Adventitious agents,comprising unintentionally introduced microorganisms in the production of biological products,pose a significant challenge in ensuring the safety of gene therapy products.The revised International ...Adventitious agents,comprising unintentionally introduced microorganisms in the production of biological products,pose a significant challenge in ensuring the safety of gene therapy products.The revised International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use(ICH)guildline Q5A(R2)from September 2022 highlights the inclusion of viral vector-based gene therapy products in safety discussions,emphasizing controls in material sourcing,testing,and viral clearance[1].Detecting adventitious virus contamination is complex due to the unique characteristics of gene therapy products and the limitations of routine testing methods.The US Food and Drug Administration(FDA)recommends incorporating routine and specific virus detection methods,including those outlined in various pharmacopeias.Existing control methods have limitations,prompting the need for highly sensitive and broad-spectrum detection approaches.Unlike traditional biological products,gene therapy products primarily consist of live viruses,necessitating methods that distinguish between the main virus and adventitious viruses.Current virus detection techniques,such as polymerase chain reaction(PCR),sequencing,mass spectrometry,and DNA microarrays[2e4],have their drawbacks.展开更多
DNA repair enzymes are important in the repair of DNA lesions for maintaining the genome stability,and their abnormal expression induced various human cancers.Simultaneous detection of these DNA enzymes could provide ...DNA repair enzymes are important in the repair of DNA lesions for maintaining the genome stability,and their abnormal expression induced various human cancers.Simultaneous detection of these DNA enzymes could provide convincing evidence based on the comparison of the activity of multiple enzymes than on that of single enzyme.Although fluorescence approach has been applied for the simultaneous detection both of DNA repair enzymes,the spectral overlap and multiwavelength excitation severely restrict the number of available fluorophores.Thus,it is difficult to simultaneously detect three enzymes in a single analysis by fluorescence detection.Herein,we developed a method for the simultaneous determination of three DNA repair enzymes including human flap DNA endonuclease 1(FEN1),human alkyladenine DNA glycosylase(hAAG)and uracil DNA glycosylase(UDG)based on the combination of template-free amplification system with capillary electrophoresis-laser induced fluorescence(CE-LIF)detection.The amplification system was adopted to transfer and amplify the enzymatic products into different length DNA fragments which could be separated effectively by CE-LIF without the complicated modification of the capillary inner wall or labeling different tails on signal probes for separation.The method demonstrated a detection limit of 0.07 U/mL(0.08-160 U/mL)for FEN1,2.40 U/mL(2.5-250U/mL)for hAAG and 2.1×10^(-4)U/mL(0.0004-2.5 U/mL)for UDG,the relative standard deviations(RSDs)of peak time and peak area for different analytes were as follows:2.50%-4,37%and 3.24%-7.18%(inter-day);1.37%-2.71%and 1.43%-3.02%(intra-day),4.28%-6.08%and 4.16%-7.57%(column to column),respectively.And it can identify the inhibitor-like drugs,evaluate enzymatic kinetics and achieve the detection of three enzymes in cell extracts,providing a simple and powerful platform for simultaneous detection of more DNA repair enzymes.展开更多
Reactive dyes with different reactive groups exhibit different hydrolysis and dyeing behaviors.This is particularly evident in the combination dyeing process,where the competion between hydrolysis and dyeing reactions...Reactive dyes with different reactive groups exhibit different hydrolysis and dyeing behaviors.This is particularly evident in the combination dyeing process,where the competion between hydrolysis and dyeing reactions increases the complexity.Therefore,developing an effective method to monitor the changes in reactive dyes during the dyeing process is important.This study aims to develop a capillary electrophoresis(CE)technique combined with an ultraviolet(UV)detector(CE-UV)for detecting three reactive dyes and their six derivatives(a total of nine analytes).The optimized CE conditions are 20.0 mmol/L sodium tetraborate(Na_(2)B_(4)O_(7)·10H_(2)O),acetonitrile(ACN)with a volume fraction of 15.0%,20.0 mmol/L α-cyclodextrin(α-CD),and at a pH value of 9.0(adjusted with 0.5 mol/L H_(3)BO_(3)).The limit of detection(LOD)(a signal-to-noise ratio of 3)for the nine analytes ranges from 1.38 to 5.06 mg/L.The relative standard deviations(RSDs)for peak areas and migration time are 2.19%-4.96%and 0.29%-2.75%,respectively.The method is capable of accurately identifying three reactive dyes and their six derivatives and monitoring alterations in composition and dyeing behavior during single and combination dyeing processes.展开更多
Capillary electrophoresis(CE),recognized for its minimal reagent dosage,rapid analysis,and high efficiency,holds significant promise for the analysis of traditional Chinese medicines(TCM).This article reviews the appl...Capillary electrophoresis(CE),recognized for its minimal reagent dosage,rapid analysis,and high efficiency,holds significant promise for the analysis of traditional Chinese medicines(TCM).This article reviews the application of CE in the determination of active ingredients in TCM.The active substances of herbal medicines have been classified and discussed based on their chemical properties,and the CE methods applied to different substances are summarized and discussed.This article also provides some suggestions for future research and improvement.展开更多
[Objective] The like-rocket immunoelectrophoresis was used to explore a new feasible electrophoresis method for single cell gel electrophoresis assay （comet assay）.[Method] The like-rocket immunoelectrophoresis was ...[Objective] The like-rocket immunoelectrophoresis was used to explore a new feasible electrophoresis method for single cell gel electrophoresis assay （comet assay）.[Method] The like-rocket immunoelectrophoresis was used for single cell gel electrophoresis assay to detect DNA damage at single cell level,then it was compared with traditional electrophoresis method to analyze its advantage and disadvantages.[Result] Under cell DNA undamaged state,the results of two electrophoresis methods were consistent.When cell DNA was damaged,the comet tail divergence of some cells under traditional electrophoresis method were drifted,however,the comet tail image of like-rocket immunoelectrophoresis was concentrated and not shifted.[Conclusion] The like-rocket immunoelectrophoresis had some advantages.展开更多
A recombinant inbred population (RI) was constructed from a cross between B5, an introgression. line from the wild rice Oryza officinalis Wall. ex Watt, and susceptible cultivar Minghui 63 ( O. sativa L.). The brown p...A recombinant inbred population (RI) was constructed from a cross between B5, an introgression. line from the wild rice Oryza officinalis Wall. ex Watt, and susceptible cultivar Minghui 63 ( O. sativa L.). The brown planthopper ( BPH) resistances of RI lines were evaluated. Based on bulked segregant analysis (BSA), two protein bulks were made by extracting proteins from equally mixed seedlings of extremely resistant and susceptible plants selected from the RI population, respectively. Two-dimensional electrophoresis was used to detect the changes of polypeptide pattern. Results showed that a protein P40 ( pI 6.3, Mw 40 kD) was significantly reduced or vanished after BPH infestation for 48 h in the susceptible bulk, while it remained uninfluenced in the resistant bulk. In connection with the physiological changes of the resistant and susceptible lines subjected to BPH sucking, we suppose that the protein P40 is related to the interaction responses of lice plants to BPH infestation.展开更多
Polyacrylamide gel electrophoresis (PAGE) and biochemical staining method were used in this study for the analysis on malate dehydrogenase (MDH,E.C. 1.1.1.37) isozymes zymogram in 11 different types of tissues of male...Polyacrylamide gel electrophoresis (PAGE) and biochemical staining method were used in this study for the analysis on malate dehydrogenase (MDH,E.C. 1.1.1.37) isozymes zymogram in 11 different types of tissues of male and female Varicorhinus macrolepis. It had been found for the first time that the phenotype of malate dehydrogenase (MDH),acid phosphatase (ACP) and superoxide dismutase (SOD) showed difference between male and female V. macrolepis,and there was no difference among different individuals in the same sex. Therefore,the electrophoresis band of malate dehydrogenase,acid phosphatase and superoxide dismutase could be used as an indicator for the identification of gender and tissues of V. macrolepis,which would provide basic data for the developmental genetics,variety improvement and directed breeding of V. macrolepis groups,thus facilitating the development and protection of this valuable fish species.展开更多
[Objective] The research aimed to establish the two-dimensional electrophoresis（2-DE）technology which was suitable for the rapeseed proteome research.[Method] Xiangyou 17 was as the material.The sample preparation m...[Objective] The research aimed to establish the two-dimensional electrophoresis（2-DE）technology which was suitable for the rapeseed proteome research.[Method] Xiangyou 17 was as the material.The sample preparation method,gel concentration and loading amount,etc.in 2-DE technology were optimized.[Result] The best extraction method of total protein of rapeseed was TCA-acetone method,and the protein spots on 2-DE map were the most.When IPG strip（pH 3-10）and 12% gel were used,and the loading amount was 250 μg,the two-dimensional electrophoresis map with the clear background,good repeatability and high protein spot resolution was obtained.[Conclusion] The research laid the foundation for carrying out the rapeseed proteomics research.展开更多
Aim A capillary zone electrophoretic method （CZE） was used to determine the interactions between natural products and thrombin. Methods Samples containing natural products and thrombin at various ratios were incubat...Aim A capillary zone electrophoretic method （CZE） was used to determine the interactions between natural products and thrombin. Methods Samples containing natural products and thrombin at various ratios were incubated at 25 ℃ and then were separated by CZE with Tris-acetate buffer at pH 7.2. Each run could be accomplished within 5 min. Results In CZE, the peak width broadened due to the affinity interaction between natural products and thrombin. Compared with positive and negative control, the natural products （CB-1, CB-2） from Coreopsis tinctoria Nuttt. interacted with thrombin; CB-3 from Coreopsis tinctoria Nuttt. and HC-1, HC-2, HC-3 from Cistanche deserticola Ma. did not bind to thrombin. Both qualification and quantification characterizations of the binding were determined. Conclusion The established method is capable of sensitive and fast determination of natural products and thrombin interactions, it can be employed as an alternative method.展开更多
Allozymes of eleven species of Megophryinae in China were examined electrophoretically to investigate genetic diversity and phylogenetic relationships. Fourteen enzymes, presumptively coded by 24 loci were detected to...Allozymes of eleven species of Megophryinae in China were examined electrophoretically to investigate genetic diversity and phylogenetic relationships. Fourteen enzymes, presumptively coded by 24 loci were detected to be variable. Gene frequencies of each population at each locus were presented. The commonly used measure of genetic diversity, the average heterozygosity (H) were calculated based on gene frequencies. The results indicated that Megophryinae had a high level of genetic diversity in amphibians, an average H of 0.18, ranging from 0.058 to 0.28. Nei's (1978) genetic distances(Nei's D) were calculated for all possible population pairs. A dendrogram of 13 populations representing 11 species, 3 genera of Megophryinae were derived and presented by using UPGMA, based on Nei' s D. The assignment of Ophryophryne as a distinct genus were supported by an average Nei's D of 1.4067 which separated O. microstoma from all other populations.Subdivision of Brachytarsophrys from Megophrys was not supported by this study. Within Megophrys, three groups were recognized: (1)M. lateralis, M. giganticus and M. longipes; (2)M. palpebralespineosa, M. boettgeri and M. parva;(3) M. minor and M. kuatunensis. Three populations of M. omeimontis were closely related and share a clade independent from all other Megophrys, and B. feae as well.展开更多
[Objective] The aim was to optimize the SSR-PCR non-denatured polyacrylamide gel electrophoresis conditions in kernelled apricot.[Method]25 accessions of kernelled apricot and three accessions of edible apricot were s...[Objective] The aim was to optimize the SSR-PCR non-denatured polyacrylamide gel electrophoresis conditions in kernelled apricot.[Method]25 accessions of kernelled apricot and three accessions of edible apricot were selected as experimental materials to screen the repeatable SSR loci with high polymorphism by the use of SSR markers combined with non-denatured polyacrylamide gel electrophoresis.And the effect of different factors on electrophoresis conditions was compared to explore the optimal SSR-PCR non-denatured polyacrylamide gel electrophoresis conditions in kernelled apricot.[Result]The optimal non-denatured polyacrylamide gel electrophoresis conditions for SSR-PCR were established as follows：polyacrylamide gel concentration 6%,the ratio of acrylamide to bisacrylamide 29∶1,electrophoresis at 1 000 V for 2-3 h,and staining for 15 min within 0.1% AgNO3.[Conclusion]The optimum electrophoresis system has provided some technical foundations to further study the phylogenetic relationship of kernelled apricots by SSR markers.展开更多
[Objective] This study aimed to investigate the effect of zearalenone (ZEN) on DNA damage of porcine leydig cells. [Method] Porcine leydig cells cultured in vitro were collected to determine the median lethal dose (LD...[Objective] This study aimed to investigate the effect of zearalenone (ZEN) on DNA damage of porcine leydig cells. [Method] Porcine leydig cells cultured in vitro were collected to determine the median lethal dose (LD50) of ZEN with tetrazolium-based colorimetric assay (MTT assay). Comet assay was carried out to detect the DNA damage of porcine leydig cells exposed to at 0 (negative group), 1, 5, 10, 20, 40 μmol/L of ZEN. [Result] The percentage of cell tail was 16.67%, 34.00%, 40.67%, 52.00% and 64.67% under 0, 1, 5, 10 and 20 μmol/L of ZEN, respectively; the differences between the percentages of cell tail in various experimental groups had extremely significant statistical significance compared with the negative group (P<0.01), showing a significant dose-effect relationship; Tail length in various groups was 57.60±4.78, 57.75±6.25, 78.97±5.83, 100.50±6.94 and 146.83±12.31 μm, respectively; Tail DNA % in various groups was 21.29±2.25%, 22.24±2.43%, 31.21±6.27%, 37.45±4.33% and 60.68±9.83%, respectively; Tail length and Tail DNA % in experimental groups with ZEN concentration above 5 μmol/L showed significant differences (P<0.05) compared with the negative group, which showed an upward trend with the increase of ZEN concentration. [Conclusion] ZEN has genotoxic effect on porcine leydig cells, which can cause DNA damage, with a significant dose-effect relationship.展开更多
[Objective] This study was to elucidate the cellular and molecular mechanism of the development of heteromorphic leaves of Populus euphratica Oliv. [Method] By employing SDS-PAGE and 2-demensional electrophoresis (2-D...[Objective] This study was to elucidate the cellular and molecular mechanism of the development of heteromorphic leaves of Populus euphratica Oliv. [Method] By employing SDS-PAGE and 2-demensional electrophoresis (2-DE) techniques,proteins in various heteromorphic leaves from the same adult tree of P. euphratica were isolated and separated to the electrophoresis technique suitable for the separation and analysis of proteins in leaves of P. euphratica tree. [Results] There were significant differences in the expressions of proteins in various heteromorphic leaves of P. euphratica tree. SDS-PAGE pattern showed that bands of proteins with molecular weight of 57.2,13.2,30.2,23.9 and 33.3 kDa were remarkably different. 2-D electrophoresis pattern presented that proteins in leaves of P. euphratica tree mainly belong to acidic proteins distributed at pH value of 5.0-6.5 and with molecular weight of 20-40 kDa; totally 73 different protein spots were observed,of which 51 were up expressed and other 22 were down expressed in the serrated ovate leaves. [Conclusion] Based on these results,we speculate that regulated gene expression in leaves of P. euphratica tree results in the generation of different shapes of leaves,in order to adapt to the surroundings better.展开更多
With high performance capillary electrophoresis using -cyclodextrin or its deriveatives as the chiral selectors, five pairs of drug enantiomers were separated, The PH of the back- gmund electrolyte and the chiral sele...With high performance capillary electrophoresis using -cyclodextrin or its deriveatives as the chiral selectors, five pairs of drug enantiomers were separated, The PH of the back- gmund electrolyte and the chiral selector concentrations were optimized; and the effect of organic modifier on separation of chlorpheniramine enantiomers was also inver展开更多
Experiments of poly（dT）20 electrophoresis throughα-hemolysin nanopores were performed to unveil the electrophoretic transport mechanism of DNA through nanopores in high concentration potassium chloride solution. It...Experiments of poly（dT）20 electrophoresis throughα-hemolysin nanopores were performed to unveil the electrophoretic transport mechanism of DNA through nanopores in high concentration potassium chloride solution. It is found that there are two obvious current blockades induced by poly（dT）20 translocation and collision events. Both blockade currents increase linearly with the applied bias voltage. However, the normalized blockade currents are almost kept the same although variable bias voltages are applied. The collision time of poly（dT）20 in the luminal site of the pore remains constant for different voltages. The translocation speed of poly（dT）20through the nanopore decreases with the increase of bias voltage. It is because as the potential increases, the drag force on the homopolymer helps it to crumple into a cluster much easier due to the poor stacking of thymine residues compared with homopolymers consisting of other nucleotides. Molecular dynamics simulations further confirm the experimental results. Increasing the applied bias voltage can slowdown the translocation velocity of the flexible poly（dT）20, which favors increasing the precision of single molecule detection by using nanopores.展开更多
Mycophenolic acid （MPA）, an immunosuppressive drug, was determined in human plasma by a simple capillary zone electrophoresis （CZE） method. A bare fused-silica capillary with an internal diameter （i.d.） of 75/am...Mycophenolic acid （MPA）, an immunosuppressive drug, was determined in human plasma by a simple capillary zone electrophoresis （CZE） method. A bare fused-silica capillary with an internal diameter （i.d.） of 75/am and an effective length of 50 cm was used for the separation. A 200 }aL plasma sample was deproteinized with 400 μL acetonitrile, and the supematant was directly injected into the capillary after a water plug at a pressure of 0.5 psi for 10 s. Boric acid （H3BO3, 200 mmol/L） solution adjusted to a pH of 8.60 with 1 mol/L sodium hydroxide （NaOH） solution was selected as the background electrolyte （BGE） through a uniform design. Calibration curve established on each day was linear over the MPA concentration range of 0.625-30.0 Bg/mL in human plasma with the correlation coefficient （r） larger than 0.9997. The limit of detection （LOD） was 0.200 ~tg/mL. Intra- and inter-day precisions at three concentrations within the calibration range were less than 4.45%. Plasma samples collected from 14 renal transplant recipients treated with the prodrug mycophenolate mofetil （MMF） were analyzed successfully by both the developed CZE technique and a validated high-performance liquid chromatography-tandem mass spectrometry （HPLC-MS/MS） technique. The mean absolute and relative differences between the concentrations measured by the two methods were -0.17 and -0.20%, respectively. In conclusion, the CZE method described here was validated to be accurate, precise, and economical for the clinical quantification of MPA.展开更多
A peptides migration model based on the principle of mechanics is presented in capillary zone electrophoresis (CZE). It is shown that the migration that the (tr) is a function of electric (Q), relative molecular mass ...A peptides migration model based on the principle of mechanics is presented in capillary zone electrophoresis (CZE). It is shown that the migration that the (tr) is a function of electric (Q), relative molecular mass (Mr), conformation parameter (Rc) of peptides and electrophoretic condition parameter(A). The conformation parameter is introduced to characterize multifarious shapes owing to the complex conlormation and the various kinds of macromolecules, where Rc≥1/3. The parameters A and Rc can be obtained from experimental data. The times of migration of the nine standard peptides were measured in pH 2.5buffer on different electrophoretic conditions in CZE. The experimental results agreed well with the theoretical prediction.展开更多
Capillary zone electrophoresis has been applied to the analysis of nucleotides. The effects of buffer concentration. pH and other operating conditions on the separation were investigated and optimized. By using the me...Capillary zone electrophoresis has been applied to the analysis of nucleotides. The effects of buffer concentration. pH and other operating conditions on the separation were investigated and optimized. By using the method, separation and identification of nuclotides in swine tissues were completed.展开更多
Rhizoma Polygoni Bistortae has been widely used as an important medicinal herb in the Far East for a few thousand years, and it has gained more recognition in the last few decades. In the present study, we used a high...Rhizoma Polygoni Bistortae has been widely used as an important medicinal herb in the Far East for a few thousand years, and it has gained more recognition in the last few decades. In the present study, we used a high-performance capillary electrophoresis （HPCE） method for determination of the gallic acid （GA） in Quanshen （QS） pieces. The analysis of GA in QS pieces was successfully achieved in less than 6 min by capillary electrophoresis with UV detection at a wavelength of 290 nm. The background electrolyte was 20 mmol/L sodium borate solution （pH 9.00）. The separation was performed in an uncoated fused-silica capillary （52 cm^50 lam） with an effective length of 40 cm, and the applied voltage was 20 kV. Sample solutions were injected for l0 s at an atmospheric pressure of 5 KPa. The result showed good resolution and precision （the intra-day RSD was 0.37%）. The calibration curves of the standard solutions were linear in the range of 0.22-2.25 mg/mL, with correlation coefficients greater than 0.9990. The recovery of the method ranged from 100.6% to 107.9%, and the average recovery of GA was 105.9% with an RSD of 3.63%. HPCE is a powerful technology, and it is the first time to successfully apply such a method for the determination of GA in the traditional Chinese medicine.展开更多
The present paper covers the separation and determination of cytosine , 9-N-di- hydroxypropyltheophylline , theobromine , adenine , guanine , uracil , theophylline , 1-hydroxyethyl-5-fluorouracil , xanthine , uric aci...The present paper covers the separation and determination of cytosine , 9-N-di- hydroxypropyltheophylline , theobromine , adenine , guanine , uracil , theophylline , 1-hydroxyethyl-5-fluorouracil , xanthine , uric acid , 6-mercaptopurine , 5-fluo- rouracil and 1 -propionyloxyuracil by capillary zone electrophoresis. In an uncoated silica capillary(70 cm ×75 μm i. d. effective length 63 cm) , thirteen bases could be separated within 1 1 min under the optimum conditions : 0. 01 mol/L(pH 10. 0) bo- rate buffer, 40℃ , 15 kV, 260 nm and 2 s load. Efficiencies up to 3. 1 ×105 theoreti- cal plates (4. 9 ×105 plates/m) were achieved.展开更多
基金financially supported by Beijing Municipal Science&Technology Commission,China(Grant No.:Z221100007922015)Youth Development Research Foundation of National Institutes for Food and Drug Control,China(Grant No.:2020B1).
文摘Adventitious agents,comprising unintentionally introduced microorganisms in the production of biological products,pose a significant challenge in ensuring the safety of gene therapy products.The revised International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use(ICH)guildline Q5A(R2)from September 2022 highlights the inclusion of viral vector-based gene therapy products in safety discussions,emphasizing controls in material sourcing,testing,and viral clearance[1].Detecting adventitious virus contamination is complex due to the unique characteristics of gene therapy products and the limitations of routine testing methods.The US Food and Drug Administration(FDA)recommends incorporating routine and specific virus detection methods,including those outlined in various pharmacopeias.Existing control methods have limitations,prompting the need for highly sensitive and broad-spectrum detection approaches.Unlike traditional biological products,gene therapy products primarily consist of live viruses,necessitating methods that distinguish between the main virus and adventitious viruses.Current virus detection techniques,such as polymerase chain reaction(PCR),sequencing,mass spectrometry,and DNA microarrays[2e4],have their drawbacks.
基金supported by the National Natural Science Foundation of China(Nos.21874060 and 22174058,U21A20282)the Science and Technology program of Gansu Province(No.22JR5RA476)。
文摘DNA repair enzymes are important in the repair of DNA lesions for maintaining the genome stability,and their abnormal expression induced various human cancers.Simultaneous detection of these DNA enzymes could provide convincing evidence based on the comparison of the activity of multiple enzymes than on that of single enzyme.Although fluorescence approach has been applied for the simultaneous detection both of DNA repair enzymes,the spectral overlap and multiwavelength excitation severely restrict the number of available fluorophores.Thus,it is difficult to simultaneously detect three enzymes in a single analysis by fluorescence detection.Herein,we developed a method for the simultaneous determination of three DNA repair enzymes including human flap DNA endonuclease 1(FEN1),human alkyladenine DNA glycosylase(hAAG)and uracil DNA glycosylase(UDG)based on the combination of template-free amplification system with capillary electrophoresis-laser induced fluorescence(CE-LIF)detection.The amplification system was adopted to transfer and amplify the enzymatic products into different length DNA fragments which could be separated effectively by CE-LIF without the complicated modification of the capillary inner wall or labeling different tails on signal probes for separation.The method demonstrated a detection limit of 0.07 U/mL(0.08-160 U/mL)for FEN1,2.40 U/mL(2.5-250U/mL)for hAAG and 2.1×10^(-4)U/mL(0.0004-2.5 U/mL)for UDG,the relative standard deviations(RSDs)of peak time and peak area for different analytes were as follows:2.50%-4,37%and 3.24%-7.18%(inter-day);1.37%-2.71%and 1.43%-3.02%(intra-day),4.28%-6.08%and 4.16%-7.57%(column to column),respectively.And it can identify the inhibitor-like drugs,evaluate enzymatic kinetics and achieve the detection of three enzymes in cell extracts,providing a simple and powerful platform for simultaneous detection of more DNA repair enzymes.
基金Research Foundation from National Innovation Center of Advanced Dyeing&Finishing Technology,China(No.2022GCJJ15)。
文摘Reactive dyes with different reactive groups exhibit different hydrolysis and dyeing behaviors.This is particularly evident in the combination dyeing process,where the competion between hydrolysis and dyeing reactions increases the complexity.Therefore,developing an effective method to monitor the changes in reactive dyes during the dyeing process is important.This study aims to develop a capillary electrophoresis(CE)technique combined with an ultraviolet(UV)detector(CE-UV)for detecting three reactive dyes and their six derivatives(a total of nine analytes).The optimized CE conditions are 20.0 mmol/L sodium tetraborate(Na_(2)B_(4)O_(7)·10H_(2)O),acetonitrile(ACN)with a volume fraction of 15.0%,20.0 mmol/L α-cyclodextrin(α-CD),and at a pH value of 9.0(adjusted with 0.5 mol/L H_(3)BO_(3)).The limit of detection(LOD)(a signal-to-noise ratio of 3)for the nine analytes ranges from 1.38 to 5.06 mg/L.The relative standard deviations(RSDs)for peak areas and migration time are 2.19%-4.96%and 0.29%-2.75%,respectively.The method is capable of accurately identifying three reactive dyes and their six derivatives and monitoring alterations in composition and dyeing behavior during single and combination dyeing processes.
文摘Capillary electrophoresis(CE),recognized for its minimal reagent dosage,rapid analysis,and high efficiency,holds significant promise for the analysis of traditional Chinese medicines(TCM).This article reviews the application of CE in the determination of active ingredients in TCM.The active substances of herbal medicines have been classified and discussed based on their chemical properties,and the CE methods applied to different substances are summarized and discussed.This article also provides some suggestions for future research and improvement.
基金Supported by Natural Science Foundation of Hebei Province(C2008000591)~~
文摘[Objective] The like-rocket immunoelectrophoresis was used to explore a new feasible electrophoresis method for single cell gel electrophoresis assay （comet assay）.[Method] The like-rocket immunoelectrophoresis was used for single cell gel electrophoresis assay to detect DNA damage at single cell level,then it was compared with traditional electrophoresis method to analyze its advantage and disadvantages.[Result] Under cell DNA undamaged state,the results of two electrophoresis methods were consistent.When cell DNA was damaged,the comet tail divergence of some cells under traditional electrophoresis method were drifted,however,the comet tail image of like-rocket immunoelectrophoresis was concentrated and not shifted.[Conclusion] The like-rocket immunoelectrophoresis had some advantages.
文摘A recombinant inbred population (RI) was constructed from a cross between B5, an introgression. line from the wild rice Oryza officinalis Wall. ex Watt, and susceptible cultivar Minghui 63 ( O. sativa L.). The brown planthopper ( BPH) resistances of RI lines were evaluated. Based on bulked segregant analysis (BSA), two protein bulks were made by extracting proteins from equally mixed seedlings of extremely resistant and susceptible plants selected from the RI population, respectively. Two-dimensional electrophoresis was used to detect the changes of polypeptide pattern. Results showed that a protein P40 ( pI 6.3, Mw 40 kD) was significantly reduced or vanished after BPH infestation for 48 h in the susceptible bulk, while it remained uninfluenced in the resistant bulk. In connection with the physiological changes of the resistant and susceptible lines subjected to BPH sucking, we suppose that the protein P40 is related to the interaction responses of lice plants to BPH infestation.
基金Supported by National Natural Science Foundation of China(30700071 )Natural Science Foundation of Shandong Province(Y2008D03 )Science and Technology Program of Qingdao City(08-1-27-jch)~~
文摘Polyacrylamide gel electrophoresis (PAGE) and biochemical staining method were used in this study for the analysis on malate dehydrogenase (MDH,E.C. 1.1.1.37) isozymes zymogram in 11 different types of tissues of male and female Varicorhinus macrolepis. It had been found for the first time that the phenotype of malate dehydrogenase (MDH),acid phosphatase (ACP) and superoxide dismutase (SOD) showed difference between male and female V. macrolepis,and there was no difference among different individuals in the same sex. Therefore,the electrophoresis band of malate dehydrogenase,acid phosphatase and superoxide dismutase could be used as an indicator for the identification of gender and tissues of V. macrolepis,which would provide basic data for the developmental genetics,variety improvement and directed breeding of V. macrolepis groups,thus facilitating the development and protection of this valuable fish species.
基金Supported by National 863 Project(2010AA101503)National Science and Technology Support Planning Item(2006BAD05A12)Student Innovation Fund Item of Hefei University of Technology(XS2010100)~~
文摘[Objective] The research aimed to establish the two-dimensional electrophoresis（2-DE）technology which was suitable for the rapeseed proteome research.[Method] Xiangyou 17 was as the material.The sample preparation method,gel concentration and loading amount,etc.in 2-DE technology were optimized.[Result] The best extraction method of total protein of rapeseed was TCA-acetone method,and the protein spots on 2-DE map were the most.When IPG strip（pH 3-10）and 12% gel were used,and the loading amount was 250 μg,the two-dimensional electrophoresis map with the clear background,good repeatability and high protein spot resolution was obtained.[Conclusion] The research laid the foundation for carrying out the rapeseed proteomics research.
文摘Aim A capillary zone electrophoretic method （CZE） was used to determine the interactions between natural products and thrombin. Methods Samples containing natural products and thrombin at various ratios were incubated at 25 ℃ and then were separated by CZE with Tris-acetate buffer at pH 7.2. Each run could be accomplished within 5 min. Results In CZE, the peak width broadened due to the affinity interaction between natural products and thrombin. Compared with positive and negative control, the natural products （CB-1, CB-2） from Coreopsis tinctoria Nuttt. interacted with thrombin; CB-3 from Coreopsis tinctoria Nuttt. and HC-1, HC-2, HC-3 from Cistanche deserticola Ma. did not bind to thrombin. Both qualification and quantification characterizations of the binding were determined. Conclusion The established method is capable of sensitive and fast determination of natural products and thrombin interactions, it can be employed as an alternative method.
基金This work was financially supported by Natural Science Foundation of China.
文摘Allozymes of eleven species of Megophryinae in China were examined electrophoretically to investigate genetic diversity and phylogenetic relationships. Fourteen enzymes, presumptively coded by 24 loci were detected to be variable. Gene frequencies of each population at each locus were presented. The commonly used measure of genetic diversity, the average heterozygosity (H) were calculated based on gene frequencies. The results indicated that Megophryinae had a high level of genetic diversity in amphibians, an average H of 0.18, ranging from 0.058 to 0.28. Nei's (1978) genetic distances(Nei's D) were calculated for all possible population pairs. A dendrogram of 13 populations representing 11 species, 3 genera of Megophryinae were derived and presented by using UPGMA, based on Nei' s D. The assignment of Ophryophryne as a distinct genus were supported by an average Nei's D of 1.4067 which separated O. microstoma from all other populations.Subdivision of Brachytarsophrys from Megophrys was not supported by this study. Within Megophrys, three groups were recognized: (1)M. lateralis, M. giganticus and M. longipes; (2)M. palpebralespineosa, M. boettgeri and M. parva;(3) M. minor and M. kuatunensis. Three populations of M. omeimontis were closely related and share a clade independent from all other Megophrys, and B. feae as well.
基金Supported by the Scientific and Technological Program of Educational Department of Hebei Province(No.ZH2007116)~~
文摘[Objective] The aim was to optimize the SSR-PCR non-denatured polyacrylamide gel electrophoresis conditions in kernelled apricot.[Method]25 accessions of kernelled apricot and three accessions of edible apricot were selected as experimental materials to screen the repeatable SSR loci with high polymorphism by the use of SSR markers combined with non-denatured polyacrylamide gel electrophoresis.And the effect of different factors on electrophoresis conditions was compared to explore the optimal SSR-PCR non-denatured polyacrylamide gel electrophoresis conditions in kernelled apricot.[Result]The optimal non-denatured polyacrylamide gel electrophoresis conditions for SSR-PCR were established as follows：polyacrylamide gel concentration 6%,the ratio of acrylamide to bisacrylamide 29∶1,electrophoresis at 1 000 V for 2-3 h,and staining for 15 min within 0.1% AgNO3.[Conclusion]The optimum electrophoresis system has provided some technical foundations to further study the phylogenetic relationship of kernelled apricots by SSR markers.
文摘[Objective] This study aimed to investigate the effect of zearalenone (ZEN) on DNA damage of porcine leydig cells. [Method] Porcine leydig cells cultured in vitro were collected to determine the median lethal dose (LD50) of ZEN with tetrazolium-based colorimetric assay (MTT assay). Comet assay was carried out to detect the DNA damage of porcine leydig cells exposed to at 0 (negative group), 1, 5, 10, 20, 40 μmol/L of ZEN. [Result] The percentage of cell tail was 16.67%, 34.00%, 40.67%, 52.00% and 64.67% under 0, 1, 5, 10 and 20 μmol/L of ZEN, respectively; the differences between the percentages of cell tail in various experimental groups had extremely significant statistical significance compared with the negative group (P<0.01), showing a significant dose-effect relationship; Tail length in various groups was 57.60±4.78, 57.75±6.25, 78.97±5.83, 100.50±6.94 and 146.83±12.31 μm, respectively; Tail DNA % in various groups was 21.29±2.25%, 22.24±2.43%, 31.21±6.27%, 37.45±4.33% and 60.68±9.83%, respectively; Tail length and Tail DNA % in experimental groups with ZEN concentration above 5 μmol/L showed significant differences (P<0.05) compared with the negative group, which showed an upward trend with the increase of ZEN concentration. [Conclusion] ZEN has genotoxic effect on porcine leydig cells, which can cause DNA damage, with a significant dose-effect relationship.
文摘[Objective] This study was to elucidate the cellular and molecular mechanism of the development of heteromorphic leaves of Populus euphratica Oliv. [Method] By employing SDS-PAGE and 2-demensional electrophoresis (2-DE) techniques,proteins in various heteromorphic leaves from the same adult tree of P. euphratica were isolated and separated to the electrophoresis technique suitable for the separation and analysis of proteins in leaves of P. euphratica tree. [Results] There were significant differences in the expressions of proteins in various heteromorphic leaves of P. euphratica tree. SDS-PAGE pattern showed that bands of proteins with molecular weight of 57.2,13.2,30.2,23.9 and 33.3 kDa were remarkably different. 2-D electrophoresis pattern presented that proteins in leaves of P. euphratica tree mainly belong to acidic proteins distributed at pH value of 5.0-6.5 and with molecular weight of 20-40 kDa; totally 73 different protein spots were observed,of which 51 were up expressed and other 22 were down expressed in the serrated ovate leaves. [Conclusion] Based on these results,we speculate that regulated gene expression in leaves of P. euphratica tree results in the generation of different shapes of leaves,in order to adapt to the surroundings better.
文摘With high performance capillary electrophoresis using -cyclodextrin or its deriveatives as the chiral selectors, five pairs of drug enantiomers were separated, The PH of the back- gmund electrolyte and the chiral selector concentrations were optimized; and the effect of organic modifier on separation of chlorpheniramine enantiomers was also inver
基金The National Natural Science Foundation of China(No.51435003,51375092)Research Program of Chongqing Municipal Education Commission(No.KJ1401030)+1 种基金the Research & Innovation Program for Graduate Student in Universities of Jiangsu Province(No.KYLX_0100)the Scientific Research Foundation of Graduate School of Southeast University(No.YBJJ1540)
文摘Experiments of poly（dT）20 electrophoresis throughα-hemolysin nanopores were performed to unveil the electrophoretic transport mechanism of DNA through nanopores in high concentration potassium chloride solution. It is found that there are two obvious current blockades induced by poly（dT）20 translocation and collision events. Both blockade currents increase linearly with the applied bias voltage. However, the normalized blockade currents are almost kept the same although variable bias voltages are applied. The collision time of poly（dT）20 in the luminal site of the pore remains constant for different voltages. The translocation speed of poly（dT）20through the nanopore decreases with the increase of bias voltage. It is because as the potential increases, the drag force on the homopolymer helps it to crumple into a cluster much easier due to the poor stacking of thymine residues compared with homopolymers consisting of other nucleotides. Molecular dynamics simulations further confirm the experimental results. Increasing the applied bias voltage can slowdown the translocation velocity of the flexible poly（dT）20, which favors increasing the precision of single molecule detection by using nanopores.
基金Plan of Vitalizing Education Action Geared to the 21st Century(Plan 985)the 1st Phase Fund of Peking University 985 Project.
文摘Mycophenolic acid （MPA）, an immunosuppressive drug, was determined in human plasma by a simple capillary zone electrophoresis （CZE） method. A bare fused-silica capillary with an internal diameter （i.d.） of 75/am and an effective length of 50 cm was used for the separation. A 200 }aL plasma sample was deproteinized with 400 μL acetonitrile, and the supematant was directly injected into the capillary after a water plug at a pressure of 0.5 psi for 10 s. Boric acid （H3BO3, 200 mmol/L） solution adjusted to a pH of 8.60 with 1 mol/L sodium hydroxide （NaOH） solution was selected as the background electrolyte （BGE） through a uniform design. Calibration curve established on each day was linear over the MPA concentration range of 0.625-30.0 Bg/mL in human plasma with the correlation coefficient （r） larger than 0.9997. The limit of detection （LOD） was 0.200 ~tg/mL. Intra- and inter-day precisions at three concentrations within the calibration range were less than 4.45%. Plasma samples collected from 14 renal transplant recipients treated with the prodrug mycophenolate mofetil （MMF） were analyzed successfully by both the developed CZE technique and a validated high-performance liquid chromatography-tandem mass spectrometry （HPLC-MS/MS） technique. The mean absolute and relative differences between the concentrations measured by the two methods were -0.17 and -0.20%, respectively. In conclusion, the CZE method described here was validated to be accurate, precise, and economical for the clinical quantification of MPA.
文摘A peptides migration model based on the principle of mechanics is presented in capillary zone electrophoresis (CZE). It is shown that the migration that the (tr) is a function of electric (Q), relative molecular mass (Mr), conformation parameter (Rc) of peptides and electrophoretic condition parameter(A). The conformation parameter is introduced to characterize multifarious shapes owing to the complex conlormation and the various kinds of macromolecules, where Rc≥1/3. The parameters A and Rc can be obtained from experimental data. The times of migration of the nine standard peptides were measured in pH 2.5buffer on different electrophoretic conditions in CZE. The experimental results agreed well with the theoretical prediction.
文摘Capillary zone electrophoresis has been applied to the analysis of nucleotides. The effects of buffer concentration. pH and other operating conditions on the separation were investigated and optimized. By using the method, separation and identification of nuclotides in swine tissues were completed.
基金supported by Beijing Kaiao Technology Development Co., Ltd., China and Beijing Friendship Hospital, Capital University of Medical Sciences Beijing
文摘Rhizoma Polygoni Bistortae has been widely used as an important medicinal herb in the Far East for a few thousand years, and it has gained more recognition in the last few decades. In the present study, we used a high-performance capillary electrophoresis （HPCE） method for determination of the gallic acid （GA） in Quanshen （QS） pieces. The analysis of GA in QS pieces was successfully achieved in less than 6 min by capillary electrophoresis with UV detection at a wavelength of 290 nm. The background electrolyte was 20 mmol/L sodium borate solution （pH 9.00）. The separation was performed in an uncoated fused-silica capillary （52 cm^50 lam） with an effective length of 40 cm, and the applied voltage was 20 kV. Sample solutions were injected for l0 s at an atmospheric pressure of 5 KPa. The result showed good resolution and precision （the intra-day RSD was 0.37%）. The calibration curves of the standard solutions were linear in the range of 0.22-2.25 mg/mL, with correlation coefficients greater than 0.9990. The recovery of the method ranged from 100.6% to 107.9%, and the average recovery of GA was 105.9% with an RSD of 3.63%. HPCE is a powerful technology, and it is the first time to successfully apply such a method for the determination of GA in the traditional Chinese medicine.
文摘The present paper covers the separation and determination of cytosine , 9-N-di- hydroxypropyltheophylline , theobromine , adenine , guanine , uracil , theophylline , 1-hydroxyethyl-5-fluorouracil , xanthine , uric acid , 6-mercaptopurine , 5-fluo- rouracil and 1 -propionyloxyuracil by capillary zone electrophoresis. In an uncoated silica capillary(70 cm ×75 μm i. d. effective length 63 cm) , thirteen bases could be separated within 1 1 min under the optimum conditions : 0. 01 mol/L(pH 10. 0) bo- rate buffer, 40℃ , 15 kV, 260 nm and 2 s load. Efficiencies up to 3. 1 ×105 theoreti- cal plates (4. 9 ×105 plates/m) were achieved.
