Analyzing the structure and function of the brain's neural network is critical for identifying the working principles of the brain and the mechanisms of brain diseases.Recombinant rabies viral vectors allow for th...Analyzing the structure and function of the brain's neural network is critical for identifying the working principles of the brain and the mechanisms of brain diseases.Recombinant rabies viral vectors allow for the retrograde labeling of projection neurons and cell type-specific trans-monosynaptic tracing,making these vectors powerful candidates for the dissection of synaptic inputs.Although several attenuated rabies viral vectors have been developed,their application in studies of functional networks is hindered by the long preparation cycle and low yield of these vectors.To overcome these limitations,we developed an improved production system for the rapid rescue and preparation of a high-titer CVS-N2c-ΔG virus.Our results showed that the new CVS-N2c-ΔG-based toolkit performed remarkably:(1)N2cG-coated CVS-N2c-ΔG allowed for efficient retrograde access to projection neurons that were unaddressed by rAAV9-Retro,and the efficiency was six times higher than that of rAAV9-Retro;(2)the trans-monosynaptic efficiency of oG-mediated CVS-N2c-ΔG was 2–3 times higher than that of oG-mediated SAD-B19-ΔG;(3)CVS-N2c-ΔG could delivery modified genes for neural activity monitoring,and the time window during which this was maintained was 3 weeks;and(4)CVS-N2c-ΔG could express sufficient recombinases for efficient transgene recombination.These findings demonstrate that new CVS-N2c-ΔG-based toolkit may serve as a versatile tool for structural and functional studies of neural circuits.展开更多
The new double projecting neurons were found in the cat spinal dorsal horn by the double retrograde fluorescent tracing technique.Fast Blue(FB)was injected into unilateral dorsal column nucleus(DCN)of adult cats anest...The new double projecting neurons were found in the cat spinal dorsal horn by the double retrograde fluorescent tracing technique.Fast Blue(FB)was injected into unilateral dorsal column nucleus(DCN)of adult cats anesthetized with pentobarbital.Nuclear Yellow(NY)was injected ipsilaterally into the lateral cervical nucleus(LCN)8-9 days later.After an additional 18-30 hrs.展开更多
A new near-infrared fluorescent dye, 9-N-(2-hydroxyethyl)-N-methylamino-6-carbethoxy- 5 H-benzo[a] phenoxazin-5-one 1, was prepared from the reaction of N - (2 - hydroxyethyl)-N -methyl -4 -nitrosoaniline hydrochlorid...A new near-infrared fluorescent dye, 9-N-(2-hydroxyethyl)-N-methylamino-6-carbethoxy- 5 H-benzo[a] phenoxazin-5-one 1, was prepared from the reaction of N - (2 - hydroxyethyl)-N -methyl -4 -nitrosoaniline hydrochloride and ethyl 1, 3 - dihydroxynaphthoate. Five more fluorescent compounds were synthesized by the reaction of the resulting dye 1 with appropriate amino acid or carboxylic acids.展开更多
The purpose of this study was to investigate the effect of four fluorescent dyes, True Blue(TB), Fluoro-Gold(FG), Fluoro-Ruby(FR), and 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate(Di I), in re...The purpose of this study was to investigate the effect of four fluorescent dyes, True Blue(TB), Fluoro-Gold(FG), Fluoro-Ruby(FR), and 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate(Di I), in retrograde tracing of rat spinal motor neurons. We transected the muscle branch of the rat femoral nerve and applied each tracer to the proximal stump in single labeling experiments, or combinations of tracers(FG-Di I and TB-Di I) in double labeling experiments. In the single labeling experiments, significantly fewer labeled motor neurons were observed after FR labeling than after TB, FG, or Di I, 3 days after tracer application. By 1 week, there were no significant differences in the number of labeled neurons between the four groups. In the double-labeling experiment, the number of double-labeled neurons in the FG-Di I group was not significantly different from that in the TB-Di I group 1 week after tracer application. Our findings indicate that TB, FG, and Di I have similar labeling efficacies in the retrograde labeling of spinal motor neurons in the rat femoral nerve when used alone. Furthermore, combinations of Di I and TB or FG are similarly effective. Therefore, of the dyes studied, TB, FG and Di I, and combinations of Di I with TB or FG, are the most suitable for retrograde labeling studies of motor neurons in the rat femoral nerve.展开更多
In order to solve the problems of low accuracy and poor variety adaptability of the current measurement method of the permeability and coating property of sizing paste in textile industry, taking starch and polyvinyl ...In order to solve the problems of low accuracy and poor variety adaptability of the current measurement method of the permeability and coating property of sizing paste in textile industry, taking starch and polyvinyl alcohol(PVA) as the representatives of the most commonly used textile sizes, various concentrations of fluorescent molecules fluorescein isothiocyanate(FITC) were used to label starch and PVA to prepare fluorescein(F)-starch and F-PVA fluorescent sizes with different degrees of labeling(DLs) for the first time, respectively. Then the starch and PVA derivatives were employed to size pure cotton warp yarns. After preparing the sections of the sized yarns, the permeability and coating percentage of starch and PVA paste to the yarns were calculated by using a fluorescence microscope and the Photoshop software, respectively. The results demonstrate that F-starch and F-PVA with appropriate DL of 0.791% and 0.161%, respectively, exhibit good fluorescence property and similar sizing performance to the sizing performance of unlabeled starch and PVA. It is considered that fluorescence labeling of sizing agents with fluorescein units can provide an innovative way for the accurate determination of the permeability and coating property of sizing paste to warp yarns.展开更多
Two fluorescent indocyanine dyes containing at least one p-carboxybenzyl group on the nitrogen atoms in the heterocyclic rings were designed and synthesized. Their absorption maxima were 549 nm and 551 nm in water res...Two fluorescent indocyanine dyes containing at least one p-carboxybenzyl group on the nitrogen atoms in the heterocyclic rings were designed and synthesized. Their absorption maxima were 549 nm and 551 nm in water respectively. They had good water solubility and photostability.展开更多
Objective The assay of DNA mismatch repair (MMR) activity can be used as a biomarker for environmental condition detection and human disease diagnosis.Radioactive 32P-endlabeled DNA containing mismatch is extensively ...Objective The assay of DNA mismatch repair (MMR) activity can be used as a biomarker for environmental condition detection and human disease diagnosis.Radioactive 32P-endlabeled DNA containing mismatch is extensively used as the substrate for MMR activity analyses.The aim of the present study is to develop a simple non-radioactive,but equally specific and sensitive method for the MMR activity assay.Methods A fluorescent label was chosen to replace the radioactive isotope label.Sensitive evaluation of the fluorescent label was carried out for the first time,and then the fluorescent label was compared with the isotope label in the MMR activity and DNA binding assays.Result LOD (limit of detection) of the fluorescent label was about 0.1 fmol and the relative signal strength displayed a pretty good linear relationship.Moreover,the fluorescent label method has equivalent sensitivity and performance as compared with the classical radioactive method in experiments.Conclusion In light of the sensitivity,reproducibility,safety,rapidity and long lifespan of the fluorescent label,this improved method can be applied to evaluation of biologic and toxic effects of environmental pollutants on man and other forms of life.展开更多
The green fluorescent protein (GFP) from jellyfish Aequorea victoria has unique superiority as a kind of biological label. GFP has been widely used in all fields of biology based on the recent studies on its structure...The green fluorescent protein (GFP) from jellyfish Aequorea victoria has unique superiority as a kind of biological label. GFP has been widely used in all fields of biology based on the recent studies on its structure, characteristic and mechanism of bioluninescence.展开更多
1) In order to achieve the visibility of the chitosan macromolecule for the UV optical system of the analytical ultracentrifuge on investigation of the molecular characteristics and polymers interactions, the labeling...1) In order to achieve the visibility of the chitosan macromolecule for the UV optical system of the analytical ultracentrifuge on investigation of the molecular characteristics and polymers interactions, the labeling of chitosan by a new fluorophore of fluorescein-5-isothiocyanat was carried out. 2) Samples of fluorescent chitosan with two different degrees of fluorophore substitution and various degrees of acetylation were obtained. 3) The labeled chitosans with the fluorescein-5-isothiocyanat allowed estimating the sedimentation coefficient and the molecular characteristic in the analytical ultracentrifuge. 4) The sensitivity of the UV-optical system of the analytical ultracentrifuge for the obtained fluorescent samples of chitosan relatively to the fixation of the meniscus and the influence of the wavelength and rotation speed were estimated.展开更多
A relatively sensitive, specific, and photostable method for the detection of cytokeratin of cancer cells via conjugation with cadmium telluride quantum dots(CdTe QDs) was described. Water soluble CdTe QDs were conjug...A relatively sensitive, specific, and photostable method for the detection of cytokeratin of cancer cells via conjugation with cadmium telluride quantum dots(CdTe QDs) was described. Water soluble CdTe QDs were conjugated to anti-pan-cytokeratin(CK) monoclonal antibody(MAb) through coupling reagent [1-ethyl-3-(3-dimethyla- mino propyl)carbodiimide, EDC] and the conjugates were purified by dialysis. The expression of pan CK protein in HepG2 cells was observed by immunocytochemistry and direct immunofluorescence via QDs-Ab conjugates respectively. Fluorescence intensity and photostability of QDs were compared with those of FITC(fluorescein isothiocyanate). The results show that the QDs-Ab conjugates recognized specifically pan CK protein in HepG2 cells. Compared with FITC, CdTe QDs had higher brightness and photostability without obvious photobleaching under continuous exciting light illumination for 30 min and after the placement at room temperature for 3 d. The results indicate that conjugates of CdTe quantum dot with anti-pan CK MAb can be used for labeling cancer cells derived from epithelial tissues, which provides the basis for the detection of circulating tumor cells(CTCs).展开更多
Fluoro-ruby was injected into the posterior funiculus of the spinal cord in the cervical(C5-T2) and lumbar(L3-6) segments of adult guinea pigs.The spinal cord was cut into serial frozen sections.The Fluoro-ruby labeli...Fluoro-ruby was injected into the posterior funiculus of the spinal cord in the cervical(C5-T2) and lumbar(L3-6) segments of adult guinea pigs.The spinal cord was cut into serial frozen sections.The Fluoro-ruby labeling was clearly delineated from the surrounding structure.The labeling traversed the cervical,thoracic and lumbar segments,and was located on the ventral portion of the posterior funiculus on the injected side,proximal to the intermediate zone of the dorsal gray matter.The fluorescence area narrowed rostro-caudally.The spinal cord,spinal cord gray matter and corticospinal tract were reconstructed using 3D-DOCTOR 4.0 software,resulting in a robust three-dimensional profile.Using functionality provided by the reconstruction software,free multi-angle observation and sectioning could be conducted on the spinal cord and corticospinal tract.Our experimental findings indicate that the Fluoro-ruby retrograde fluorescent tracing technique can accurately display the anatomical location of corticospinal tract in the guinea pig and that three-dimensional reconstruction software can be used to provide a three-dimensional image of the corticospinal tract.展开更多
The labelling and imaging of tumor cells were investigated via arginine-glycine-aspartic acidcysteine(RGDC) peptide-labelled quantum dots(QDs). The results show that RGDC modified QDs can label SMMC-7721 tumor cells a...The labelling and imaging of tumor cells were investigated via arginine-glycine-aspartic acidcysteine(RGDC) peptide-labelled quantum dots(QDs). The results show that RGDC modified QDs can label SMMC-7721 tumor cells and adhere to cellular membrane. In constrast, the unmodified QDs are mainly dispersed around the cell. We also found that the RGDC-QDs can penetrate into the cell at 2 h of incubation. After 6 h of incubation, RGDC-QDs can accumulate in a unique intracellular region.展开更多
基金supported by the National Natural Science Foundation of China,Nos.32100899(to KZL),31830035(to FQX),31771156(to FQX),21921004(to FQX)the National Science and Technology Innovation 2030,No.2021ZD0201003(to FQX)+2 种基金the Key-Area Research and Development Program of Guangdong Province,No.2018B030331001(to FQX)the Strategic Priority Research Program of the Chinese Academy of Sciences,No.XDB32030200(to FQX)the Shenzhen Key Laboratory of Viral Vectors for Biomedicine,No.ZDSYS20200811142401005(to FQX)。
文摘Analyzing the structure and function of the brain's neural network is critical for identifying the working principles of the brain and the mechanisms of brain diseases.Recombinant rabies viral vectors allow for the retrograde labeling of projection neurons and cell type-specific trans-monosynaptic tracing,making these vectors powerful candidates for the dissection of synaptic inputs.Although several attenuated rabies viral vectors have been developed,their application in studies of functional networks is hindered by the long preparation cycle and low yield of these vectors.To overcome these limitations,we developed an improved production system for the rapid rescue and preparation of a high-titer CVS-N2c-ΔG virus.Our results showed that the new CVS-N2c-ΔG-based toolkit performed remarkably:(1)N2cG-coated CVS-N2c-ΔG allowed for efficient retrograde access to projection neurons that were unaddressed by rAAV9-Retro,and the efficiency was six times higher than that of rAAV9-Retro;(2)the trans-monosynaptic efficiency of oG-mediated CVS-N2c-ΔG was 2–3 times higher than that of oG-mediated SAD-B19-ΔG;(3)CVS-N2c-ΔG could delivery modified genes for neural activity monitoring,and the time window during which this was maintained was 3 weeks;and(4)CVS-N2c-ΔG could express sufficient recombinases for efficient transgene recombination.These findings demonstrate that new CVS-N2c-ΔG-based toolkit may serve as a versatile tool for structural and functional studies of neural circuits.
文摘The new double projecting neurons were found in the cat spinal dorsal horn by the double retrograde fluorescent tracing technique.Fast Blue(FB)was injected into unilateral dorsal column nucleus(DCN)of adult cats anesthetized with pentobarbital.Nuclear Yellow(NY)was injected ipsilaterally into the lateral cervical nucleus(LCN)8-9 days later.After an additional 18-30 hrs.
文摘A new near-infrared fluorescent dye, 9-N-(2-hydroxyethyl)-N-methylamino-6-carbethoxy- 5 H-benzo[a] phenoxazin-5-one 1, was prepared from the reaction of N - (2 - hydroxyethyl)-N -methyl -4 -nitrosoaniline hydrochloride and ethyl 1, 3 - dihydroxynaphthoate. Five more fluorescent compounds were synthesized by the reaction of the resulting dye 1 with appropriate amino acid or carboxylic acids.
基金supported by a grants from the National Program on Key Basic Research Project(973 Program),No.2014CB542200the Innovative Research Team by the Ministry of Education,No.IRT1201+1 种基金the National Natural Science Foundation of China,No.31271284,31171150,81171146,30971526,31040043,31371210,81372044,31471144the Beijing Municipal Natural Science Foundation of China,No.7142164
文摘The purpose of this study was to investigate the effect of four fluorescent dyes, True Blue(TB), Fluoro-Gold(FG), Fluoro-Ruby(FR), and 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate(Di I), in retrograde tracing of rat spinal motor neurons. We transected the muscle branch of the rat femoral nerve and applied each tracer to the proximal stump in single labeling experiments, or combinations of tracers(FG-Di I and TB-Di I) in double labeling experiments. In the single labeling experiments, significantly fewer labeled motor neurons were observed after FR labeling than after TB, FG, or Di I, 3 days after tracer application. By 1 week, there were no significant differences in the number of labeled neurons between the four groups. In the double-labeling experiment, the number of double-labeled neurons in the FG-Di I group was not significantly different from that in the TB-Di I group 1 week after tracer application. Our findings indicate that TB, FG, and Di I have similar labeling efficacies in the retrograde labeling of spinal motor neurons in the rat femoral nerve when used alone. Furthermore, combinations of Di I and TB or FG are similarly effective. Therefore, of the dyes studied, TB, FG and Di I, and combinations of Di I with TB or FG, are the most suitable for retrograde labeling studies of motor neurons in the rat femoral nerve.
基金National Natural Science Foundation of China(Nos.51573095 and 51873187)Project of Key Laboratory of Clean Dyeing and Finishing Technology of Zhejiang Province,China(No.QJRZ1902)+1 种基金Technological Research Project for Public Welfare of Zhejiang Province,China(No.LGG21E030005)Postdoctoral Research Program of Zhejiang Province in 2021,China。
文摘In order to solve the problems of low accuracy and poor variety adaptability of the current measurement method of the permeability and coating property of sizing paste in textile industry, taking starch and polyvinyl alcohol(PVA) as the representatives of the most commonly used textile sizes, various concentrations of fluorescent molecules fluorescein isothiocyanate(FITC) were used to label starch and PVA to prepare fluorescein(F)-starch and F-PVA fluorescent sizes with different degrees of labeling(DLs) for the first time, respectively. Then the starch and PVA derivatives were employed to size pure cotton warp yarns. After preparing the sections of the sized yarns, the permeability and coating percentage of starch and PVA paste to the yarns were calculated by using a fluorescence microscope and the Photoshop software, respectively. The results demonstrate that F-starch and F-PVA with appropriate DL of 0.791% and 0.161%, respectively, exhibit good fluorescence property and similar sizing performance to the sizing performance of unlabeled starch and PVA. It is considered that fluorescence labeling of sizing agents with fluorescein units can provide an innovative way for the accurate determination of the permeability and coating property of sizing paste to warp yarns.
基金the National Natural Science Foundation of China and The Ministry of Education of China for providing financial support for this project.
文摘Two fluorescent indocyanine dyes containing at least one p-carboxybenzyl group on the nitrogen atoms in the heterocyclic rings were designed and synthesized. Their absorption maxima were 549 nm and 551 nm in water respectively. They had good water solubility and photostability.
基金grants from the Chinese National Natural Sciences Foundation(30821006,50973046)the Doctoral Station Science Foundation from the Chinese Ministry of Education(200802840023)the Jiangsu Provincial Natural Science Foundation (BK2010046,BK2008138,BK2008072,BY2009147)
文摘Objective The assay of DNA mismatch repair (MMR) activity can be used as a biomarker for environmental condition detection and human disease diagnosis.Radioactive 32P-endlabeled DNA containing mismatch is extensively used as the substrate for MMR activity analyses.The aim of the present study is to develop a simple non-radioactive,but equally specific and sensitive method for the MMR activity assay.Methods A fluorescent label was chosen to replace the radioactive isotope label.Sensitive evaluation of the fluorescent label was carried out for the first time,and then the fluorescent label was compared with the isotope label in the MMR activity and DNA binding assays.Result LOD (limit of detection) of the fluorescent label was about 0.1 fmol and the relative signal strength displayed a pretty good linear relationship.Moreover,the fluorescent label method has equivalent sensitivity and performance as compared with the classical radioactive method in experiments.Conclusion In light of the sensitivity,reproducibility,safety,rapidity and long lifespan of the fluorescent label,this improved method can be applied to evaluation of biologic and toxic effects of environmental pollutants on man and other forms of life.
文摘The green fluorescent protein (GFP) from jellyfish Aequorea victoria has unique superiority as a kind of biological label. GFP has been widely used in all fields of biology based on the recent studies on its structure, characteristic and mechanism of bioluninescence.
文摘1) In order to achieve the visibility of the chitosan macromolecule for the UV optical system of the analytical ultracentrifuge on investigation of the molecular characteristics and polymers interactions, the labeling of chitosan by a new fluorophore of fluorescein-5-isothiocyanat was carried out. 2) Samples of fluorescent chitosan with two different degrees of fluorophore substitution and various degrees of acetylation were obtained. 3) The labeled chitosans with the fluorescein-5-isothiocyanat allowed estimating the sedimentation coefficient and the molecular characteristic in the analytical ultracentrifuge. 4) The sensitivity of the UV-optical system of the analytical ultracentrifuge for the obtained fluorescent samples of chitosan relatively to the fixation of the meniscus and the influence of the wavelength and rotation speed were estimated.
基金Supported by the Major Project Fund of Jilin Provincial Science and Technology DepartmentChina(No.20082123)+1 种基金the Province University Union Fund of Jilin Province China(No.20082011)
文摘A relatively sensitive, specific, and photostable method for the detection of cytokeratin of cancer cells via conjugation with cadmium telluride quantum dots(CdTe QDs) was described. Water soluble CdTe QDs were conjugated to anti-pan-cytokeratin(CK) monoclonal antibody(MAb) through coupling reagent [1-ethyl-3-(3-dimethyla- mino propyl)carbodiimide, EDC] and the conjugates were purified by dialysis. The expression of pan CK protein in HepG2 cells was observed by immunocytochemistry and direct immunofluorescence via QDs-Ab conjugates respectively. Fluorescence intensity and photostability of QDs were compared with those of FITC(fluorescein isothiocyanate). The results show that the QDs-Ab conjugates recognized specifically pan CK protein in HepG2 cells. Compared with FITC, CdTe QDs had higher brightness and photostability without obvious photobleaching under continuous exciting light illumination for 30 min and after the placement at room temperature for 3 d. The results indicate that conjugates of CdTe quantum dot with anti-pan CK MAb can be used for labeling cancer cells derived from epithelial tissues, which provides the basis for the detection of circulating tumor cells(CTCs).
基金supported by a grant from the Priority Academic Program Development of Jiangsu Higher Education Institutions
文摘Fluoro-ruby was injected into the posterior funiculus of the spinal cord in the cervical(C5-T2) and lumbar(L3-6) segments of adult guinea pigs.The spinal cord was cut into serial frozen sections.The Fluoro-ruby labeling was clearly delineated from the surrounding structure.The labeling traversed the cervical,thoracic and lumbar segments,and was located on the ventral portion of the posterior funiculus on the injected side,proximal to the intermediate zone of the dorsal gray matter.The fluorescence area narrowed rostro-caudally.The spinal cord,spinal cord gray matter and corticospinal tract were reconstructed using 3D-DOCTOR 4.0 software,resulting in a robust three-dimensional profile.Using functionality provided by the reconstruction software,free multi-angle observation and sectioning could be conducted on the spinal cord and corticospinal tract.Our experimental findings indicate that the Fluoro-ruby retrograde fluorescent tracing technique can accurately display the anatomical location of corticospinal tract in the guinea pig and that three-dimensional reconstruction software can be used to provide a three-dimensional image of the corticospinal tract.
基金Supported by the National Natural Science Foundation of China(Nos.30970719, 21043002)the Social Development Project of Science and Technology Department of Jilin Province, China(Nos.20106031, 20090133)+1 种基金the Project of Science and Technology Department of Changchun City, China(No.09SF02)the Fundamental Research Funds of Jilin University, China (No.200903098)
文摘The labelling and imaging of tumor cells were investigated via arginine-glycine-aspartic acidcysteine(RGDC) peptide-labelled quantum dots(QDs). The results show that RGDC modified QDs can label SMMC-7721 tumor cells and adhere to cellular membrane. In constrast, the unmodified QDs are mainly dispersed around the cell. We also found that the RGDC-QDs can penetrate into the cell at 2 h of incubation. After 6 h of incubation, RGDC-QDs can accumulate in a unique intracellular region.
