N-Conjugated antigen was synthesized and polyclonal antibody with high specificity was obtained from immunizing animals. With this polyclonal antibody, a rapid and efficient CEIA-LIF method was developed to determine ...N-Conjugated antigen was synthesized and polyclonal antibody with high specificity was obtained from immunizing animals. With this polyclonal antibody, a rapid and efficient CEIA-LIF method was developed to determine the free morphine in urine of abusers. The detection limit was calculated to be 40 ng/mL. Simulated urine samples were analyzed with good recoveries, which showed the feasibility of its application in specific morphine determination in urine of morphine abusers.展开更多
A novel approach for analysis of amino acids in individual erythrocytes was established. In this method, the derivatization reagent was introduced into the living cells by electroporation. After derivatization, the am...A novel approach for analysis of amino acids in individual erythrocytes was established. In this method, the derivatization reagent was introduced into the living cells by electroporation. After derivatization, the amino acids in a single cell were determined by capillary electrophoresis with laser-induced fluorescence detection.展开更多
Objective Lower respiratory tract infections continue to pose a significant threat to human health. It is important to accurately and rapidly detect respiratory bacteria. To compensate for the limits of current respir...Objective Lower respiratory tract infections continue to pose a significant threat to human health. It is important to accurately and rapidly detect respiratory bacteria. To compensate for the limits of current respiratory bacteria detection methods, we developed a combination of multiplex polymerase chain reaction （PCR） and capillary electrophoresis （MPCE） assay to detect thirteen bacterial pathogens responsible for lower respiratory tract infections, including Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catorrholis, Pseudomonas aeruginosa, Klebsiella pneumoniae, Escherichia coli, Staphylococcus aureus, Mycoplasma pneumoniae, Legionella spp., Bordetella pertussis, Mycobacterium tuberculosis complex, Corynebactefium diphthefiae, and Streptococcus pyogenes. Methods Three multiplex PCR reactions were built, and the products were analyzed by capillary electrophoresis using the high-throughput DNA analyzer. The specificity of the MPCE assay was examined and the detection limit was evaluated using DNA samples from each bacterial strain and the simulative samples of each strain. This assay was further evaluated using 152 clinical specimens and compared with real-time PCR reactions. For this assay, three nested-multiplex-PCRs were used to detect these clinical specimens. Results The detection limits of the MPCE assay for the 13 pathogens were very low and ranged from 10-7 to 10-2 ng/μL. Furthermore, analysis of the 252 clinical specimens yielded a specificity ranging from 96.5%-100.0%, and a sensitivity of 100.0% for the 13 pathogens. Conclusion This study revealed that the MPCE with high specificity and sensitivity. This assay survey of respiratory pathogens. assay is a rapid, reliable, and high-throughput method has great potential in the molecular epidemiological.展开更多
A simple method was developed to quantify DNA fragments such as PCR (polymerase chain reaction) products by capillary electrophoresis. Restraint fragments with different lengths were employed as internal standards in ...A simple method was developed to quantify DNA fragments such as PCR (polymerase chain reaction) products by capillary electrophoresis. Restraint fragments with different lengths were employed as internal standards in the study, which makes it possible for the evaluation of the quantity of PCR product in a single run.展开更多
The main goal of this work was to quantify the detection of colistin at low levels in urine samples through the practical application of mixed surfactant micellar electrokinetic chromatography–laser-induced fluoresce...The main goal of this work was to quantify the detection of colistin at low levels in urine samples through the practical application of mixed surfactant micellar electrokinetic chromatography–laser-induced fluorescence (MEKC-LIF) analysis method using its advantage of sensitivity and to examine direct injection of biological samples. Colistin (po- lymyxin E) has neither strong UV chromophore nor fluorophore. So, its assay for metabolism, pharmacokinetics studies for bioavailability and bioequivalence are difficult because of poor detectability. Therefore an enhanced UV or fluores-cence detection by chemical derivatization is required. MEKC-LIF method was proposed for colistin with a 488/520 nm argon-ion laser using a pre-CE derivatization with fluorescein isothiocyanate (FITC). Borate buffer was used as background buffer (BGB). The different parameters affecting the proposed derivatization reaction including concentration of the derivatizing reagent, reaction time and temperature were studied and optimized. The derivative was stable for up to 3 days. Different micelles (TX-100 and SDS) were examined as BGB additives separately but negative-charged mixed micelles (SDS/TX-100) were shown to be the best additive to BGB for the analysis of colistin particularly in human urine as they enhance both selectivity and sensitivity of the proposed method. BGB was used with pH 9.5, 10 kV, 8 s inj time, capillary length 75 cm × 75 μm ID (66 cm effective length), detection was LIF Ex 488 nm;Em 520 nm. The method was applied to colistin analysis in human urine and the recovery was > 98% (n = 5). LOD and LOQ in urine after pre-column derivatization using FITC were 100 and 250 ng/ml, respectively. Urine samples were analysed by direct injection without sample pre-treatment. The mechanism of enhancement of fluorescence of the derivative by surfactant was proposed.展开更多
Human erythropoietin （hEPO）, an endogenous glycoprotein, plays a fundamental role in erythropoiesis controlling the formation of red blood cells. Production of recombinant human erythropoietin （rhEPO） has made it ...Human erythropoietin （hEPO）, an endogenous glycoprotein, plays a fundamental role in erythropoiesis controlling the formation of red blood cells. Production of recombinant human erythropoietin （rhEPO） has made it possible for its abuse in competitive sports. In this work, pre-capillary and on-capillary derivatization by 5-furoylquinoline-3-carboxaldehyde （FQ） and fluorescein isothiocyanate （FITC） for the detection of rhEPO by capillary electrophoresis with laser-induced fluorescence detection （CE-LIF） were compared. FQ pre-capillary labeling improves sensitivity but degrades the glycoforms separation due to the inhomogeneity of the reaction products from multiple labeling. Compared with FITC pre-capillary derivatization with the excess fluorescent background, the on-capillary FQ derivatization method can provide shorter analysis time, lower background, and better selectivity. It is demonstrated that, through optimizing reaction conditions of FQ on-capillary derivatization, both high sensitivity and satisfactory resolution for the analysis of the be used for the glycoforms profiling and quality control of rhEPO doping control analysis. glycoforms of rhEPO could be obtained. This method can It may be used as a candidate method for fast screening in展开更多
This work described a sensitive method for determination of metoprolol in rabbit plasma.The method involved purification by ultrafiltration,derivatization with fluorescein isothiocyanate,determination by capillary ele...This work described a sensitive method for determination of metoprolol in rabbit plasma.The method involved purification by ultrafiltration,derivatization with fluorescein isothiocyanate,determination by capillary electrophoresis（CE） coupled with laser-induced fluorescence（LIF） detector.Other components in plasma including a variety of amino acids and proteins did not interfere with the determination of metoprolol in experimental condition.The assay had a wide range（2.0-500 ng/mL） of linearity and a detection limit of 0.8 ng/mL.The intra- and inter-day precisions were satisfactory with relative standard deviation（RSD） less than 10.0%and accuracy within 10.0%.This method was successfully applied to pharmacokinetic study of metoprolol in rabbit blood.展开更多
FITC labeled amino acids have been separated using a home-built capillary electrophoresis with a laser-induced fluorescence detection (CE-LIF) system. Seventeen peaks can now be generated from the twenty common amino ...FITC labeled amino acids have been separated using a home-built capillary electrophoresis with a laser-induced fluorescence detection (CE-LIF) system. Seventeen peaks can now be generated from the twenty common amino acids. The key conditions lie in the optimization of pH, buffer electrolytes and buffer additives.展开更多
Proteomics is becoming more and more mature, but the detection of low abundance proteins is still a difficult task. Laser-induced fluorescence（LIF） detection is one of the most sensitive detection methods in a capil...Proteomics is becoming more and more mature, but the detection of low abundance proteins is still a difficult task. Laser-induced fluorescence（LIF） detection is one of the most sensitive detection methods in a capillary electrophoresis（CE） system. However, most proteins do not exhibit favourable native fluorescence, a derivatization procedure is necessary for LIF detection of proteins. Since the derivatization reaction between protein and fluorescent reagent takes place after the separation of protein, the separation cannot be compromised by multiple derivatization products, the post-column derivatization becomes an attractive method for derivatization in CE-LIF system.展开更多
文摘N-Conjugated antigen was synthesized and polyclonal antibody with high specificity was obtained from immunizing animals. With this polyclonal antibody, a rapid and efficient CEIA-LIF method was developed to determine the free morphine in urine of abusers. The detection limit was calculated to be 40 ng/mL. Simulated urine samples were analyzed with good recoveries, which showed the feasibility of its application in specific morphine determination in urine of morphine abusers.
基金supported by the National Natural Science Foundation of China,the State Key Laboratory of Microbial Technology,Shandong University.
文摘A novel approach for analysis of amino acids in individual erythrocytes was established. In this method, the derivatization reagent was introduced into the living cells by electroporation. After derivatization, the amino acids in a single cell were determined by capillary electrophoresis with laser-induced fluorescence detection.
基金supported by grants from the Priority Project on Infectious Disease Control and Prevention(2012ZX10004215,2013ZX10004610)from Ministry of Health,China,and the Science Foundation for the State Key Laboratory for Infectious Disease Prevention and Control from China(Grant No.2015SKLID508)the National Natural Science Foundation of China(Grant No.81671985)and(Grant No.81170009)
文摘Objective Lower respiratory tract infections continue to pose a significant threat to human health. It is important to accurately and rapidly detect respiratory bacteria. To compensate for the limits of current respiratory bacteria detection methods, we developed a combination of multiplex polymerase chain reaction （PCR） and capillary electrophoresis （MPCE） assay to detect thirteen bacterial pathogens responsible for lower respiratory tract infections, including Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catorrholis, Pseudomonas aeruginosa, Klebsiella pneumoniae, Escherichia coli, Staphylococcus aureus, Mycoplasma pneumoniae, Legionella spp., Bordetella pertussis, Mycobacterium tuberculosis complex, Corynebactefium diphthefiae, and Streptococcus pyogenes. Methods Three multiplex PCR reactions were built, and the products were analyzed by capillary electrophoresis using the high-throughput DNA analyzer. The specificity of the MPCE assay was examined and the detection limit was evaluated using DNA samples from each bacterial strain and the simulative samples of each strain. This assay was further evaluated using 152 clinical specimens and compared with real-time PCR reactions. For this assay, three nested-multiplex-PCRs were used to detect these clinical specimens. Results The detection limits of the MPCE assay for the 13 pathogens were very low and ranged from 10-7 to 10-2 ng/μL. Furthermore, analysis of the 252 clinical specimens yielded a specificity ranging from 96.5%-100.0%, and a sensitivity of 100.0% for the 13 pathogens. Conclusion This study revealed that the MPCE with high specificity and sensitivity. This assay survey of respiratory pathogens. assay is a rapid, reliable, and high-throughput method has great potential in the molecular epidemiological.
文摘A simple method was developed to quantify DNA fragments such as PCR (polymerase chain reaction) products by capillary electrophoresis. Restraint fragments with different lengths were employed as internal standards in the study, which makes it possible for the evaluation of the quantity of PCR product in a single run.
文摘The main goal of this work was to quantify the detection of colistin at low levels in urine samples through the practical application of mixed surfactant micellar electrokinetic chromatography–laser-induced fluorescence (MEKC-LIF) analysis method using its advantage of sensitivity and to examine direct injection of biological samples. Colistin (po- lymyxin E) has neither strong UV chromophore nor fluorophore. So, its assay for metabolism, pharmacokinetics studies for bioavailability and bioequivalence are difficult because of poor detectability. Therefore an enhanced UV or fluores-cence detection by chemical derivatization is required. MEKC-LIF method was proposed for colistin with a 488/520 nm argon-ion laser using a pre-CE derivatization with fluorescein isothiocyanate (FITC). Borate buffer was used as background buffer (BGB). The different parameters affecting the proposed derivatization reaction including concentration of the derivatizing reagent, reaction time and temperature were studied and optimized. The derivative was stable for up to 3 days. Different micelles (TX-100 and SDS) were examined as BGB additives separately but negative-charged mixed micelles (SDS/TX-100) were shown to be the best additive to BGB for the analysis of colistin particularly in human urine as they enhance both selectivity and sensitivity of the proposed method. BGB was used with pH 9.5, 10 kV, 8 s inj time, capillary length 75 cm × 75 μm ID (66 cm effective length), detection was LIF Ex 488 nm;Em 520 nm. The method was applied to colistin analysis in human urine and the recovery was > 98% (n = 5). LOD and LOQ in urine after pre-column derivatization using FITC were 100 and 250 ng/ml, respectively. Urine samples were analysed by direct injection without sample pre-treatment. The mechanism of enhancement of fluorescence of the derivative by surfactant was proposed.
基金National Natural Science Foundation of China(Grant No.20635001 and 21175005)
文摘Human erythropoietin （hEPO）, an endogenous glycoprotein, plays a fundamental role in erythropoiesis controlling the formation of red blood cells. Production of recombinant human erythropoietin （rhEPO） has made it possible for its abuse in competitive sports. In this work, pre-capillary and on-capillary derivatization by 5-furoylquinoline-3-carboxaldehyde （FQ） and fluorescein isothiocyanate （FITC） for the detection of rhEPO by capillary electrophoresis with laser-induced fluorescence detection （CE-LIF） were compared. FQ pre-capillary labeling improves sensitivity but degrades the glycoforms separation due to the inhomogeneity of the reaction products from multiple labeling. Compared with FITC pre-capillary derivatization with the excess fluorescent background, the on-capillary FQ derivatization method can provide shorter analysis time, lower background, and better selectivity. It is demonstrated that, through optimizing reaction conditions of FQ on-capillary derivatization, both high sensitivity and satisfactory resolution for the analysis of the be used for the glycoforms profiling and quality control of rhEPO doping control analysis. glycoforms of rhEPO could be obtained. This method can It may be used as a candidate method for fast screening in
基金supported by the National Natural Science Foundation of China(No30470886)
文摘This work described a sensitive method for determination of metoprolol in rabbit plasma.The method involved purification by ultrafiltration,derivatization with fluorescein isothiocyanate,determination by capillary electrophoresis（CE） coupled with laser-induced fluorescence（LIF） detector.Other components in plasma including a variety of amino acids and proteins did not interfere with the determination of metoprolol in experimental condition.The assay had a wide range（2.0-500 ng/mL） of linearity and a detection limit of 0.8 ng/mL.The intra- and inter-day precisions were satisfactory with relative standard deviation（RSD） less than 10.0%and accuracy within 10.0%.This method was successfully applied to pharmacokinetic study of metoprolol in rabbit blood.
基金Project supported by The National Natural Science Foundation of China (Grant No. 29825112) and CAS (No. KJ951-A1-507).
文摘FITC labeled amino acids have been separated using a home-built capillary electrophoresis with a laser-induced fluorescence detection (CE-LIF) system. Seventeen peaks can now be generated from the twenty common amino acids. The key conditions lie in the optimization of pH, buffer electrolytes and buffer additives.
基金Supported by the Surface Project of the National Natural Science Foundation of China（No.21075039/B050106） and the Spe- cialized Research Fund for the Doctoral Program of Higher Education of China（No.20100074110016）.
文摘Proteomics is becoming more and more mature, but the detection of low abundance proteins is still a difficult task. Laser-induced fluorescence（LIF） detection is one of the most sensitive detection methods in a capillary electrophoresis（CE） system. However, most proteins do not exhibit favourable native fluorescence, a derivatization procedure is necessary for LIF detection of proteins. Since the derivatization reaction between protein and fluorescent reagent takes place after the separation of protein, the separation cannot be compromised by multiple derivatization products, the post-column derivatization becomes an attractive method for derivatization in CE-LIF system.
