples because of the adverse effects on reproductive health and causing hormonal imbalance. Herein, we have developed optical photoluminescence (PL) method implementing carbon dots (CDs) and graphene oxide (GO) based fluorescence resonance energy transfer (FRET) mechanism for progesterone hormone sensing. In this method, firstly progesterone-specific antibodies are covalently functionalized to CDs whose fluorescence has been quenched (turn off) by adding GO. However, the presence of progesterone increases the distance between donor and acceptor moieties and turns-on the fluorescence by displacing GO off the surface of CDs which in turn substantially reduces the FRET efficiency. Various optimization steps have been performed under control experiments to tune the fluorescence of CDs and the concentration of graphene oxide for maximum efficiency. In conclusion, a linear relationship is achieved in between PL intensity and different progesterone concentrations (10-900 nM) with 13.8 nM detection limits (R2=0.974). The structural and morphological analysis (XPS, and FT-IR) revealed the structural and surface functionalities of CDs. Finally, the potential of the developed biosensor has been explored in water matrices.

Structural Dynamics Role of AGG Interruptions in Inhibition CGG Repeat Expansion Associated with Fragile X Syndrome.

Abnormal expansion of trinucleotide CGG repeats is responsible for Fragile X syndrome. AGG interruptions in CGG repeat tracts were found in most healthy individuals, suggesting a crucial role in preventing disease-prone repeat expansion. Previous biophysics studies emphasize a difference in the secondary structure affected by AGG interruptions. However, the mechanism of how AGG interruptions impede repeat expansion remains elusive. We utilized single-molecule fluorescence resonance energy transfer spectroscopy to investigate the structural dynamics of CGG repeats and their AGG-interrupted variants. Tandem CGG repeats fold into a stem-loop hairpin structure with the capability to undergo a conformational rearrangement to modulate the length of the overhang. However, this conformational rearrangement is much more retarded when two AGG interruptions are present. Considering the significance of hairpin slippage in repeat expansion, we present a molecular basis suggesting that the internal loop created by two AGG interruptions acts as a barrier, obstructing the hairpin slippage reconfiguration. This impediment potentially plays a crucial role in curbing abnormal expansion, thereby contributing to the genomic stability.

Novel fluorescence nano-orbital biosensor for highly sensitive microRNA detection

BackgroundMicroRNAs play an important role in regulating cell function and gene expression. Early prevention and clinical diagnosis of diseases have high requirements for high-sensitivity detection of microRNAs. Due to the limitations of tedious operation and large sample size, miRNA with small molecular weight and low expression abundance cannot be accurately detected in traditional miRNA detection. To improve the sensitivity and accuracy of detection, we established a novel biosensor based on nucleic acid circuit of signal amplification, which converted miRNA recognition into a fluorescence signal for amplification. ResultsWe designed a biosensor based on an exponential amplification reaction with cascaded HCR and DNAzyme nucleic acid circuit (named E-NOF biosensor) by amplicon sub-fragments to trigger the construction of fluorescence nano-orbitals (NOF), which could be used to detect miRNA ultrasensitively. By modifying two fluorophores (Cy3 and Cy5) on the chain of constructing nano-orbitals, when the amplicon triggered the construction of nano-orbitals, fluorescence resonance energy transfer (FRET) occurred between Cy3 and Cy5, and then two fluorescence signals with different trends could be observed. Therefore, through the ratio of the two signals, we could quantitatively and quickly detect the miRNA from 1 fM to 100 nM, and the E-NOF biosensor detection limit was as low as 0.129 fM. Furthermore, the HCR nucleic acid circuit cascaded with DNAzyme could enrich the fluorophores on the nano-orbitals and significantly enhance the fluorescence signal by accelerating the reaction rate. SignificanceAccording to our understanding, the E-NOF biosensor is the first strategy to cascade EXPAR with HCR and DNAzyme nucleic acid circuit for miRNA-1246 detection. Accurate results can be obtained in only 120 min. Compared with the traditional HCR system, the sensitivity of the new E-NOF biosensor is increased by 1 × 109 times. Furthermore, the biosensor can also detect biomarkers in human serum samples. It has great potential in miRNA detection and identification.

A bubble-enhanced lanthanide-doped up/down-conversion platform with tumor microenvironment response for dual-modal photoacoustic and near-infrared-II fluorescence imaging

As an important tumor diagnosis strategy in precision medicine, multimodal imaging has been widely studied. However, the weak imaging signal with low spatial resolution and the constant signal of lack of specific activation severely limit its disease diagnosis. Herein, a bubble-enhanced lanthanide-based up/down-conversion platform with tumor microenvironment response for dual-mode imaging, LDNP@DMSN-Au@CaCO3 nanoparticles (named as LDAC NPs) were successfully developed. Combining the advantages of photoacoustic imaging (PAI) and the second near-infrared window (NIR-II) fluorescence imaging (FI), significantly improved the accuracy of diseases diagnosis. LDAC NPs with flower-like structure were synthesized through the encapsulation of uniform lanthanide-doped nanoparticles (NaYbF4:Ce,Er@NaYF4 named LDNPs) with dendritic mesoporous silica (DMSN). The gold nanoparticles (Au NPs) were then in situ grown on the surface of DMSN and the surface were finally coated with a layer of calcium carbonate (CaCO3). Under the excitation of the 980 nm laser, LDNPs showed strong emission of NIR-II at 1550 nm due to the doping of Ce and Er ions, showcasing excellent spatial resolution and deep tissue penetration characteristics, while the resulting visible light emission (540 nm) enables Au NPs to generate PAI signals with the aid of LDNPs via the fluorescence resonance energy transfer effect. In acidic tumoral environment, CaCO3 layer could produce CO2 microbubbles, and the PAI signals of LDAC NPs could be further enhanced with the generation of CO2 bubbles due to the bubble cavitation effect. Simultaneously, the NIR-II FI of LDAC NPs was self-enhanced with the degradation of the CaCO3. This intelligent nanoparticle with stimulus-activated dual-mode imaging capability holds great promise in future precision diagnostics.

Rationally Designed Dual Base Pair Mismatch Enables Toehold-Mediated Strand Displacement to Efficiently Recognize Single-Nucleotide Polymorphism without Enzymes.

The efficiency of the enzyme-free toehold-mediated strand displacement (TMSD) technique is often insufficient to detect single-nucleotide polymorphism (SNP) that possesses only single base pair mismatch discrimination. Here, we report a novel dual base pair mismatch strategy enabling TMSD biosensing for SNP detection under enzyme-free conditions when coupled with catalytic hairpin assembly (CHA) and fluorescence resonance energy transfer (FRET). The strategy is based on a competitive strand displacement reaction mechanism, affected by the thermodynamic stability originating from rationally designed dual base pair mismatch, for the specific recognition of mutant-type DNA. In particular, enzyme-free nucleic acid circuits, such as CHA, emerge as a powerful method for signal amplification. Eventually, the signal transduction of this proposed biosensor was determined by FRET between streptavidin-coated 605 nm emission quantum dots (605QDs, donor) and Cy5/biotin hybridization (acceptor, from CHA) when incubated with each other. The proposed biosensor displayed high sensitivity to the mutant target (MT) with a detection concentration down to 4.3 fM and led to high discrimination factors for all types of mismatches in multiple sequence contexts. As such, the application of this proposed biosensor to investigate mechanisms of the competitive strand displacement reaction further illustrates the versatility of our dual base pair mismatch strategy, which can be utilized for the creation of a new class of biosensors.

Micellular fluorescence resonance energy transfer based fluorescent ratiometric response to hydrocarbon analytes.

Fluorescence resonance energy transfer (FRET) has been utilised to develop numerous selective and sensitive fluorescent ratiometric sensors. Typically, FRET-based fluorescent ratiometric sensors rely on chemical interactions between the sensor and analyte to illicit a response, thus unreactive hydrocarbons are a neglected analyte and a source for new sensors. By containing an unbound donor-acceptor system within micelles, energy transfer is enabled by spatial confinement. This offers the potential of a ratiometric response as a hydrocarbon analyte is added. Introducing a hydrocarbon analyte to this system causes micelles to swell, increasing the donor-acceptor distance and thus reducing the amount of observed energy transfer. We present InP/ZnS quantum dot donors interacting with a Nile Red acceptor, confined by cetyltrimethylammonium bromide (CTAB)-based micelles. We alleviated spatial confinement of the pair within micelles using common laboratory solvents to represent hydrocarbons, (toluene, hexane and octadecene). We constructed calibration curves for each solvent and found effective sensing ranges of 0.009-0.21, 0.008-0.27 and 0.003-0.06 M for toluene, hexane and octadecene, respectively. This study contributes towards the development of new hydrocarbon sensors utilising this new mechanism.

Pathogenic Variants in USH1G/SANS Alter Protein Interaction with Pre-RNA Processing Factors PRPF6 and PRPF31 of the Spliceosome.

Pre-mRNA splicing is an essential process orchestrated by the spliceosome, a dynamic complex assembled stepwise on pre-mRNA. We have previously identified that USH1G protein SANS regulates pre-mRNA splicing by mediating the intranuclear transfer of the spliceosomal U4/U6.U5 tri-snRNP complex. During this process, SANS interacts with the U4/U6 and U5 snRNP-specific proteins PRPF31 and PRPF6 and regulates splicing, which is disturbed by variants of USH1G/SANS causative for human Usher syndrome (USH), the most common form of hereditary deaf-blindness. Here, we aim to gain further insights into the molecular interaction of the splicing molecules PRPF31 and PRPF6 to the CENTn domain of SANS using fluorescence resonance energy transfer assays in cells and in silico deep learning-based protein structure predictions. This demonstrates that SANS directly binds via two distinct conserved regions of its CENTn to the two PRPFs. In addition, we provide evidence that these interactions occur sequentially and a conformational change of an intrinsically disordered region to a short α-helix of SANS CENTn2 is triggered by the binding of PRPF6. Furthermore, we find that pathogenic variants of USH1G/SANS perturb the binding of SANS to both PRPFs, implying a significance for the USH1G pathophysiology.

A novel self-assembled dual-emissive ratiometric fluorescent nanoprobe for alkaline phosphatase sensing

BackgroundAlkaline phosphatase (ALP) is widely found in various organs and tissues of the human body which could assist in the verification of the presence of various diseases through its content in the blood. In the past few years, many analytical methods for ALP activity assays have been explored. However, a simple and economical method with high sensitivity and specificity also remains great challenge. Therefore, the development of sensitive and efficient approach for ALP analysis is of great significance in biomedical studies. ResultsHerein, we constructed a highly sensitive and label-free ratiometric fluorometric biosensing platform for the determination of ALP activity, which utilizing lysozyme(Ly)-functionalized 5-methyl-2-thiouracil(MTU)-modified gold nanoclusters (MTU-Ly@Au NC) and poly-dopamine (PDA) as signal indicators. Dopamine (DA) can self-polymerizes to form PDA under alkaline conditions that can further quenched the fluorescence of MTU-Ly@Au NC at 525nm due to fluorescence resonance energy transfer (FRET) and absorption competition quenching (ACQ) effects. In this process, the PDA fluorescence intensity at 325nm was nearly unchanged. After the addition of ALP, ascorbic acid (AA) which can alleviate the self-polymerization process of DA was generated from the substrate ascorbic acid 2-phosphate (AAP), thus changing ratiometric fluorescence intensity of I525/I325. Hence, by monitoring the fluorescence ratio (I525/I325), a ratiometric fluorescence biosensing platform for ALP was established with the linear calibration in the range of 0.5–8 U L−1 and the limit of detection of 0.157 U L−1. SignificanceThis work not only synthesized a novel fluorescence probe with simple preparation and low cost for ALP which has excellent anti-interference properties and selectivity. Furthermore, this biosensing platform was successfully applied for the determination of ALP activity in human serum samples. This work provided a potential tool for biomedical diagnostics in the future.

A fluorescent sensor based on Zn(II)-coodination polymers bearing mixed ligands for the sensitive detection of riboflavine, imidacloprid and nitenpyram

Two new 2D Zn(II) coordination polymers (BAO28 and BAO29), formulated as [Zn(DIN)(HPT)]DMF·2H2O (BAO28) and [Zn(DIN)(HPT)]·2H2O (BAO29) (H2HPT = 5‑hydroxy-isophthalic acid, DIN = 1,4-di(imidazol-1-yl)naphthalene and DMF = N,N-dimethylformamide), were successfully synthesized under solvothermal conditions. Fluorescence analysis showed that BAO28 proceeded with a DIN ligand-based emissive fluorescence. More importantly, the recoverable multi-responsive fluorescent sensor BAO28 was used for fluorescent detection of imidacloprid (IMI), nitenpyram (NTP), and riboflavine (VB2) in a neutral buffer solution with high sensitivity and a low detection limit. Moreover, the sensing of the pesticides by BAO28 was mainly due to the competitive absorption of excitation and emission light by the sensor with that of the pesticides. Furthermore, for the sensing of riboflavine, in addition to the internal filtration effect (IFE) and fluorescence resonance energy transfer (FRET) mechanisms, the sensing mechanism was simultaneously attributed to photoinduced electron transfer.

First Full-Length Cloning of Human NaV1.7 in S. Cerevisiae-Based Novel D-Crypt Platform for its High-Scale Production

NaV1.7, a voltage-gated sodium channel, induces chronic pain. Current research on the discovery of inhibitors against NaV1.7 is advancing with the hope of treating chronic pain conditions in patients suffering from various diseases. However, to characterize NaV1.7 and inhibitor interactions, a higher yield of expression is required to obtain sufficient protein. Molecular cloning of hNaV1.7 was done using the homologous recombination method in our novel S. cerevisiae-based D-cryptÔ platform. The platform was designed for the high-yield production of ‘difficult-to-express’ proteins (DTE-Ps) up to 500 L. The changes in the cell membrane potential of hNaV1.7 were determined using a fluorescence resonance energy transfer (FRET) assay with tetracaine (hNaV1.7 inhibitor). Expression analysis of hNaV1.7 showed 2 bands of size 250 and 280 kDa that were absent in untransfected yeast cells. Immunofluorescence images revealed the presence of hNaV1.7 on the membrane of hNaV1.7-expressing yeast cells. Tetracaine exhibited concentration-dependent inhibition of the FRET ratio, with the order of potency (IC50 = 0.46 µM) being approximately the same as previously reported, suggesting the functionality of the channel protein expressed in the D-cryptÔ platform. Cloning of NaV1.7 in the D-cryptÔ platform will help to scale up the production of channel proteins, which will ultimately help in the structural and functional characterization of the binding interactions between toxins and the NaV1.7 channel to identify more specific NaV1.7 inhibitors. HIGHLIGHTS 7 is a voltage-gated sodium channel and induces chronic pain 7 inhibitors can advance the treatment of chronic pain associated with various diseases To characterize interaction of Nav1.7 with its inhibitors, higher yield of the NaV1.7 is required D-crypt is a platform designed for the high-yield production of ‘difficult-to-express’ proteins (DTE-Ps) up to 500 L Successful cloning of NaV1.7 in D-crypt platform suggest that the D-crypt platform is a suitable platform for expressing full-length functionally active hNaV1.7 channel at larger scale. It will further aid in the screening of NaV1.7 inhibitors GRAPHICAL ABSTRACT

High dispersibility ratiometric fluorescence sensor designed by functionalized mesoporous silica nanopraticles for sensing and imaging of hydrogen peroxide

Herein, a mesoporous silica nanoparticle (MSN) based ratiometric fluorescence nanosensor (MSN-BA-SiCD) with improved performance was designed for monitoring hydrogen peroxide (H2O2). Specifically, H2O2 response molecule (BA) with AIE feature was synthesized and encapsulated into MSN channel as the fluoresent recognition site. Afterwards, silane-modified carbon dot (SiCD) was covalently connected to the MSN surface which can effectively prevent the leakage of BA and improve the water dispersibility of the nanosensor. The presence of H2O2 could result in a “turn-on” fluorescence of BA as well as concurrent variation in the fluorescence resonance energy transfer (FRET) signal between SiCD and BA. MSN-BA-SiCD displays a specific blue-to-green resolved emission change in response to H2O2 and the detection limit can be as low as 4.78 × 10−6 M. In addition, the nanosensor exhibited enhanced photostability due to the protection of the material framework. Moreover, the nanosensing system was successfully applied to detect H2O2 in beer, orange juice and living cells. The excellent water dispersibility and photostability of the nanosensor, together with high specificity and biocompatibility, constituted an advanced set of characteristics among existing MSNs, which was expected to further promote the development of H2O2 sensors in the environmental and biological fields.