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.

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.

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°C,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.

Temperature-controlled DCP Fluorescent Probe Based on Hydrogel-immobilized Quantum Dots Composite

A composite was created by incorporating the quantum dot-enhanced SiO_(2)nanoparticles within this hydrogel.Based on this composite,a temperature-controlled fluorescent probe for DCP was developed.A meticulous examination of this probe revealed its attributes and factors affecting its performance.By using temperature modulation,the probe was adept at detecting DCP concentrations ranging between 1.0×10^(-6)and 9.0×10^(-6)mol/L.Such a probe offers remarkable selectivity,repeatability,and robust stability,so that the detection of DCP can be carried out at different temperatures,and a fast,reliable,sensitive and low-cost intelligent detection method is realized.

Monitoring Xylem Transport in the Stem of Lilium lancifolium Using Fluorescent Dye 5(6)-Carboxyfluorescein Diacetate

The xylem undergoes physiological changes in response to various environmental conditions during the process of plant growth.To understand these physiological changes,it is extremely important to observe the transport of xylem.In this study,the distribution and structure of vascular bundle in Lilium lancifolium were observed using the method of semithin section.Methods for introducing a fluorescent tracer into the xylem of the stems were evaluated.Then,the transport rule of 5(6)-Carboxyfluorescein diacetate(CFDA)in the xylem of the stem of L.lancifolium was studied by fluorescence dye in live cells tracer technology.The results showed that the vascular bundles of L.lancifolium were scattered in the basic tissue,the peripheral vascular bundles were smaller and densely distributed,and the closer to the center,the larger the volume of vascular bundles and the more sparsely distributed.The vascular bundles of L.lancifolium are limited external tenacity vascular bundles,which are composed of phloem and xylem.The most suitable method for CFDA labeling the xylem of isolated stem segments of L.lancifolium was solution soaking for 24 h.The running speed of CF in the isolated stem was 0.3 cm/h,which was consistent with the running speed of the material in the field.CF could be transported between the xylem and parenchyma cells,indicating that the material transport in the xylem could be through the symplastic pathway.The above results laid a foundation for the study of the xylem transport mechanism and the xylem pathogen disease of lily.

3,6-Bis-β-Dicarbonylsubstituted Carbazoles Bearing N-Spacers and Their Eu(III) Complexes as Immunofluorescent Labelling Agents

New reagents for immunofluorescence analysis of carbazole series containing fluorinated β-dicarbonyl fragments and carboxylic substituent groups separated by spacers of different lengths from the light-gathering carbazole scaffold have been developed. The markers in complex with Eu3+ ions possess stability in the aqueous phase, intense and prolonged luminescence (τ 550 - 570 μs) with characteristic emission maxima in the region of 615 nm and excitation wavelengths in the region of 380 - 390 nm, which distinguishes them from most of the analogs used. In the study of marker conjugation with streptavidin, a reagent containing 4 - 5 europium labeling complexes based on spacer-containing carbazole tetraketone was obtained. The marker-doped silicate nanoparticles exhibit intense and long-lived luminescence in the characteristic region.

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.

Metal-organic cage as fluorescent probe for LiPF_(6)in lithium batteries

Lithium hexafluorophosphate(LiPF_(6)),the most commonly used lithium battery electrolyte salt,is vulnerable to heat and humidity.Quantitative and qualitative determination the variation of LiPF_(6)have always relied on advanced equipment.Herein,we develop a fast,convenient,high-selective fluorescence detection method based on metal-organic cages(MOC),whose emission is enhanced by nearly 20 times in the presence of LiPF_(6)with good stability and photobleaching resistance.The fluorescent probe can also detect moisture in battery electrolyte.We propose and verify that the luminescence enhancement is due to the presence of hydrogen bond-induced enhanced emission effect in cages.Fluorescent excitation-emission matrix spectra and variable-temperature nuclear magnetic resonance spectroscopy are employed to clarify the role of hydrogen bonds in guest-loaded cages.Density functional theory(DFT)calculation is applied to simulate the structure of host-guest complexes and estimate the adsorption energy involved in the system.The precisely matched lock-and-key model paves a new way for designing and fabricating novel host structures,enabling specific recognition of other target compounds.

A highly sensitive fluorescent probe RN-NA reveals peroxynitrite as a novel biomarker for primary open angle glaucoma

AIM:To directly quantify peroxynitrite(ONOO-)using a highly sensitive fluorescence resonance energy transfer probe RN-NA,investigate the association between ONOOand primary open angle glaucoma(POAG),and clarify whether RN-NA could be used as a potential tool for POAG diagnosis.METHODS:Plasma and aqueous humor(AH)samples were collected from POAG patients(n=100,age:59.70±6.87y)and age-related cataract(ARC)patients(n=100,age:61.15±4.60y)admitted to our hospital.Next,RN-NA was used to detect ONOO-in plasma and AH samples,and the relationship between ONOO-level and POAG was analyzed using binary logistic regression.Besides,Pearson correlation analysis was applied to characterize the correlation of the levels of ONOO-with the patients’age,intraocular pressure(IOP),and mean deviation of visual field testing.The ONOO-scavenger MnTMPyP was employed to treat the 3-morpholinosyndnomine(SIN-1)-induced ocular hypertension in mice(n=7,6-8wk).Finally,the IOP and ONOO-in both eyes were measured 30min after the last drug treatment.RESULTS:ONOO-levels of AH and plasma were significantly higher in the POAG group than in the ARC group(P<0.01).Additionally,ONOO-levels were closely correlated with POAG in a binary logistic regression analysis[odds ratio(OR)=1.008,95%confidence interval(CI):1.002-1.013,P<0.01 for AH;OR=1.004,95%CI:1.002-1.006,P<0.001 for plasma].Pearson correlation analysis showed that ONOO-levels in AH or plasma were positively associated with visual field defects(R=0.51,P<0.01 for AH;R=0.45,P<0.001 for plasma),and ONOO-levels in plasma and AH were correlated in the POAG group(R=0.69,P<0.001).However,administering MnTMPyP to mouse eyes reversed the elevated IOP caused by SIN-1(P<0.05).CONCLUSION:ONOO-levels in AH and plasma,detected by RN-NA,are significantly related to POAG and positively correlated with visual field defects in POAG patients.Hence,ONOO-is a potential biomarker of POAG,especially advanced POAG.Besides,anti-nitration compounds may be novel ocular hypotensive agents based on the animal study.

A ratiometric fluorescent probe for hypoxanthine detection in aquatic products based on the enzyme mimics and fluorescence of cobalt-doped carbon nitride

A ratiometric fluorescent probe for hypoxanthine(Hx)detection was established based on the mimic enzyme and fluorescence characteristics of cobalt-doped graphite-phase carbon nitride(Co doped g-C_(3)N_(4)).In addition to emitting strong fluorescence,the peroxidase activity of Co doped g-C_(3)N_(4)can catalyze the reaction of O-phenylenediamine and H_(2)O_(2)to produce diallyl phthalate which can emit yellow fluorescence at 570 nm.Through the decomposition of Hx by xanthine oxidase,Hx can be indirectly detected by the generating hydrogen peroxide based on the measurement of fluorescent ratio I(F_(570)/F_(370)).The linear range was 1.7-272.2 mg/kg(R^(2)=0.997),and the detection limit was 1.52 mg/kg(3σ/K,n=9).The established method was applied to Hx detection in bass,grass carp,and shrimp,and the data were verified by HPLC.The result shows that the established probe is sensitive,accurate,and reliable,and can be used for Hx detection in aquatic products.

Fluorescent probes for imaging and detection of plant hormones and their receptors

Exploring plant behavior at the cellular scale in a minimally invasive manner is critical to understanding plant adaptation to the environment.Phytohormones play vital regulatory roles in multiple aspects of plant growth and development and acclimation to environmental changes.Since the biosynthesis,modification,transportation,and degradation of plant hormones in plants change with time and space,their content level and distribution are highly dynamic.To monitor the production,transport,perception,and distribution of phytohormones within undamaged tissues,we require qualitative and quantitative tools endowed with remarkably high temporal and spatial resolution.Fluorescent probes are regarded as excellent tools for widespread plant imaging because of their high sensitivity and selectivity,reproducibility,real-time in situ detection,and uncomplicated mechanism elucidation.In this review,we provide a systematical overview of the progress in the sensing and imaging of phytohormone fluorescent probes and fluorescently labeled phytohormones to their receptors in plants.Moreover,forthcoming viewpoints and possible applications of these fluorescent probes within the realm of plants are also presented.We hold the conviction that the new perspective brought by this paper can promote the development of fluorescent probes,enabling them to have better detection performance in plant hormone imaging.

Manipulable multipurpose nanothermometers based on a fluorescent hybrid glass fiber microsphere cavity

Fluorescent nanothermometers for remote temperature measurement at the micro/nanoscale have stimulated growing efforts in developing efficient temperature-responsive materials and detection procedures.However,the efficient collection and transmission of optical signals have been a tremendous challenge for practical applications of these nanothermometers.Herein,we design an all-fiberized thermometry based on a fiber-coupled microsphere cavity coated with thermo-sensitive NaYF_(4)∶20%Yb^(3+);2%Er^(3+)@NaYF_(4)nanocrystals(NCs),allowing for spatial temperature sensing with resolution down to the few-micrometer scale.In our design,the microsphere efficiently excites the NCs and collects their upconversion emissions,and the use of a fiber splitter coupled with the microsphere allows for lossless routing of excitation and emitted light.We demonstrate the use of this all-fiber temperature sensor in diverse environments,especially in strongly acidic and alkaline conditions.Leveraging the high flexibility of commercial silica fiber,this all-fiber temperature ensor was employed for stable fixed-point real-time temperature measurement and multipurpose temperature recording/mapping in opaque environments,microscale areas,various solutions,and complicated bent structures.Thus,the demonstrated design could have strong implications for the practical use of nanothermometers in various possible scenarios,especially monitoring temperatures in diverse physiological settings.

Fluorescent Features of Germline Cysts of Fruit Fly (Drosophila melanogaster) after Fixation and Staining

[Objective] In order to provide firsthand information for the development of fluorescent histology,the paper studied the fluorescent features of ovarioles and germline cysts of fruit fly after common fixation and staining. [Method] With ovaries of Drosophila melanogaster as materials,after fixation with Bouin＇s fluid,the ovaries were sectioned and stained with HE,haematoxylin or eosin,respectively. The specimens were observed and photographed under fluorescent microscope in bright field,and fluorescent fields after excitation with green,blue and UV light,respectively. [Result] After staining by three methods,germ cells and somatic cells emitted different colors of fluorescence after excitation by different lights; lipids,nucleic acids and proteins in cells could also emit their special fluorescence. [Conclusion] Conventional dyes could give different fluorescence characteristics to germ cells and somatic cells,which can also give special fluorescence characteristics to different cellular components. Thus,the fluorescence histology will provide broad prospect for more convenient study on different cell types and cellular components.

Life prediction for vacuum fluorescent display using maximum likelihood estimation

In order to obtain the life information of the vacuum fluorescent display （VFD） in a short time, a model of constant stress accelerated life tests （CSALT） is established with its filament temperature increased, and four constant stress tests are conducted. The Weibull function is applied to describe the life distribution of the VFD, and the maximum likelihood estimation （MLE） and its iterative flow chart are used to calculate the shape parameters and the scale parameters. Furthermore, the accelerated life equation is determined by the least square method, the Kolmogorov-Smirnov test is performed to verify whether the VFD life meets the Weibull distribution or not, and selfdeveloped software is employed to predict the average life and the reliable life. Statistical data analysis results demonstrate that the test plans are feasible and versatile, that the VFD life follows the Weibull distribution, and that the VFD accelerated model satisfies the linear Arrhenius equation. The proposed method and the estimated life information of the VFD can provide some significant guideline to its manufacturers and customers.

Real-time fluorescent quantitative PCR非特异性扩增的研究进展

Real-time fluorescent quantitative PCR(RT-qPCR)因其高效快速、操作简便、高度敏感等优点获得广泛应用,但因其强大的放大功能而容易出现非特异性扩增的问题,尤其是荧光染料法中更加明显。文章对RT-qPCR的原理、非特异性扩增的发生因素、解决方案及该技术的应用前景进行了综述。

Production of Transgenic Pig Clone Embryo Expressed the Red Fluorescent Protein by Using the Somatic Cell Nucleus Transplantation Technology

[Objective] The research aimed to lay the foundation for producing the transgenic clone pig.[Method] The pig fetus fibroblast with the red fluorescent protein（RFP）gene that was transfected by the retrovirus was as the donor of nucleus transplantation.By using the somatic cell cloning technology,the development situation in vitro of clone embryo with RFP was studied.[Result] The fusion rate of RFP transgenic cell was 83.87% which had no significant difference with 80.56% of non-transgenic cell（P0.05）.The blastula rate in vitro of RFP transgenic somatic cell reconstructed embryo was 8.67% which had no significant difference with 6.56% of non-transgenic cell（P0.05）.After the reconstructed embryo of RFP transgenic somatic cell was transplanted into fifteen receptors,there was no conception individual.[Conclusion] The transgenic cell with the red fluorescent protein as the donor could successfully clone the transgenic embryo and obtain the transgenic blastula.

Application of Real-time Fluorescent Quantitative PCR in Plant

Real-time fluorescent quantitative PCR （RQ-PCR） is a detection method by adding fluorescent dye or fluorescent probe into the PCR reaction system, using fluorescent signal accumulation to monitor amplification reactions of PCR reaction process, and finally the unknown template can be quantitatively analyzed through the standard curve. So the detection level of PCR has improved from the qualitative to the quantitative. In order to provide a theoretical reference for further application, the principle, classification, advantages and disadvantages of RQ-PCR were intro- duced, and its application and progress in plants in recent years were reviewed.

Comparison of Multiplex Fluorescent PCR with Serum Type-specific Antibody Detection in Diagnosis of Genital Herpes

Objectives: To compare multiplex fluorescent PCRwith serum type-specific antibody detection in thediagnosis of herpes simplex virus (HSV) infection andto evaluate its significance in the diagnosis of genitalherpes.Methods: We detected HSV infection in 121 speci-mens collected from patients with genital herpesusing both multiplex fluorescent PCR and serum type-specific antibody detection. HSV viral isolation wasused as the standard control.Results: When compared with the viral isolation, thesensitivity and specificity for multiplex fluorescentPCR were 100% and 88.89%, respectively afterdiscrepant analysis. The sensitivity and specificity fortype-specific antibody detection was 77.68 % and77.78 %, respectively. However, the type-specificantibody detected HSV in two asymptomatic patientswhile the multiplex fluorescent PCR couldn’t detectany HSV DNA from those specimens.Conclusions: Multiplex fluorescent PCR is a verysensitive and specific method for detection and typingof HSV in the lesion of genital herpes, it failed todetect HSV DNA from the asymptomatic patients.Serum type-specific antibody detection was a lesssensitive and specific test but could detect the specificantibody from some asymptomatic patients. Thecombination of these two techniques would allow rapid,sensitive and accurate detection and typing of HSVand help clinical diagnosis and epidemiologic survey-ing of genital herpes.

Detecting H2S Gas Concentration by 1,8-Naphthalimides Fluorescent Probe

针对传统硫化氢检测方法灵敏度低的问题,以1,8-萘酰亚胺为荧光基团,基于H_2S的还原性,通过在荧光分子结构上引入具有氧化性的硝基,合成一种可与硫化氢气体发生氧化还原反应生成有荧光响应的小分子荧光探针。该探针本身荧光十分微弱,且荧光峰值在λ=467nm和λ=522nm处。与H_2S反应之后,522nm处的荧光效应消失,467nm处的荧光效应显著增强。测定小分子荧光探针在通入H_2S气体前后的荧光光谱,分析467nm处的荧光强度与气体浓度关系。结果表明:荧光光谱法检测出的H_2S气体浓度与荧光强度之间存在很强的线性关系,相关系数为0.979 3,最低可检测极限可达0.88×10^(-6 )mol·L^(-1)量级。表明基于1,8-萘酰亚胺衍生物的荧光光谱检测法可为油气田H_2S气体浓度的的快速测定提供参考。

Affinity and fluorescent detection of surfactants/ssDNA and single-walled carbon nanotube

A new biosensor platform was explored for detection of surfactant based on fluorescence changes from single strand DNA （ssDNA） and single-walled carbon nanotubes （SWNTs）. Thermodynamics assay was performed to value the stability of probe. The affinities of SWNT to five common surfactants （SDS, DBS, Triton X-100, Tween-20 and Tween-80） were investigated by real-time fluorescence method. The effects of Mg^2＋ and pH on the fluorescence intensity of self-assembled quenched sensor were performed. The fluorescent emission spectra were used to measure the responses of self-assembled quenched fluorescent of ssDNA/SWNTs to different concentration surfactant（Triton X-100）. The FAM-DNA wrapped SWNTs probe was stable in a wide temperature range （5 ℃ to 80℃）. The binding strength of surfactants and single-stranded DNA （ssDNA） on SWNTs surfaces was shown as follows： Triton X-100〉DBS〉Tween-20〉Tween-80〉ssDNA〉SDS, and the optimized reaction conditions included pH 7.4 and 10 mmol/L Mg2＋. The fluorescence of FAM-ssDNA wrapped SWNTs was proportionally recovered as a result of adding different concentrations of Triton X- 100, which realizes the quantitative detection of Triton X- 100.