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2016 Volume 44 Issue 8
2016, 44(8): 1157-1164
doi: 10.11895/j.issn.0253-3820.151031
Abstract:
A molecularly imprinted film (MIF) was prepared on the surface plasmon resonance (SPR) sensor chip for the detection of testosterone. The nanometer imprinted film was synthesized by surface grafting approach. First, the gold sensor chip was modified with an iniferter of benzyl diethyldithiocarbamate to create the "grafting from" polymeric surface. Grafting of the MIF onto the SPR chip was subsequently achieved through ultraviolet light-initiated copolymerization of methacrylic acid (functional monomer) and ethylene glycol dimethacrylate (crosslinker) in the presence of testosterone as a model template. The template molecules were then removed to form a MIF with specific recognition sites for testosterone. With this iniferter technique, self-aggregation in the reaction solution phase was avoided and grafting polymerization was confined to the exterior of the chip surface. The grafting process was implemented by in situ monitoring with SPR, which permitted the thickness of the film to be easily and strictly controlled. The chip surface modified with a testosterone-imprinted film was characterized by the methods of polarization modulation infrared reflection absorption spectroscopy (PM-IRRAS) and atomic force microscopy (AFM). According to the results, testosterone-imprinted film modification of the SPR chip was achieved by the distribution of numerous homogeneous, nanoscale holes on the surface. The testosterone-imprinted film was applied on a SPR sensor chip for the detection of testosterone in the range of 2.5×10-16 to 2.5×10-6 mol/L in acetonitrile with the lowest determining concentration of 2.5×10-16 mol/L. Adsorption experiments showed that there were two kinds of binding sites in the MIF, and the linear correlations between the changes in reflectivity and the concentration of testosterone were y=19.69+1.21x(R2=0.9913) and y=11.5+0.45x(R2=0.9895), respectively. Control experiments utilizing estradiol, estriol, and progesterone as analogues showed impressive selectivity and specificity for testosterone determination. The testosterone-imprinted film reproducibility was evaluated with five cycles of rinsing-rebinding and the RSD was 16.8% and 11.2% for the SPR angle changes in rinsing and rebinding respectively, demonstrating that the MIF-modified SPR sensor had good reproducibility and repeatability. Finally, the SPR sensor was successfully used to determine testosterone in artificial urine samples with recoveries from 85.2% to 92.8%.
A molecularly imprinted film (MIF) was prepared on the surface plasmon resonance (SPR) sensor chip for the detection of testosterone. The nanometer imprinted film was synthesized by surface grafting approach. First, the gold sensor chip was modified with an iniferter of benzyl diethyldithiocarbamate to create the "grafting from" polymeric surface. Grafting of the MIF onto the SPR chip was subsequently achieved through ultraviolet light-initiated copolymerization of methacrylic acid (functional monomer) and ethylene glycol dimethacrylate (crosslinker) in the presence of testosterone as a model template. The template molecules were then removed to form a MIF with specific recognition sites for testosterone. With this iniferter technique, self-aggregation in the reaction solution phase was avoided and grafting polymerization was confined to the exterior of the chip surface. The grafting process was implemented by in situ monitoring with SPR, which permitted the thickness of the film to be easily and strictly controlled. The chip surface modified with a testosterone-imprinted film was characterized by the methods of polarization modulation infrared reflection absorption spectroscopy (PM-IRRAS) and atomic force microscopy (AFM). According to the results, testosterone-imprinted film modification of the SPR chip was achieved by the distribution of numerous homogeneous, nanoscale holes on the surface. The testosterone-imprinted film was applied on a SPR sensor chip for the detection of testosterone in the range of 2.5×10-16 to 2.5×10-6 mol/L in acetonitrile with the lowest determining concentration of 2.5×10-16 mol/L. Adsorption experiments showed that there were two kinds of binding sites in the MIF, and the linear correlations between the changes in reflectivity and the concentration of testosterone were y=19.69+1.21x(R2=0.9913) and y=11.5+0.45x(R2=0.9895), respectively. Control experiments utilizing estradiol, estriol, and progesterone as analogues showed impressive selectivity and specificity for testosterone determination. The testosterone-imprinted film reproducibility was evaluated with five cycles of rinsing-rebinding and the RSD was 16.8% and 11.2% for the SPR angle changes in rinsing and rebinding respectively, demonstrating that the MIF-modified SPR sensor had good reproducibility and repeatability. Finally, the SPR sensor was successfully used to determine testosterone in artificial urine samples with recoveries from 85.2% to 92.8%.
2016, 44(8): 1165-1170
doi: 10.11895/j.issn.0253-3820.160103
Abstract:
Gram-negative (G-) bacteria, such as denitrifying bacteria and anaerobic ammonia oxidation bacteria, are highly social organisms capable of sophisticated cooperative behavior mediated via quorum sensing. As signal molecules of the chemical communication, N-acyl-homoserine lactones (AHLs) can mediate the quorum sensing of the functional microbial population and regulate the population density. To understand the growth of functional microbial population and the mechanism for biological nitrogen removal in upflow microaerobic sludge reactors (UMSRs) treating organic wastewater with low ratio of chemical oxygen demand to total nitrogen, a method was established to simultaneously detect AHLs in the microaerobic processes. Water-sludge mixtures sampled from the UMSRs were pretreated in sequence by liquid-liquid extraction using ethyl acetate, rotary evaporation, constant volume with methanol, separation by C18 column. Gradient elution was carried out using 5 mmol/L ammonium acetate (containing 0.1% formic acid) and methanol as mobile phases. On the base of multiple reaction monitoring analysis, a triple quadrupole mass spectrometer with an electrospray ionization was introduced to detect the target compounds. Nine kinds of AHLs were used to evaluate the established method and the results showed that the detection limits were 0.01-0.5 μg/L and all of the AHLs presented excellent linearity with the concentration ranging from 0.5 to 100 μg/L. The recovery and relative standard deviation ranged from 62.5% to 118.1% and 2.9% to 12.1%, respectively. The analysis could be finished within 6.5 min. The rapid, accurate and precise method for detecting AHLs provided a new insight into the growth and metabolic activity of functional microbial population in the activated sludge processes to understand the mechanism of biological nitrogen removal, suggesting a good application in regulation and operation of wastewater biological treatment processes.
Gram-negative (G-) bacteria, such as denitrifying bacteria and anaerobic ammonia oxidation bacteria, are highly social organisms capable of sophisticated cooperative behavior mediated via quorum sensing. As signal molecules of the chemical communication, N-acyl-homoserine lactones (AHLs) can mediate the quorum sensing of the functional microbial population and regulate the population density. To understand the growth of functional microbial population and the mechanism for biological nitrogen removal in upflow microaerobic sludge reactors (UMSRs) treating organic wastewater with low ratio of chemical oxygen demand to total nitrogen, a method was established to simultaneously detect AHLs in the microaerobic processes. Water-sludge mixtures sampled from the UMSRs were pretreated in sequence by liquid-liquid extraction using ethyl acetate, rotary evaporation, constant volume with methanol, separation by C18 column. Gradient elution was carried out using 5 mmol/L ammonium acetate (containing 0.1% formic acid) and methanol as mobile phases. On the base of multiple reaction monitoring analysis, a triple quadrupole mass spectrometer with an electrospray ionization was introduced to detect the target compounds. Nine kinds of AHLs were used to evaluate the established method and the results showed that the detection limits were 0.01-0.5 μg/L and all of the AHLs presented excellent linearity with the concentration ranging from 0.5 to 100 μg/L. The recovery and relative standard deviation ranged from 62.5% to 118.1% and 2.9% to 12.1%, respectively. The analysis could be finished within 6.5 min. The rapid, accurate and precise method for detecting AHLs provided a new insight into the growth and metabolic activity of functional microbial population in the activated sludge processes to understand the mechanism of biological nitrogen removal, suggesting a good application in regulation and operation of wastewater biological treatment processes.
2016, 44(8): 1171-1177
doi: 10.11895/j.issn.0253-3820.160102
Abstract:
Mitochondria play a central role in the regulation of energy metabolism and signal transduction in eukaryotic cells. Although many fluorescent labeling strategies have been developed for mitochondrial studies, the methods that enable labeling efficiency assessment at the single-mitochondrion level are still lacking. By employing the unique advantages of high sensitivity flow cytometry (HSFCM) in the sensitive, rapid, and quantitative multiparameter analysis of individual mitochondria, here we examined the performance of several different mitochondrial labeling strategies from the perspectives of brightness, labeling ratio, and stability. Mitochondria isolated from HeLa cells transfected with pAcGFP1-Mito plasmid upon transient or stable transfections, and mitochondria directly labeled with MitoTracker Green or SYTO 62 were analyzed by a laboratory-built three-channel HSFCM. Upon the quantitative measurement of fluorescence brightness, it was found that the fluorescence intensity of green fluorescent protein (GFP) in mitochondria isolated from cells with stable transfection was about 17.7-fold higher than the transient transfection ones, and was approximately two orders of magnitude brighter than mitochondria labeled with MitoTracker Green. On the other hand, the fluorescence signal of SYTO 62 labeling decreased upon washing, indicating its rapid dissociation rate. The strong fluorescence intensity and good labeling stability make stable transfection an efficient method to label mitochondria. The experimental results demonstrates that HSFCM provides a powerful analytical tool to assess the performance of mitochondrial fluorescence labeling via high throughput single mitochondria analysis.
Mitochondria play a central role in the regulation of energy metabolism and signal transduction in eukaryotic cells. Although many fluorescent labeling strategies have been developed for mitochondrial studies, the methods that enable labeling efficiency assessment at the single-mitochondrion level are still lacking. By employing the unique advantages of high sensitivity flow cytometry (HSFCM) in the sensitive, rapid, and quantitative multiparameter analysis of individual mitochondria, here we examined the performance of several different mitochondrial labeling strategies from the perspectives of brightness, labeling ratio, and stability. Mitochondria isolated from HeLa cells transfected with pAcGFP1-Mito plasmid upon transient or stable transfections, and mitochondria directly labeled with MitoTracker Green or SYTO 62 were analyzed by a laboratory-built three-channel HSFCM. Upon the quantitative measurement of fluorescence brightness, it was found that the fluorescence intensity of green fluorescent protein (GFP) in mitochondria isolated from cells with stable transfection was about 17.7-fold higher than the transient transfection ones, and was approximately two orders of magnitude brighter than mitochondria labeled with MitoTracker Green. On the other hand, the fluorescence signal of SYTO 62 labeling decreased upon washing, indicating its rapid dissociation rate. The strong fluorescence intensity and good labeling stability make stable transfection an efficient method to label mitochondria. The experimental results demonstrates that HSFCM provides a powerful analytical tool to assess the performance of mitochondrial fluorescence labeling via high throughput single mitochondria analysis.
2016, 44(8): 1178-1184
doi: 10.11895/j.issn.0253-3820.160148
Abstract:
A rapid gas chromatographic (GC) method was established for the determination of short-chain fatty acids (SCFAs) in human feces. Feces samples were directly extracted by 1% HCl-75% ethanol solution, and then centrifuged at high speed for GC analysis. The chromatographic separation was performed on a DB-FFAP capillary column (30 m×0.25 mm×0.25 μm) with a temperature program (initial temperature at 50℃ held for 1 min, ramped to 190℃ at 10℃/min). The injection port temperature was 250℃ with split ratio at 50:1. The carrier gas was high purity nitrogen with a constant linear velocity at 1.0 mL/min. A flame ionization detector was employed to quantify SCFAs. The proposed method had been certified by systematic method validation, and used to determine feces samples. Subsequently, the health volunteer and colorectal cancer patient groups could be distinguished successfully by multivariate statistical analysis. Compared with health volunteers, the acetic acid and butyric acid of feces from colorectal cancer patients were reduced obviously, indicating that SCFAs particularly butyric acid could be considered as candidate markers for colorectal cancer diagnosis. In summary, this study provides a rapid method for the determination of SCFAs in feces form health volunteers and colorectal cancer patients. The method had a prospect for screening and diagnosing colorectal cancer rapidly.
A rapid gas chromatographic (GC) method was established for the determination of short-chain fatty acids (SCFAs) in human feces. Feces samples were directly extracted by 1% HCl-75% ethanol solution, and then centrifuged at high speed for GC analysis. The chromatographic separation was performed on a DB-FFAP capillary column (30 m×0.25 mm×0.25 μm) with a temperature program (initial temperature at 50℃ held for 1 min, ramped to 190℃ at 10℃/min). The injection port temperature was 250℃ with split ratio at 50:1. The carrier gas was high purity nitrogen with a constant linear velocity at 1.0 mL/min. A flame ionization detector was employed to quantify SCFAs. The proposed method had been certified by systematic method validation, and used to determine feces samples. Subsequently, the health volunteer and colorectal cancer patient groups could be distinguished successfully by multivariate statistical analysis. Compared with health volunteers, the acetic acid and butyric acid of feces from colorectal cancer patients were reduced obviously, indicating that SCFAs particularly butyric acid could be considered as candidate markers for colorectal cancer diagnosis. In summary, this study provides a rapid method for the determination of SCFAs in feces form health volunteers and colorectal cancer patients. The method had a prospect for screening and diagnosing colorectal cancer rapidly.
2016, 44(8): 1185-1192
doi: 10.11895/j.issn.0253-3820.160038
Abstract:
Ion mobility spectrometry (IMS) based explosives detectors is wieldy deployed at the check points of airport, etc, so the development of new ionization method to replace the radioactive 63Ni ion source in IMS is highly demanded. In this study, a novel negative corona discharge was developed for ion mobility spectrometer for rapid detection of trace explosives, which was running in the unidirectional mode to efficiently remove nitrogen oxides and ozone produced from the discharge process. The diameter of target electrode was 3 mm, the distance between the needle and the target electrode was 2 mm, the discharge voltage was 2400 V and the flow rate of drift gas was 1200 mL/min. Under the optimized conditions, the dominant reactant ions was O2-(H2O)n and could be used to directly detect the common explosives, such as trinitrotoluene (TNT), ammonium nitrate (AN), nitroglycerin (NG), pentaerythritol tetranitrate (PETN) and hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX). The limit of detection of TNT was 200 pg/μL. These results indicated that negative corona discharge ionization source, with the advantanges of the high sensibility, simple structure and no radioactivity, could be used as a potential and promising ionization source for ion mobility spectrometry to detect explosives.
Ion mobility spectrometry (IMS) based explosives detectors is wieldy deployed at the check points of airport, etc, so the development of new ionization method to replace the radioactive 63Ni ion source in IMS is highly demanded. In this study, a novel negative corona discharge was developed for ion mobility spectrometer for rapid detection of trace explosives, which was running in the unidirectional mode to efficiently remove nitrogen oxides and ozone produced from the discharge process. The diameter of target electrode was 3 mm, the distance between the needle and the target electrode was 2 mm, the discharge voltage was 2400 V and the flow rate of drift gas was 1200 mL/min. Under the optimized conditions, the dominant reactant ions was O2-(H2O)n and could be used to directly detect the common explosives, such as trinitrotoluene (TNT), ammonium nitrate (AN), nitroglycerin (NG), pentaerythritol tetranitrate (PETN) and hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX). The limit of detection of TNT was 200 pg/μL. These results indicated that negative corona discharge ionization source, with the advantanges of the high sensibility, simple structure and no radioactivity, could be used as a potential and promising ionization source for ion mobility spectrometry to detect explosives.
2016, 44(8): 1193-1199
doi: 10.11895/j.issn.0253-3820.160139
Abstract:
Formaldehyde has been widely employed to immobilize clinical tissue specimens, inactivate toxins and viruses in biomedical fields. Formaldehyde can react with active groups in bio-molecules such as proteins, resulting in protein cross-linking, inactivation, and immobilization. By using several standard peptides and tryptic peptides from matrix protein of influenza virus as experimental models, we studied the chemical modifications of peptides and proteins with formaldehyde by matrix-assisted laser desorption ionization time-of-flight mass spectrometry and nano-electrospray quadruple time-of-flight tandem mass spectrometry. The reaction between formaldehyde and peptides was performed under the same conditions as those during inactivation of virus (4℃, 0.025% Formalin (V/V), 37% formaldehyde solution (w/w), and 72 h). The results indicated that under above conditions, formaldehyde could react with amino group of N-terminus of standard peptide to generate a methylol adduct, which was further condensed into an imine to generate +12 Da product. Besides, formaldehyde could react with side chain of two amino acids such as arginine and lysine, yielding +12 Da product respectively. The analysis of the reaction between formaldehyde and tryptic peptides from matrix protein of influenza virus showed that +24 Da products could be detected in most peptides due to combinational contribution from N-terminus of peptide (+12 Da) and side chain of C-terminal arginine or lysine (+12 Da). Moreover, a +36 Da product was detected for a peptide with miss-cut site. The results indicated that low-concentration formaldehyde primarily reacted with amino group on N-termini of peptides and proteins, as well as the side chains of arginine and lysine residues. The present study suggested an effective mass spectrometry-based method for analyzing the reaction between low-concentration formaldehyde and peptides and proteins, thus provided strategies for interpretation for the mass spectra of reaction products.
Formaldehyde has been widely employed to immobilize clinical tissue specimens, inactivate toxins and viruses in biomedical fields. Formaldehyde can react with active groups in bio-molecules such as proteins, resulting in protein cross-linking, inactivation, and immobilization. By using several standard peptides and tryptic peptides from matrix protein of influenza virus as experimental models, we studied the chemical modifications of peptides and proteins with formaldehyde by matrix-assisted laser desorption ionization time-of-flight mass spectrometry and nano-electrospray quadruple time-of-flight tandem mass spectrometry. The reaction between formaldehyde and peptides was performed under the same conditions as those during inactivation of virus (4℃, 0.025% Formalin (V/V), 37% formaldehyde solution (w/w), and 72 h). The results indicated that under above conditions, formaldehyde could react with amino group of N-terminus of standard peptide to generate a methylol adduct, which was further condensed into an imine to generate +12 Da product. Besides, formaldehyde could react with side chain of two amino acids such as arginine and lysine, yielding +12 Da product respectively. The analysis of the reaction between formaldehyde and tryptic peptides from matrix protein of influenza virus showed that +24 Da products could be detected in most peptides due to combinational contribution from N-terminus of peptide (+12 Da) and side chain of C-terminal arginine or lysine (+12 Da). Moreover, a +36 Da product was detected for a peptide with miss-cut site. The results indicated that low-concentration formaldehyde primarily reacted with amino group on N-termini of peptides and proteins, as well as the side chains of arginine and lysine residues. The present study suggested an effective mass spectrometry-based method for analyzing the reaction between low-concentration formaldehyde and peptides and proteins, thus provided strategies for interpretation for the mass spectra of reaction products.
2016, 44(8): 1200-1208
doi: 10.11895/j.issn.0253-3820.160047
Abstract:
A chiral separation and residue determination method for cis-epoxiconazole enantiomers in apple, grape and tea samples was developed and validated by ultra performance convergence chromatography combined with quadrupole time-of-flight mass spectrometry (UPC2-QTOF/MS). The Chrial CCA column was used to separate cis-epoxiconazole enantiomers and the chromatography conditions (mobile phase modifier and proportion, column temperature, automated backpressure regulator, and auxiliary solvent) were optimized. Samples were extracted by acetonitrile, and respectively purified by Cleanert TPT or Pesti-Carb solid phase extraction (SPE) columns, then analyzed by UPC2-QTOF/MS. The optimum conditions were as follows:mobile phase was CO2/isopropanol(95:5, V/V), flow-rate was 2.0 mL/min, automated backpressure regulator (ABPR) was 13.79 MPa, column temperature was 30℃, with a post-column mauxiliary solvent of methanol/water (1:1, V/V) containing 2 mmol/L ammonium formate. The analyte was quantified by matrix external standard method. The results showed that linear range of this method was 0.01-1.00 mg/L, and the correlation coefficients were above 0.99. The recoveries of cis-epoxiconazole enantiomers at three spiked levels (0.005, 0.025 and 0.25 mg/kg) in fruit matrix were 67.9%-92.8% with relative standard deviations (RSDs, n=6) less than 10%, and the limit of quantification (LOQ) of enantiomers was 0.005 mg/kg. The recoveries of cis-epoxiconazole enantiomers at three spiked levels (0.01, 0.05 and 0.5 mg/kg) in black tea were 74.1%-84.0% with RSDs (n=6) less than 8%, and the LOQ for these two enantiomers was 0.01 mg/kg. This method is rapid, convenient and reliable, and could meet the requirement of residue analysis.
A chiral separation and residue determination method for cis-epoxiconazole enantiomers in apple, grape and tea samples was developed and validated by ultra performance convergence chromatography combined with quadrupole time-of-flight mass spectrometry (UPC2-QTOF/MS). The Chrial CCA column was used to separate cis-epoxiconazole enantiomers and the chromatography conditions (mobile phase modifier and proportion, column temperature, automated backpressure regulator, and auxiliary solvent) were optimized. Samples were extracted by acetonitrile, and respectively purified by Cleanert TPT or Pesti-Carb solid phase extraction (SPE) columns, then analyzed by UPC2-QTOF/MS. The optimum conditions were as follows:mobile phase was CO2/isopropanol(95:5, V/V), flow-rate was 2.0 mL/min, automated backpressure regulator (ABPR) was 13.79 MPa, column temperature was 30℃, with a post-column mauxiliary solvent of methanol/water (1:1, V/V) containing 2 mmol/L ammonium formate. The analyte was quantified by matrix external standard method. The results showed that linear range of this method was 0.01-1.00 mg/L, and the correlation coefficients were above 0.99. The recoveries of cis-epoxiconazole enantiomers at three spiked levels (0.005, 0.025 and 0.25 mg/kg) in fruit matrix were 67.9%-92.8% with relative standard deviations (RSDs, n=6) less than 10%, and the limit of quantification (LOQ) of enantiomers was 0.005 mg/kg. The recoveries of cis-epoxiconazole enantiomers at three spiked levels (0.01, 0.05 and 0.5 mg/kg) in black tea were 74.1%-84.0% with RSDs (n=6) less than 8%, and the LOQ for these two enantiomers was 0.01 mg/kg. This method is rapid, convenient and reliable, and could meet the requirement of residue analysis.
2016, 44(8): 1209-1214
doi: 10.11895/j.issn.0253-3820.150937
Abstract:
Eight mouse hybridoma cell lines which stably secreted monoclonal antibodies (McAbs) against human prostate-specific antigen-α1-antichymotrypsin complex (PSA-ACT) were obtained through hybridoma technique. After purification, the immunological characters of 8 McAbs were identified and classified by epitopes analysis through indirect enzyme-linked immunosorbent assay (ELISA). A pair of McAbs was chosen from above 8 McAbs, based on which a highly sensitive, simple and rapid chemiluminescence enzyme immunoassay (CLEIA) was developed for determination of PSA-ACT in human serums using the lumino-H2O2 reaction catalyzed by horseradish peroxidase (HRP) as the chemiluminescence system. Several experiment factors such as coating buffer, coating concentration, dilution ratio of PSA-ACT-HRP complex, incubation time, immunoreaction protocol and chemiluminescence reaction time were optimized. The results showed that the linear range of the proposed method for PSA-ACT determination was 0-40 ng/mL (R2=0.9943), with the detection limit of 0.53 ng/mL. The inter-assay relative standard deviations (RSDs) were 4.6%-6.6%, and intra-assay RSDs were 5.7%-8.0%.The recoveries of PSA-ACT at three spiked levels in serum samples were 95.4%-104.2%. The proposed method exhibited a cross-reactivity of 0.6% with free-PSA. The proposed method is stable, sensitive, rapid and simple, and provides a foundation for the development of PSA-ACT CLEIA kit and shows great value in clinical auxiliary diagnosis of prostate cancer.
Eight mouse hybridoma cell lines which stably secreted monoclonal antibodies (McAbs) against human prostate-specific antigen-α1-antichymotrypsin complex (PSA-ACT) were obtained through hybridoma technique. After purification, the immunological characters of 8 McAbs were identified and classified by epitopes analysis through indirect enzyme-linked immunosorbent assay (ELISA). A pair of McAbs was chosen from above 8 McAbs, based on which a highly sensitive, simple and rapid chemiluminescence enzyme immunoassay (CLEIA) was developed for determination of PSA-ACT in human serums using the lumino-H2O2 reaction catalyzed by horseradish peroxidase (HRP) as the chemiluminescence system. Several experiment factors such as coating buffer, coating concentration, dilution ratio of PSA-ACT-HRP complex, incubation time, immunoreaction protocol and chemiluminescence reaction time were optimized. The results showed that the linear range of the proposed method for PSA-ACT determination was 0-40 ng/mL (R2=0.9943), with the detection limit of 0.53 ng/mL. The inter-assay relative standard deviations (RSDs) were 4.6%-6.6%, and intra-assay RSDs were 5.7%-8.0%.The recoveries of PSA-ACT at three spiked levels in serum samples were 95.4%-104.2%. The proposed method exhibited a cross-reactivity of 0.6% with free-PSA. The proposed method is stable, sensitive, rapid and simple, and provides a foundation for the development of PSA-ACT CLEIA kit and shows great value in clinical auxiliary diagnosis of prostate cancer.
2016, 44(8): 1215-1220
doi: 10.11895/j.issn.0253-3820.160143
Abstract:
Atomic force microscopy (AFM) is one of important tools for studying the structure and function of biomolecules due to its nanoscale capability. A well prepared sample was the primary key issue for high-resolution imaging macromolecules. Here, by using self-assembly technology of DNA origami, antigens were modified on DNA origami. Afterward, antibodies reacted specifically with antigens by a molecular recognition way. Then, a nanostructure formed from DNA origami and antigen-antibody complexes. By the aid of adsorption character of DNA origami, individual antibodies could selectively adsorb on mica surface. As a result, the ultrastructural morphology image of a single IgG molecule, the characteristic "Y-shaped" domains, was resolved in liquid. The way for adsorbing biomolecules on the mica surface in a liquid environment was simple and convenient, and it may be useful to detect and measure biomoleculers at the singe-molecule level with AFM.
Atomic force microscopy (AFM) is one of important tools for studying the structure and function of biomolecules due to its nanoscale capability. A well prepared sample was the primary key issue for high-resolution imaging macromolecules. Here, by using self-assembly technology of DNA origami, antigens were modified on DNA origami. Afterward, antibodies reacted specifically with antigens by a molecular recognition way. Then, a nanostructure formed from DNA origami and antigen-antibody complexes. By the aid of adsorption character of DNA origami, individual antibodies could selectively adsorb on mica surface. As a result, the ultrastructural morphology image of a single IgG molecule, the characteristic "Y-shaped" domains, was resolved in liquid. The way for adsorbing biomolecules on the mica surface in a liquid environment was simple and convenient, and it may be useful to detect and measure biomoleculers at the singe-molecule level with AFM.
2016, 44(8): 1221-1226
doi: 10.11895/j.issn.0253-3820.160053
Abstract:
Rapid detection and classification of bacteria colonies (Escherichia coli, Listeria monocytogens and Staphylococcus aureus) were investigated by using hyperspectral imaging. The hyperspectral reflectance images (390-1040 nm) of bacterial colonies on agar plates were collected. Bacterial spectra were extracted automatically based on the masks produced by segmenting a band difference image using the OTSU method. Full wavelength and simplified PLS-DA models were established for classification of bacterial colonies. For the full wavelength model, the overall correct classification rate (OCCR) and confident OCCR for the prediction set were 100% and 95.9%, respectively. Besides, competitive adaptive reweighted sampling (CARS), genetic algorithm (GA) and least angle regression-least absolute shrinkage and selection operator (LARS-Lasso) were used to select feature wavelengths for the development of simplified models. Among them, the CARS-model outperformed the other two in terms of precision, stability and classification accuracy with OCCR and confident OCCR of 100% and 98.0% for the prediction set, respectively. It was demonstrated that hyperspectral imaging was an effective technology for nondestructive detection of bacterial colonies with high accuracy and high speed. The allocated feature wavelengths by CARS could lay theoretical basis for developing low cost multispectral imaging systems for bacterial colony detection.
Rapid detection and classification of bacteria colonies (Escherichia coli, Listeria monocytogens and Staphylococcus aureus) were investigated by using hyperspectral imaging. The hyperspectral reflectance images (390-1040 nm) of bacterial colonies on agar plates were collected. Bacterial spectra were extracted automatically based on the masks produced by segmenting a band difference image using the OTSU method. Full wavelength and simplified PLS-DA models were established for classification of bacterial colonies. For the full wavelength model, the overall correct classification rate (OCCR) and confident OCCR for the prediction set were 100% and 95.9%, respectively. Besides, competitive adaptive reweighted sampling (CARS), genetic algorithm (GA) and least angle regression-least absolute shrinkage and selection operator (LARS-Lasso) were used to select feature wavelengths for the development of simplified models. Among them, the CARS-model outperformed the other two in terms of precision, stability and classification accuracy with OCCR and confident OCCR of 100% and 98.0% for the prediction set, respectively. It was demonstrated that hyperspectral imaging was an effective technology for nondestructive detection of bacterial colonies with high accuracy and high speed. The allocated feature wavelengths by CARS could lay theoretical basis for developing low cost multispectral imaging systems for bacterial colony detection.
2016, 44(8): 1227-1235
doi: 10.11895/j.issn.0253-3820.150938
Abstract:
A method based on dispersive solid phase extraction-high performance liquid chromatography-tandem mass spectrometry for determination of 19 kinds of nonprotein nitrogen compounds including melamine, cyromazine, amidinourea, aminotriazine, 3-aminotriazole, 4-aminotriazole, allantoin, cyanuric acid, dicyandiamide, thiourea, semicarbazide, L-leucine, L-isoleucine, L-arginine, L-hydroxyproline, L-theanine, ammeline, ammelide and guanidine in powdered formulas was presented. The nonprotein nitrogen compounds were degreased by chloroform and extracted by acetonitrile, with MgSO4 to remove water and C18 to clean up. The samples were separated on Merck ZIC HILIC column (150 mm×2.1 mm, 5 μm, 20 nm) with gradient elution. The electrospray ionization was operated in the positive mode and negative mode, and monitored by the multiple reaction monitoring (MRM) mode. Allation was quantified by external standard method and the other 18 kinds of nonprotein nitrogen compounds were quantified by internal standard method. All of the correlation coefficients (r) were higher than 0.99. The limits of quantitation (LOQ) were 0.05-5.0 mg/kg, the average recoveries were between 82.2% and 115.0%, and the relative standard deviations were less than 20%.
A method based on dispersive solid phase extraction-high performance liquid chromatography-tandem mass spectrometry for determination of 19 kinds of nonprotein nitrogen compounds including melamine, cyromazine, amidinourea, aminotriazine, 3-aminotriazole, 4-aminotriazole, allantoin, cyanuric acid, dicyandiamide, thiourea, semicarbazide, L-leucine, L-isoleucine, L-arginine, L-hydroxyproline, L-theanine, ammeline, ammelide and guanidine in powdered formulas was presented. The nonprotein nitrogen compounds were degreased by chloroform and extracted by acetonitrile, with MgSO4 to remove water and C18 to clean up. The samples were separated on Merck ZIC HILIC column (150 mm×2.1 mm, 5 μm, 20 nm) with gradient elution. The electrospray ionization was operated in the positive mode and negative mode, and monitored by the multiple reaction monitoring (MRM) mode. Allation was quantified by external standard method and the other 18 kinds of nonprotein nitrogen compounds were quantified by internal standard method. All of the correlation coefficients (r) were higher than 0.99. The limits of quantitation (LOQ) were 0.05-5.0 mg/kg, the average recoveries were between 82.2% and 115.0%, and the relative standard deviations were less than 20%.
2016, 44(8): 1236-1242
doi: 10.11895/j.issn.0253-3820.160090
Abstract:
A reliable UPLC-MS method was developed for the simultaneous determination of 4 chemicals (Sudan Ⅰ, tetrabromobisphenol A, tris(1,3-dichloroisopropyl)phosphate and tris(chlorisopropyl)Phosphate) in children products. The samples were ultrasonic extracted with acetonitrile, and then the four chemicals were separated on a C18 column in 3 min. Results showed that the limit of quantification of the method was between 5 and 500 μg/kg. The calibration curves were linear within 2-3 orders of magnitude with typical correlation coefficient above 0.9995. The recoveries ranged from 83.7% to 97.8% with three addition levels. The sensitivity, recovery and selectivity of the method could fully meet the requirements of practical work.
A reliable UPLC-MS method was developed for the simultaneous determination of 4 chemicals (Sudan Ⅰ, tetrabromobisphenol A, tris(1,3-dichloroisopropyl)phosphate and tris(chlorisopropyl)Phosphate) in children products. The samples were ultrasonic extracted with acetonitrile, and then the four chemicals were separated on a C18 column in 3 min. Results showed that the limit of quantification of the method was between 5 and 500 μg/kg. The calibration curves were linear within 2-3 orders of magnitude with typical correlation coefficient above 0.9995. The recoveries ranged from 83.7% to 97.8% with three addition levels. The sensitivity, recovery and selectivity of the method could fully meet the requirements of practical work.
2016, 44(8): 1243-1249
doi: 10.11895/j.issn.0253-3820.150999
Abstract:
The three-in-one immunoaffinity column (IAC) for the determination of aflatoxin B1 (AFB1)-zearalenone (ZEN)-deoxynivalenol (DON) was prepared with rProtein A-sepharose 4B as the column matrix. The comprehensive performance (such as nonspecific adsorption, column blank, column capacity, column efficiency and sample standard addition recovery rate) was evaluated and investigated. The results showed that, the column capacities of AFB1, ZEN, DON were 295 ng per 0.25 mL gel, 905 ng per 0.25 mL gel, 2342 ng per 0.25 mL gel, respectively, and the column blank was 0. The average recoveries of AFB1, ZEN and DON were 97.4%, 98.0% and 98.4%, respectively. By optimizing conditions, the samples were extracted using the mixture of methanol and water (80:20, V/V), and diluted with phosphate buffered saline (contain 0.1% Tween-20, PBST). The detection results of FAPAS (Food Analysis Performance Assessment Scheme) by different batch three-in-one columns were close to the target value. The prepared three-in-one immunoaffinity column which could take the place of conventional single immunoaffinity column was able to meet the requirement for treatment of food and feed samples, and lay a foundation for one step enrichment, purification and detection of multi-mycotoxins.
The three-in-one immunoaffinity column (IAC) for the determination of aflatoxin B1 (AFB1)-zearalenone (ZEN)-deoxynivalenol (DON) was prepared with rProtein A-sepharose 4B as the column matrix. The comprehensive performance (such as nonspecific adsorption, column blank, column capacity, column efficiency and sample standard addition recovery rate) was evaluated and investigated. The results showed that, the column capacities of AFB1, ZEN, DON were 295 ng per 0.25 mL gel, 905 ng per 0.25 mL gel, 2342 ng per 0.25 mL gel, respectively, and the column blank was 0. The average recoveries of AFB1, ZEN and DON were 97.4%, 98.0% and 98.4%, respectively. By optimizing conditions, the samples were extracted using the mixture of methanol and water (80:20, V/V), and diluted with phosphate buffered saline (contain 0.1% Tween-20, PBST). The detection results of FAPAS (Food Analysis Performance Assessment Scheme) by different batch three-in-one columns were close to the target value. The prepared three-in-one immunoaffinity column which could take the place of conventional single immunoaffinity column was able to meet the requirement for treatment of food and feed samples, and lay a foundation for one step enrichment, purification and detection of multi-mycotoxins.
2016, 44(8): 1250-1256
doi: 10.11895/j.issn.0253-3820.160063
Abstract:
A novel method was proposed for the quantification of nitrite ion in aqueous samples by the combination of an advanced quantitative fluorescence model with ratiometric fluorescence indicator 2,3-diaminonaphthalene. The proposed method was tentatively applied to direct quantitative determination of nitrite ion in turbid environmental water samples containing both scatterers and absorbers; and its performance was thoroughly investigated and evaluated. Experimental results showed that the proposed method could realize accurate quantitative determination of nitrite ion in environmental water samples with recovery rates in the range of 90.8%-103.0%, which were comparable to the corresponding values of HPLC-DAD experiments. The limit of detection and limit of quantification of the proposed method were estimated to be 1.9 and 5.8 nmo/L, respectively.
A novel method was proposed for the quantification of nitrite ion in aqueous samples by the combination of an advanced quantitative fluorescence model with ratiometric fluorescence indicator 2,3-diaminonaphthalene. The proposed method was tentatively applied to direct quantitative determination of nitrite ion in turbid environmental water samples containing both scatterers and absorbers; and its performance was thoroughly investigated and evaluated. Experimental results showed that the proposed method could realize accurate quantitative determination of nitrite ion in environmental water samples with recovery rates in the range of 90.8%-103.0%, which were comparable to the corresponding values of HPLC-DAD experiments. The limit of detection and limit of quantification of the proposed method were estimated to be 1.9 and 5.8 nmo/L, respectively.
2016, 44(8): 1257-1262
doi: 10.11895/j.issn.0253-3820.150841
Abstract:
A method based on photogeneration of OH radicals in water from TiO2 nanoparticles was developed to study the kinetics of oxidation of organic molecules which were used as biological antioxidants. The kinetics of oxidation of terephthalic acid as a reference probe was monitored by fluorescence measurements of the concentration of its oxidized form, 2-hydroterephthalic acid (λex=315 nm, λem=425 nm). The kinetics of oxidation of other antioxidant molecules was then deduced from the radical scavenging competition. The antioxidant properties of normal antioxidants were compared basedon this kinetic model. And the antioxidant kinetic decreased in the order:lipoic acid, gallic acid, glutathione, uric acid, vitamin C, vitamin E, trolox and bilirubin.
A method based on photogeneration of OH radicals in water from TiO2 nanoparticles was developed to study the kinetics of oxidation of organic molecules which were used as biological antioxidants. The kinetics of oxidation of terephthalic acid as a reference probe was monitored by fluorescence measurements of the concentration of its oxidized form, 2-hydroterephthalic acid (λex=315 nm, λem=425 nm). The kinetics of oxidation of other antioxidant molecules was then deduced from the radical scavenging competition. The antioxidant properties of normal antioxidants were compared basedon this kinetic model. And the antioxidant kinetic decreased in the order:lipoic acid, gallic acid, glutathione, uric acid, vitamin C, vitamin E, trolox and bilirubin.
2016, 44(8): 1263-1269
doi: 10.11895/j.issn.0253-3820.160095
Abstract:
An economic and facilely prepared dopamine (DA) sensor have been successfully fabricated by the electrodeposition of copper on single-walled carbon nanotubes (SWNTs)/Nafion-modified glassy carbon electrode. The morphology of the material was observed by scanning electron microscopy (SEM) and element composition of the material was investigated by energy dispersive X-ray spectroscopy (EDX). Tests with various scan rates and pH conditions indicated that an adsorption-controlled process occured in the electrochemical system. The mechanism of the electrode reaction of dopamine involved a two-electron process which was accompanied by a deprotonation step. Electrochemical parameters were calculated with the electron transfer number as 2.67, the charge transfer coefficients as 0.6, the apparent heterogeneous electron transfer rate constant as 1.38 s-1. Under the optimal conditions with differential pulse voltammetric measurement, the linear equation was Ipa(μA)=-0.054c (μmol/L)-3.82(R2=0.9988), with linear range of 5-100 μmol/L and detection limit of 0.0135 μmol/L (S/N=3). The main advantages of sensor included facile fabrication approach, high sensitivity, good stability and high reproducibility. The sensor was applied to the detection of DA in volunteer urine by differential pulse voltammetry with favorable recoveries of 96.5%-100.4% and relative standard deviations (RSDs) of 1.2%-2.4%.
An economic and facilely prepared dopamine (DA) sensor have been successfully fabricated by the electrodeposition of copper on single-walled carbon nanotubes (SWNTs)/Nafion-modified glassy carbon electrode. The morphology of the material was observed by scanning electron microscopy (SEM) and element composition of the material was investigated by energy dispersive X-ray spectroscopy (EDX). Tests with various scan rates and pH conditions indicated that an adsorption-controlled process occured in the electrochemical system. The mechanism of the electrode reaction of dopamine involved a two-electron process which was accompanied by a deprotonation step. Electrochemical parameters were calculated with the electron transfer number as 2.67, the charge transfer coefficients as 0.6, the apparent heterogeneous electron transfer rate constant as 1.38 s-1. Under the optimal conditions with differential pulse voltammetric measurement, the linear equation was Ipa(μA)=-0.054c (μmol/L)-3.82(R2=0.9988), with linear range of 5-100 μmol/L and detection limit of 0.0135 μmol/L (S/N=3). The main advantages of sensor included facile fabrication approach, high sensitivity, good stability and high reproducibility. The sensor was applied to the detection of DA in volunteer urine by differential pulse voltammetry with favorable recoveries of 96.5%-100.4% and relative standard deviations (RSDs) of 1.2%-2.4%.
2016, 44(8): 1270-1276
doi: 10.11895/j.issn.0253-3820.160247
Abstract:
A method for determination of trace mercury in water was established. The trace mercury in water was adsorbed quantitatively by activated carbon, and then determined by electrical pyrolysis atomic absorption spectrometry. In comparison with the detection methods of total mercury in water at present, the method avoids the steps of digestion, reduces the mercury pollution and the loss of the mercury, and is simple in operation. The effects of particle size of activated carbon, acid treatment method, acid medium and enrichment time on the enrichment efficiency were investigated. The effect of the pyrolysis temperature and the interfering ions on the determination results was investigated. Three standard addition procedures including activated carbon blank addition, solution blank addition and environmental water samples addition were studied. Regression correlation coefficients of three standard curves drawn by the three methods reached 0.9999. The slope of the three standard curves had no difference by statistical test, indicating that the determination of mercury in environmental water samples under the experiment conditions was not interfered by the coexistent elements, which showed that the activated carbon blank addition method could be directly used for preparing standard curve of the method. The water samples containing 5 ng/L and 50 ng/L mercury were determined by the method, and the relative standard deviation were 7.2% and 4.2% (n=11), respectively, with a detection limit of 1.2 ng/L. The recovery experiment was carried out after adding 10 ng/L mercury to the surface water and tap water samples, and the recoveries were between 92.0% and 103.0%. Analysis results were compared with ICP-MS as control and the deviation of the two methods were between 2.9% and 3.4%, indicating that the method was accurate and reliable, and had good precision.
A method for determination of trace mercury in water was established. The trace mercury in water was adsorbed quantitatively by activated carbon, and then determined by electrical pyrolysis atomic absorption spectrometry. In comparison with the detection methods of total mercury in water at present, the method avoids the steps of digestion, reduces the mercury pollution and the loss of the mercury, and is simple in operation. The effects of particle size of activated carbon, acid treatment method, acid medium and enrichment time on the enrichment efficiency were investigated. The effect of the pyrolysis temperature and the interfering ions on the determination results was investigated. Three standard addition procedures including activated carbon blank addition, solution blank addition and environmental water samples addition were studied. Regression correlation coefficients of three standard curves drawn by the three methods reached 0.9999. The slope of the three standard curves had no difference by statistical test, indicating that the determination of mercury in environmental water samples under the experiment conditions was not interfered by the coexistent elements, which showed that the activated carbon blank addition method could be directly used for preparing standard curve of the method. The water samples containing 5 ng/L and 50 ng/L mercury were determined by the method, and the relative standard deviation were 7.2% and 4.2% (n=11), respectively, with a detection limit of 1.2 ng/L. The recovery experiment was carried out after adding 10 ng/L mercury to the surface water and tap water samples, and the recoveries were between 92.0% and 103.0%. Analysis results were compared with ICP-MS as control and the deviation of the two methods were between 2.9% and 3.4%, indicating that the method was accurate and reliable, and had good precision.
2016, 44(8): 1277-1280
doi: 10.11895/j.issn.0253-3820.160240
Abstract:
A new method was established to determine the free inorganic cadmium ion (Cd2+) in marine bivalves using high performance liquid chromatography and inductively coupled plasma mass spectrometry (HPLC-ICP-MS). The free Cd2+ in shellfish was ultrasonic extracted for 40 min by 10 mmol/L Tris buffer solution (with 0.1 mol/L NaCl, pH=7.5), and the separation of Cd2+ was achieved using an IonPac CS5A analytical column with an IonPac CG5A guard column. The mobile phase consisted of 50 mmol/L C2H2O4 and 95 mmol/L LiOH. The new method had a good linear relationship with the correlation coefficient of 0.999 and the standard recoveries of Cd2+ were all above 84.6%. The free Cd2+ content in several kinds of marine bivalves was determined using the method and the results showed that the percentage of free inorganic Cd2+ to the total Cd content in samples with high Cd content was higher than those with low Cd content.
A new method was established to determine the free inorganic cadmium ion (Cd2+) in marine bivalves using high performance liquid chromatography and inductively coupled plasma mass spectrometry (HPLC-ICP-MS). The free Cd2+ in shellfish was ultrasonic extracted for 40 min by 10 mmol/L Tris buffer solution (with 0.1 mol/L NaCl, pH=7.5), and the separation of Cd2+ was achieved using an IonPac CS5A analytical column with an IonPac CG5A guard column. The mobile phase consisted of 50 mmol/L C2H2O4 and 95 mmol/L LiOH. The new method had a good linear relationship with the correlation coefficient of 0.999 and the standard recoveries of Cd2+ were all above 84.6%. The free Cd2+ content in several kinds of marine bivalves was determined using the method and the results showed that the percentage of free inorganic Cd2+ to the total Cd content in samples with high Cd content was higher than those with low Cd content.
2016, 44(8): 1281-1285
doi: 10.11895/j.issn.0253-3820.160049
Abstract:
A microscopic fluorescence spectrometric analysis (MFSA) system was set in the laboratory. A novel method for in situ determination of Anthracene (Ant) adsorbed onto root surface micro-zone of two kinks of mangrove plant, named Kandelia obovata (K. obovata) and Avicennia marina (A. marina) by MFSA was established. Fluorescence spectra of Ant adsorbed on root surface micro-zone were obtained by synchronous scanning mode. The signal to noise (S/N) of Ant (5.3 pg/μm2) adsorbed on K. obovata and A. marina root surface micro-zone increased up to 5.5 and 6.8 while wavelength offset (Δλ) both were at 60 nm, respectively. The linear dynamic ranges of established method were 5.3-63.2 pg/μm2 for K. obovata and 10.5-52.6 pg/μm2 for A. marina, with the detection limits of 1.1 pg/μm2 and 5.5 pg/μm2, respectively. The relative standard deviations were both less than 12.5% (n=9), and the recoveries were 98.1%-117.0% and 81.2%-110.9%, respectively. The result showed that the MFSA system had ability to obtain quantitative information of fluorescence spectra and fluorescence image of Ant adsorbed onto plant roots surface micro-zone.
A microscopic fluorescence spectrometric analysis (MFSA) system was set in the laboratory. A novel method for in situ determination of Anthracene (Ant) adsorbed onto root surface micro-zone of two kinks of mangrove plant, named Kandelia obovata (K. obovata) and Avicennia marina (A. marina) by MFSA was established. Fluorescence spectra of Ant adsorbed on root surface micro-zone were obtained by synchronous scanning mode. The signal to noise (S/N) of Ant (5.3 pg/μm2) adsorbed on K. obovata and A. marina root surface micro-zone increased up to 5.5 and 6.8 while wavelength offset (Δλ) both were at 60 nm, respectively. The linear dynamic ranges of established method were 5.3-63.2 pg/μm2 for K. obovata and 10.5-52.6 pg/μm2 for A. marina, with the detection limits of 1.1 pg/μm2 and 5.5 pg/μm2, respectively. The relative standard deviations were both less than 12.5% (n=9), and the recoveries were 98.1%-117.0% and 81.2%-110.9%, respectively. The result showed that the MFSA system had ability to obtain quantitative information of fluorescence spectra and fluorescence image of Ant adsorbed onto plant roots surface micro-zone.
2016, 44(8): 1286-1290
doi: 10.11895/j.issn.0253-3820.160192
Abstract:
To detect furaltadone metabolite 3-amino-5-morpholinomethyl-2-oxazolidinone (AMOZ) in fish sample, an Eu3+ labeling time-resolved fluoroimmunoassay (TRFIA) was developed. The effects of experimental conditions including AMOZA-OVA concentration, dilution of antibody, and reaction time on the sensitivity of TRFIA were explored. The results showed that the optimized assay conditions were as follows:the AMOZA-OVA concentration was 0.25 μg/mL; the antibody was diluted 5×104 folds, and the competitive reaction time was 50 min. Under optimal conditions, the method showed a detection limit of 0.01 ng/mL, an IC50 of 0.26 ng/mL and a linear range (IC20-IC80) of 0.025-2.83 ng/mL. The recoveries of AMOZ in fish at three spiked levels ranged from 78.0% to 86.0%, and the relative standard deviations were less than 15%. Good correlation between the ic-TRFIA and high performance liquid chromatography-tandem mass spectrometry was obtained for spiked food samples. The proposed ic-TRFIA method was suited for the determination of AMOZ residue in food samples.
To detect furaltadone metabolite 3-amino-5-morpholinomethyl-2-oxazolidinone (AMOZ) in fish sample, an Eu3+ labeling time-resolved fluoroimmunoassay (TRFIA) was developed. The effects of experimental conditions including AMOZA-OVA concentration, dilution of antibody, and reaction time on the sensitivity of TRFIA were explored. The results showed that the optimized assay conditions were as follows:the AMOZA-OVA concentration was 0.25 μg/mL; the antibody was diluted 5×104 folds, and the competitive reaction time was 50 min. Under optimal conditions, the method showed a detection limit of 0.01 ng/mL, an IC50 of 0.26 ng/mL and a linear range (IC20-IC80) of 0.025-2.83 ng/mL. The recoveries of AMOZ in fish at three spiked levels ranged from 78.0% to 86.0%, and the relative standard deviations were less than 15%. Good correlation between the ic-TRFIA and high performance liquid chromatography-tandem mass spectrometry was obtained for spiked food samples. The proposed ic-TRFIA method was suited for the determination of AMOZ residue in food samples.
2016, 44(8): 1291-1299
doi: 10.11895/j.issn.0253-3820.160054
Abstract:
Since γH2AX was firstly found in 1998, it has been one of the most important scientific topics and research tools in the related scientific fields. At present, a series of advanced testing methods and analytical technologies have been developed, which exhibited a quite attractive application prospect in the area of life science and medical science. This paper reviewed the latest progress about γH2AX in terms of molecular mechanism of phosphorylation/dephosphorylation, development of testing technologies, and the related applications.
Since γH2AX was firstly found in 1998, it has been one of the most important scientific topics and research tools in the related scientific fields. At present, a series of advanced testing methods and analytical technologies have been developed, which exhibited a quite attractive application prospect in the area of life science and medical science. This paper reviewed the latest progress about γH2AX in terms of molecular mechanism of phosphorylation/dephosphorylation, development of testing technologies, and the related applications.
2016, 44(8): 1300-1307
doi: 10.11895/j.issn.0253-3820.160074
Abstract:
Digital polymerase chain reaction (PCR) has been experiencing a rapid development during the past few years. Comparing with the traditional real-time quantitative PCR (RT-qPCR), using the same primer and probe, the accuracy for the absolute quantification of target gene is significantly improved. The development of digital PCR is directly related to the development of microfluidics. The integrated fluid circuit is an early combination of the microfluidics and digital PCR, which has a complicated fabrication process with high cost. Recently, researchers are trying to apply the droplet microfluidics in digital PCR, and the droplet microfluidic chip is able to generate millions of droplets within a short time. Each of these droplets containing no more than one target gene is a reaction chamber during the amplification process. After amplification, each droplet is tested to achieve the absolute quantification of the target gene. This paper reviews the recent progresses of droplet digital PCR, and the applications of droplet digital PCR in biological, medical and environmental fields.
Digital polymerase chain reaction (PCR) has been experiencing a rapid development during the past few years. Comparing with the traditional real-time quantitative PCR (RT-qPCR), using the same primer and probe, the accuracy for the absolute quantification of target gene is significantly improved. The development of digital PCR is directly related to the development of microfluidics. The integrated fluid circuit is an early combination of the microfluidics and digital PCR, which has a complicated fabrication process with high cost. Recently, researchers are trying to apply the droplet microfluidics in digital PCR, and the droplet microfluidic chip is able to generate millions of droplets within a short time. Each of these droplets containing no more than one target gene is a reaction chamber during the amplification process. After amplification, each droplet is tested to achieve the absolute quantification of the target gene. This paper reviews the recent progresses of droplet digital PCR, and the applications of droplet digital PCR in biological, medical and environmental fields.
2016, 44(8): 1308-1313
doi: 10.11895/j.issn.0253-3820.160219
Abstract:
We carried out parallel design and development of two differently structured auto sampler based on a multi-axis and multi-mode high-precision closed-loop servo control system. An integrated embedded control drive module was developed based on the idea of compatibility and inter-changeability, so that DC motor and encoder were standardized into uniform models. Meanwhile, electric and mechanical interfaces were uniformed to a same standard. This allows the direct exchange of above-mentioned components between the two models. A 1-μL manual sample injection syringe was installed on both standard 110-sample and platform 40-sample liquid auto sample injectors connected with gas chromatographer. Approximately 0.5 μL of cetane-isooctyl was sampled for 6 consecutive times at six different positions in the sample bottle. The repeatability RSDs of the injection peak areas of the two systems were 1.1% and 1.5%, respectively. A linear correlation coefficient (0.9947) of peak area with injection volume was achieved based on the gradient sampling volume of 0.1, 0.3, 0.5, 0.7 and 0.9 μL.
We carried out parallel design and development of two differently structured auto sampler based on a multi-axis and multi-mode high-precision closed-loop servo control system. An integrated embedded control drive module was developed based on the idea of compatibility and inter-changeability, so that DC motor and encoder were standardized into uniform models. Meanwhile, electric and mechanical interfaces were uniformed to a same standard. This allows the direct exchange of above-mentioned components between the two models. A 1-μL manual sample injection syringe was installed on both standard 110-sample and platform 40-sample liquid auto sample injectors connected with gas chromatographer. Approximately 0.5 μL of cetane-isooctyl was sampled for 6 consecutive times at six different positions in the sample bottle. The repeatability RSDs of the injection peak areas of the two systems were 1.1% and 1.5%, respectively. A linear correlation coefficient (0.9947) of peak area with injection volume was achieved based on the gradient sampling volume of 0.1, 0.3, 0.5, 0.7 and 0.9 μL.
