Utilizing a microfluidic device to enrich and fluorescently detect circulating tumor cells

Circulating tumor cells(CTCs)are cancer cells that have propagated from primary tumor sites,spreading into the bloodstream as the cellular origin of fatal metastasis,and to secondary tumor sites.Capturing and analyzing CTCs is a kind of‘‘liquid biopsy'of the tumor that provides information about cancer changes over time and tailoring treatment[1].CTC enrichment and detection remains technologically challenging due to their extremely low concentra-

A simple fluorescence anisotropy assay for detection of bisphenol A using fluorescently labeled aptamer

Bisphenol A(BPA)is one of the environmental endocrine disruptors(EDCs),and BPA contamination in environment can cause high risks to human health.Rapid determination of BPA on sites is in high demand in environmental analysis.Taking advantage of aptamers as affinity ligands and fluorescence anisotropy(FA)analysis,we developed a simple and rapid FA assay for BPA by employing a single tetramethylrhodamine(TMR)labeled short 35-mer DNA aptamer against BPA.The assay is based on the BPA-binding induced conformation change of TMR-labeled aptamer and alteration of interaction between TMR and guanine bases,resulting in change of FA signals.We screened the FA change of aptamer probes having TMR label on a specific site of the aptamer upon BPA addition.The aptamer with a TMR label on the 22nd T base showed large FA-decreasing response to BPA and maintained good binding affinity to BPA.By using this TMR-labeled aptamer,we achieved FA detection of BPA with a detection limit of 0.5μmol/L under the optimized conditions.This assay was selective towards BPA and enabled the detection of BPA spiked in tap water sample,showing the potential applications on water samples.

Hypoxia-activatable nano-prodrug for fluorescently tracking drug release in mice

Chemotherapy is one of the commonly used methods to treat various types of cancers in clinic by virtue of its high efficiency and universality. However, strong side effects and low concentration of conventional drugs at the tumor site have always been important factors that plague the chemotherapy effects of patients, further precluding their practical applications. Thereof, to solve the above dilemma, by integration of anticancer drug(nitrogen mustard, NM) into an NIR fluorophore(a dicyanoisophorone derivative), an intelligent prodrug NIR-NM was developed via molecular engineering strategy. Prodrug NIR-NM stimulated in hypoxia condition exhibits significantly higher toxicity to cancer cells than normal cells, essentially reducing the collateral damage to healthy cells and tissues of nitrogen mustard. More importantly, the nanoparticle prodrug FA-lip@NIR-NM showed the advantages of the high accumulation of drug at tumor site and long-circulation capacity in vivo, which endowed it the ability to track the release of the active chemotherapeutic drug and further treat solid tumors.

NIR-II fluorescence imaging in liver tumor surgery: A narrative review

In liver tumor surgery,the recognition of tumor margin and radical resection of microcancer focis have always been the crucial points to reduce postoperative recurrence of tumor.However,naked-eye inspection and palpation have limited effectiveness in identifying tumor boundaries,and traditional imaging techniques cannot consistently locate tumors in real time.As an intraoperative real-time navigation imaging method,NIRfluorescence imaging has been extensively studied for its simplicity,reliable safety,and superior sensitivity,and is expected to improve the accuracy of liver tumor surgery.In recent years,the research focus of NIRfluorescence has gradually shifted from the-rst near-infrared window(NIR-I,700–900 nm)to the second near-infrared window(NIR-II,1000–1700 nm).Fluorescence imaging in NIR-II reduces the scattering effect of deep tissue,providing a preferable detection depth and spatial resolution while signi-cantly eliminating liver autofluorescence background to clarify tumor margin.Developingfluorophores combined with tumor antibodies will further improve the precision offluorescence-guided surgical navigation.With the development of a bunch offluorophores with phototherapy ability,NIR-II can integrate tumor detection and treatment to explore a new therapeutic strategy for liver cancer.Here,we review the recent progress of NIR-IIfluorescence technology in liver tumor surgery and discuss its challenges and potential development direction.

Visualizing Wallerian degeneration in the corticospinal tract after sensorimotor cortex ischemia in mice

Stroke can cause Wallerian degeneration in regions outside of the brain,particularly in the corticospinal tract.To investigate the fate of major glial cells and axons within affected areas of the corticospinal tract following stroke,we induced photochemical infarction of the sensorimotor cortex leading to Wallerian degeneration along the full extent of the corticospinal tract.We first used a routine,sensitive marker of axonal injury,amyloid precursor protein,to examine Wallerian degeneration of the corticospinal tract.An antibody to amyloid precursor protein mapped exclusively to proximal axonal segments within the ischemic cortex,with no positive signal in distal parts of the corticospinal tract,at all time points.To improve visualization of Wallerian degeneration,we next utilized an orthograde virus that expresses green fluorescent protein to label the corticospinal tract and then quantitatively evaluated green fluorescent protein-expressing axons.Using this approach,we found that axonal degeneration began on day 3 post-stroke and was almost complete by 7 days after stroke.In addition,microglia mobilized and activated early,from day 7 after stroke,but did not maintain a phagocytic state over time.Meanwhile,astrocytes showed relatively delayed mobilization and a moderate response to Wallerian degeneration.Moreover,no anterograde degeneration of spinal anterior horn cells was observed in response to Wallerian degeneration of the corticospinal tract.In conclusion,our data provide evidence for dynamic,pathogenic spatiotemporal changes in major cellular components of the corticospinal tract during Wallerian degeneration.

Highly sensitive ratiometric fluorescent fiber matrices for oxygen sensing with micrometer spatial resolution

Oxygen(O_(2))-sensing matrices are promising tools for the live monitoring of extracellular O_(2) consumption levels in long-term cell cultures.In this study,ratiometric O_(2)-sensing membranes were prepared by electrospinning,an easy,low-cost,scalable,and robust method for fabricating nanofibers.Poly(ε-caprolactone)and poly(dimethyl)siloxane polymers were blended with tris(4,7-diphenyl-1,10-phenanthroline)ruthenium(II)dichloride,which was used as the O_(2)-sensing probe,and rhodamine B isothiocyanate,which was used as the reference dye.The functionalized scaffolds were morphologically characterized by scanning electron microscopy,and their physicochemical profiles were obtained by Fourier transform infrared spectroscopy,thermogravimetric analysis,and water contact angle measurement.The sensing capabilities were investigated by confocal laser scanning microscopy,performing photobleaching,reversibility,and calibration curve studies toward different dissolved O_(2)(DO)concentrations.Electrospun sensing nanofibers showed a high response to changes in DO concentrations in the physiological-pathological range from 0.5%to 20%and good stability under ratiometric imaging.In addition,the sensing systems were highly biocompatible for cell growth promoting adhesiveness and growth of three cancer cell lines,namely metastatic melanoma cell line SK-MEL2,breast cancer cell line MCF-7,and pancreatic ductal adenocarcinoma cell line Panc-1,thus recreating a suitable biological environment in vitro.These O_(2)-sensing biomaterials can potentially measure alterations in cell metabolism caused by changes in ambient O_(2)content during drug testing/validation and tissue regeneration processes.

Myelin histology:a key tool in nervous system research

The myelin sheath is a lipoprotein-rich,multilayered structure capable of increasing conduction velocity in central and peripheral myelinated nerve fibers.Due to the complex structure and composition of myelin,various histological techniques have been developed over the centuries to evaluate myelin under normal,pathological or experimental conditions.Today,methods to assess myelin integrity or content are key tools in both clinical diagnosis and neuroscience research.In this review,we provide an updated summary of the composition and structure of the myelin sheath and discuss some histological procedures,from tissue fixation and processing techniques to the most used and practical myelin histological staining methods.Considering the lipoprotein nature of myelin,the main features and technical details of the different available methods that can be used to evaluate the lipid or protein components of myelin are described,as well as the precise ultrastructural techniques.

Exploring non-curative endoscopic submucosal dissection:Current treatment optimization and future indication expansion

The increasing popularity of endoscopic submucosal dissection(ESD)as a treatment for early gastric cancer has highlighted the importance of quality assessment in achieving curative resections.This article emphasizes the significance of evaluating ESD quality,not only for curative cases but also for non-curative ones.Postoperative assessment relies on the endoscopic curability(eCura)classification,but management strategies for eCuraC-1 tumour with a positive horizontal margin are unclear.Current research primarily focuses on comparing additional surgical procedures in high-risk patients,while studies specifically targeting eCuraC-1 patients are limited.Exploring management strategies and follow-up outcomes for such cases could provide valuable insights.Furthermore,the application of molecular imaging using near-infrared fluorescent tracers holds promise for precise tumour diagnosis and navigation,potentially impacting the management of early-stage gastric cancer patients.Advancing research in these areas is essential for improving the overall efficacy of endoscopic techniques and refining treatment indications.

Simple substituents make big differences in photophysical performances of 2,1,3-benzothiadizole-conjugated spiropyrans

Developing multifunctional spiropyran dyes is of particular importance in diverse applications.In the present study,we synthesized two 2,1,3-benzothiadiazole-conjugated spiropyrans(BT-SP-NO2and BTSP-NMe2)with distinct substituents.These donor-acceptor-structured spiropyrans exhibited typical twisted intramolecular charge transfer features and strong emissions in low-polarity solvents with fluorescence quantum yields(QYs)of up to 90.7%.Like traditional spiropyrans,the electron-acceptor-substituted BT-SP-NO2exhibited excellent photochromic behavior under multiple alternating UV—Vis irradiation,while the electron-donor-substituted BT-SP-NMe2was an acidochromic dye.In addition,the substituent groups distinctly affected the packing modes of these spiropyrans in the solid state.BTSP-NMe2showed a much stronger solid-state emission(QY of 59.0%)than BT-SP-NO2.Moreover,these two dyes were utilized as biocompatible probes for the specific light-up imaging of lipid droplets.

Activatable fluorescent probes for imaging and diagnosis of rheumatoid arthritis

Rheumatoid arthritis(RA)is a systemic autoimmune disease that is primarily manifested as synovitis and polyarticular opacity and typically leads to serious joint damage and irreversible disability,thus adversely affecting locomotion ability and life quality.Consequently,good prognosis heavily relies on the early diagnosis and effective therapeutic monitoring of RA.Activatable fluorescent probes play vital roles in the detection and imaging of biomarkers for disease diagnosis and in vivo imaging.Herein,we review the fluorescent probes developed for the detection and imaging of RA biomarkers,namely reactive oxygen/nitrogen species(hypochlorous acid,peroxynitrite,hydroxyl radical,nitroxyl),pH,and cysteine,and address the related challenges and prospects to inspire the design of novel fluorescent probes and the improvement of their performance in RA studies.

A novel fluorescence sensor for milk clotting enzyme chymosin using peptide as substrate and covalent organic framework nanosheet as fluorescence quencher

Chymosin is one of the critical enzymes in cheese making.Herein,we proposed a novel fluorometric assay for chymosin determination.Firstly,covalent organic frameworks(COF)were synthesized and exfoliated to 2-dimensional COF nanosheets(COF NS)by ultrasound treatment.Gold nanoparticles(Au NPs)were loaded with COF NS to prepare AuNPs/COF NS(Au@COF NS).Secondly,rhodamine B(RhB)modified substrate peptide(Pep)for chymosin was linked with Au@COF NS to construct a Pep-Au@COF NS nanocomposite.For the sensing principle,fluorescence of RhB was quenched by Au@COF NS and the fluorescence intensity was weak due to the fluorescence resonance energy transfer between COF NS and RhB of Pep.However,in the presence of chymosin,the RhB was released by specific cleavage of the substrate peptide by chymosin and resulted in the recovery of fluorescence.The increased fluorescence intensity was proportional to the increase of chymosin concentration and thus a“turn on”fluorescent sensor for chymosin was constructed.The sensor showed a linear range in the concentration of 0.05-60.00μg/mL for the detection of chymosin with a detection limit of 20 ng/mL.The sensor was used to quantify chymosin in rennet product with good selectivity,which has the potential applications in cheese manufacturing.

Self-confocal NIR-II fluorescence microscopy for multifunctional in vivo imaging

Fluorescence imaging in the second near-infrared window(NIR-II,900–1880 nm)with less scattering background in biological tissues has been combined with the confocal microscopic system for achieving deep in vivo imaging with high spatial resolution.However,the traditional NIR-IIfluorescence confocal microscope with separate excitation focus and detection pinhole makes it possess low confocal e±ciency,as well as di±cultly to adjust.Two types of upgraded NIR-IIfluorescence confocal microscopes,sharing the same pinhole by excitation and emission focus,leading to higher confocal e±ciency,are built in this work.One type is-ber-pinhole-based confocal microscope applicable to CW laser excitation.It is constructed forfluorescence intensity imaging with large depth,high stabilization and low cost,which could replace multiphotonfluorescence microscopy in some applications(e.g.,cerebrovascular and hepatocellular imaging).The other type is air-pinhole-based confocal microscope applicable to femtosecond(fs)laser excitation.It can be employed not only for NIR-IIfluorescence intensity imaging,but also for multi-channelfluorescence lifetime imaging to recognize different structures with similarfluorescence spectrum.Moreover,it can be facilely combined with multiphotonfluorescence microscopy.A single fs pulsed laser is utilized to achieve up-conversion(visible multiphotonfluorescence)and down-conversion(NIR-II one-photonfluorescence)excitation simultaneously,extending imaging spectral channels,and thus facilitates multi-structure and multi-functional observation.

Effect of dissolved organic nitrogen on the bloom of Prorocentrum donghaiense and Karenia spp. in the East China Sea coastal waters

Understan ding the mechanism of harmful algal bloom formation is vital for effectively preventing algal bloom outbreaks in coastal environments.Karenia spp.blooms in the East China Sea show a significant correlation with nutrient regimes.However,the impact of key components of nutrients,especially dissolved organic nitrogen(DON),on the blooms of Karenia spp.is not clear.Quantitative research is still lacking.In this study,the cruise observations,field mesocosm-flask culture experiments,and a multinitrogen-tri-phytoplankton-detritus model(NTPD) are combined to reveal the quantitative influence of nutrient regimes on the shift of Prorocentrum donghaiense and Karenia spp.in the East China Sea.It has a synchronism rhythm of diatom-P.donghaienseKarenia spp.-diatom loop in the field culture experiment,which is consistent with the results of the cruise observation.The results showed that the processes of terrigenous DON(TeDON) and dissolved inorganic nitrogen(DIN:NO_(3)^(-)-N,NH_(4)^(+)-N) absorption promoted P.donghaiense to become the dominant algae in the community;whereas the processes of DON from P.donghaiense absorption promoted Karenia spp.to become the dominant algae in ambient DIN exhaustion.In addition,the three-dimensional fluorescence components of humus C,tyrosine and fulvic acid can indicate the processes of growth and extinction of P.donghaiense and Karenia spp.,respectively.This study infers that P.donghaiense and Karenia spp.regime shift mechanism associated with the nutrient regime in coastal waters,which provides a scientific basis for the environmental management of coastal eco system health.

Design and synthesis of two coordination polymers for the rapid detection of ciprofloxacin based on triphenylpolycarboxylic acid ligands

Two coordination polymers were synthesized by hydrothermal reaction,namely,[Cd(H_(3)cpbda)(2,2′‑bipy)(H_(2)O)]_(n)(1)and[Mn(H_(3)cpbda)(phen)(H_(2)O)]_(n)(2),where H_(5)cpbda=5,5′‑[(5‑carboxy‑1,3‑phenyl)bis(oxy)]triisophthalic acid,2,2′‑bipy=2,2′‑bipyridine,phen=1,10‑phenanthroline.The two complexes were characterized by single‑crystal X‑ray diffraction,powder diffraction,infrared spectroscopy,and thermogravimetric analysis.Complexes 1 and 2 are“V”‑shaped 1D chains,and the molecules form 2D(1)and 3D framework(2)structures through weakπ…πstacking.Furthermore,complex 1 was dispersed in an aqueous solution and its fluorescence intensity demonstrated excellent stability.Complex 1 can specifically detect ciprofloxacin in urine with a detection limit of 1.91×10^(-8)mol·L^(-1).CCDC:2359498,1;2359499,2.

In situ visualization of the cellular uptake and sub-cellular distribution of mussel oligosaccharides

Unlike chemosynthetic drugs designed for specific molecular and disease targets,active small-molecule natural products typically have a wide range of bioactivities and multiple targets,necessitating extensive screening and development.To address this issue,we propose a strategy for the direct in situ microdynamic examination of potential drug candidates to rapidly identify their effects and mechanisms of action.As a proof-of-concept,we investigated the behavior of mussel oligosaccharide(MOS-1)by tracking the subcellular dynamics of fluorescently labeled MOS-1 in cultured cells.We recorded the entire dynamic process of the localization of fluorescein isothiocyanate(FITC)-MOS-1 to the lysosomes and visualized the distribution of the drug within the cell.Remarkably,lysosomes containing FITC-MOS-1 actively recruited lipid droplets,leading to fusion events and increased cellular lipid consumption.These drug behaviors confirmed MOS-1 is a candidate for the treatment of lipid-related diseases.Furthermore,in a high-fat HepG2 cell model and in high-fat diet-fed apolipoprotein E(ApoE)^(-/-)mice,MOS-1 significantly promoted triglyceride degradation,reduced lipid droplet accumulation,lowered serum triglyceride levels,and mitigated liver damage and steatosis.Overall,our work supports the prioritization of in situ visual monitoring of drug location and distribution in subcellular compartments during the drug development phase,as this methodology contributes to the rapid identification of drug indications.Collectively,this methodology is significant for the screening and development of selective small-molecule drugs,and is expected to expedite the identification of candidate molecules with medicinal effects.

Isomeric fluorescence sensors for wide range detection of ionizing radiations

In order to achieve a wider range of ionizing radiations detection,novel fluorescence sensing materials have been developed that utilize the fluorescence enhancement phenomenon caused by the intramolecular photoinduced electron transfer(PET)effect.Two perylene diimide isomers PDI-P and PDI-B were designed and synthesized,and their molecular structures were characterized by high-resolution Fourier transform mass spectrometry(HRMS),nuclear magnetic resonance hydrogen and carbon spectroscopy(~1H and~(13)C NMR).The interaction between ionizing radiation and fluorescent molecules was simulated by HCl titration.The results show that combining PDIs and HCl can improve fluorescence through the retro-PET process.Despite the similarities in chemical structures,the fluorescent enhancement multiple of PDI-B with aromatic amine as electron donor is much higher than that of PDI-P with alkyl amine.In the direct irradiation experiments of ionizing radiation,the emission enhancement multiples of PDI-P and PDI-B are 2.01 and 45.4,respectively.Furthermore,density functional theory(DFT)and time-dependent density functional theory(TDDFT)calculations indicate that the HOMO and HOMO-1 energy ranges of PDI-P and PDI-B are 0.54 e V and 1.13 e V,respectively.A wider energy range has a stronger driving force on electrons,which is conducive to fluorescence quenching.Both femtosecond transient absorption spectroscopy(fs-TAS)and transient fluorescence spectroscopy(TFS)tests show that PDI-B has shorter charge separation lifetime and higher electron transfer rate constant.Although both isomers can significantly reduce LOD during PET process,PDI-B with aromatic amine has a wider detection range of 0.118—240 Gy due to its larger emission enhancement,which is a leap of three orders of magnitude.It breaks through the detection range of gamma radiation reported in existing studies,and provides theoretical support for the further study of sensitive and effective new materials for ionizing radiation detection.

Straw mulching alters the composition and loss of dissolved organic matter in farmland surface runoff by inhibiting the fragmentation of soil small macroaggregates

Straw mulching is a widespread practice for reducing the soil carbon loss caused by erosion.However,the effects of straw mulching on dissolved organic matter(DOM)runoff loss from black soil are not well studied.How straw mulching affects the composition and loss of runoff DOM by changing soil aggregates remains largely unclear.Here,a straw mulching treatment was compared to a no mulching treatment(as a control)on sloping farmland with black soil erosion in Northeast China.We divided the soil into large macroaggregates(>2 mm),small macroaggregates(0.25-2 mm),and microaggregates(<0.25 mm).After five rain events,the effects of straw mulching on the concentration(characterized by dissolved organic carbon(DoC)and composition(analyzed by fluorescence spectroscopy)of runoff and soil aggregate DOM were studied.The results showed that straw mulching reduced the runoff amount by 54.7%.Therefore,although straw mulching increased the average DOc concentration in runoff,it reduced the total runoff DOM loss by 48.3%.The composition of runoff DOM is similar to that of soil,as both contain humic-like acid and protein-like components.With straw mulching treatment,the protein-like components in small macroaggregates accumulated and the protein-like components in runoff declined with rain events.Fluorescence spectroscopy technology may help in understanding the hydrological paths of rain events by capturing the dynamic changes of runoff and soil DOM characteristics.A variation partitioning analysis(VPA)indicated that the DOM concentration and composition of microaggregates explained 68.2%of the change in runoff DOM from no mulching plots,while the change in runoff DOM from straw mulching plots was dominated by small macroaggregates at a rate of 55.1%.Taken together,our results demonstrated that straw mulching reduces the fragmentation of small macroaggregates and the loss of microaggregates,thus effecting DOM compositions in soil and reducing the DOM loss in runoff.These results provide a theoretical basis for reducing carbon loss in sloping farmland.

Enhancing the Photosensitivity of Hypocrellin A by Perylene Diimide Metallacage‑Based Host–Guest Complexation for Photodynamic Therapy

The development of supramolecular hosts which can efficiently encapsulate photosensitizers to improve the photodynamic efficacy holds great promise for cancer therapy.Here,we report two perylene diimide-based metallacages that can form stable host–guest complexes with planar conjugated molecules including polycyclic aromatic hydrocarbons and photosensitizers(hypocrellin A).Such host–guest complexation not only prevents the aggregation of photosensitizers in aqueous environments,but also offers fluorescence resonance energy transfer(FRET)from the metallacage to the photosensitizers to further improve the singlet oxygen generation(Φ_(Δ)=0.66).The complexes are further assembled with amphiphilic polymers,forming nanoparticles with improved stability for anticancer study.Both in vitro and in vivo studies indicate that the nanoparticles display excellent anticancer activities upon light irradiation,showing great potential for cancer photodynamic therapy.This study provides a straightforward and effective approach for enhancing the photosensitivity of conventional photosensitizers via host–guest complexation-based FRET,which will open a new avenue for host–guest chemistry-based supramolecular theranostics.

Fluorescent Double Network Hydrogels with Ionic Responsiveness and High Mechanical Properties for Visual Detection

We developed a fluorescent double network hydrogel with ionic responsiveness and high mechanical properties for visual detection.The nanocomposite hydrogel of laponite and polyacrylamide serves as the first network,while the ionic cross-linked hydrogel of terbium ions and sodium alginate serves as the second network.The double-network structure,the introduction of nanoparticles and the reversible ionic crosslinked interactions confer high mechanical properties to the hydrogel.Terbium ions are not only used as the ionic cross-linked points,but also used as green emitters to endow hydrogels with fluorescent properties.On the basis of the “antenna effect” of terbium ions and the ion exchange interaction,the fluorescence of the hydrogels can make selective responses to various ions(such as organic acid radical ions,transition metal ions) in aqueous solutions,which enables a convenient strategy for visual detection toward ions.Consequently,the fluorescent double network hydrogel fabricated in this study is promising for use in the field of visual sensor detection.

Effects of Fluorescent Pair on the Kinetics of DNA Strand Displacement Reaction

Fluorescent labels are widely used in the characterizations of DNA-based reaction network operations.We systematically studied the effects of commonly used fluorescent pairs on thermal stabilities of signal-substrate duplex and the strand displacement kinetics.It is demonstrated that the modifications of duplex with fluorescent pairs stabilize DNA duplex by up to 3.5℃,and the kinetics of DNA strand displacement circuit is also evidently slowed down.These results highlight the importance of fluorescent pairs towards the kinetic modulation in designing nucleic acid probes and complex DNA dynamic circuits.