To establish a rapid quantification method for heparinase I during its production in recombinant Escherichia coli, a translational fusion vector was constructed by fusing the N terminus of heparinase I to the C termin...To establish a rapid quantification method for heparinase I during its production in recombinant Escherichia coli, a translational fusion vector was constructed by fusing the N terminus of heparinase I to the C terminus of a green fluorescent protein mutant （GFPmutl）. As a result, not only was the functional recombinant expression of heparinase I in E. coli accomplished, but also a linear correlation was obtained between the GFP fluorescence intensity and heparinase I activity, allowing enzyme activity to be quantified rapidly during the fermentation.展开更多
Surface-enhanced Raman scattering(SERS)spectroscopy has been employed as a rapid analysis technology for food security inspection recently.Nowadays,it is still a great challenge to rapidly quantify multiple trace anti...Surface-enhanced Raman scattering(SERS)spectroscopy has been employed as a rapid analysis technology for food security inspection recently.Nowadays,it is still a great challenge to rapidly quantify multiple trace antibiotics potentially abused in aquaculture industry.In this work,a magnetic Ti_(3)C_(2)T_(x)/Fe_(3)O_(4)/Ag substrate was prepared for the development of a reliable rapid SERS quantification method for multiple trace sulfonamides in aquatic products.This magnetic substrate had good uniformity,reproducibility,stability and SERS activity.Moreover,this substrate could integrate the magnetic separation-enrichment and matrix clean-up without cross contamination,which endowed it with good selectivity and antiinterference capability during real sample analysis.The electromagnetic enhancement and chemical enhancement mechanism of this magnetic substrate were studied in detail to reveal its good separationenrichment performance and SERS activity.Finally,a rapid SERS quantification method was established and practically applied for trace phthalic sulfathiazole(PST)and silver sulfadiazine(SSD)in aquatic products by using Ti_(3)C_(2)T_(x)/Fe_(3)O_(4)/Ag magnetic substrates.Trace PST and SSD could be actually detected and quantified as 55.9μg/kg and 64.0μg/kg in aquatic products,respectively.Good recoveries of 83.9%-116%with relative standard deviations(RSDs)of 0.5%-3.2%for PST and 80.2%-102%with RSDs of 1.3%-5.8%for SSD were obtained.This work proposed an efficient and reliable method for rapid quantification of trace multiple sulfonamides in complex aquatic samples during food security inspection.展开更多
The increase in emerging and reemerging infectious diseases has underscored the need for the prompt monitoring of intact infectious viruses and the quick assessment of their infectivity.However,molecular techniques ca...The increase in emerging and reemerging infectious diseases has underscored the need for the prompt monitoring of intact infectious viruses and the quick assessment of their infectivity.However,molecular techniques cannot distinguish between intact infectious and noninfectious viruses.Here,two distinct methodologies have been developed for the expeditious and dependable quantification of intact infectious H1N1 virus,and several experiments have been conducted to substantiate their efficacy.One is an integrated cell absorption quantitative polymerase chain reaction(qPCR)method(ICA-qPCR),and the other is a combined propidium monoazide qPCR method(PMA-qPCR).The quantification limit is 100 cell culture infective dose 50%(CCID50)/mL in ICA-qPCR following a 1.5-hour cell absorption or 126 CCID50/mL after a 15-minute incubation.For PMA-qPCR,the limit was 2,512 CCID50/mL.The number of genome copies quantified by the ICA-qPCR and PMA-qPCR methods was strongly correlated with the infectious titer determined by the CCID50 assay,thereby enabling the estimation of virus infectivity.The ICA-qPCR and PMA-qPCR methods are both suitable for the identification and quantification of intact infectious H1N1 virus in inactivated samples,wastewater,and biological materials.In conclusion,the ICA-qPCR and PMA-qPCR methods have distinct advantages and disadvantages,and can be used to quantify intact infectious viruses rapidly.These methodologies can facilitate the identification of the presence of intact infectious viruses in wastewater or on pathogen-related physical surfaces in high-level biosafety laboratories and medical facilities.Furthermore,these methodologies can also be utilized to detect other highly pathogenic pathogens.展开更多
基金Supported by the National Natural Science Foundation of China (No.20336010 and No.20176025).
文摘To establish a rapid quantification method for heparinase I during its production in recombinant Escherichia coli, a translational fusion vector was constructed by fusing the N terminus of heparinase I to the C terminus of a green fluorescent protein mutant （GFPmutl）. As a result, not only was the functional recombinant expression of heparinase I in E. coli accomplished, but also a linear correlation was obtained between the GFP fluorescence intensity and heparinase I activity, allowing enzyme activity to be quantified rapidly during the fermentation.
基金financially supported by the National Natural Science Foundation of China(Nos.22074161 and 21976213)Guangdong Basic and Applied Basic Research Foundation(No.2019A1515010107)+3 种基金the Science and Technology Planning Project of Guangzhou City(No.202102080167)the Research and Development Plan for Key Areas of Food Safety in Guangdong Province of China(No.2019B020211001)the National Key Research and Development Program of China(No.2019YFC1606101)the State Key Program of National Natural Science of China(No.22134007)。
文摘Surface-enhanced Raman scattering(SERS)spectroscopy has been employed as a rapid analysis technology for food security inspection recently.Nowadays,it is still a great challenge to rapidly quantify multiple trace antibiotics potentially abused in aquaculture industry.In this work,a magnetic Ti_(3)C_(2)T_(x)/Fe_(3)O_(4)/Ag substrate was prepared for the development of a reliable rapid SERS quantification method for multiple trace sulfonamides in aquatic products.This magnetic substrate had good uniformity,reproducibility,stability and SERS activity.Moreover,this substrate could integrate the magnetic separation-enrichment and matrix clean-up without cross contamination,which endowed it with good selectivity and antiinterference capability during real sample analysis.The electromagnetic enhancement and chemical enhancement mechanism of this magnetic substrate were studied in detail to reveal its good separationenrichment performance and SERS activity.Finally,a rapid SERS quantification method was established and practically applied for trace phthalic sulfathiazole(PST)and silver sulfadiazine(SSD)in aquatic products by using Ti_(3)C_(2)T_(x)/Fe_(3)O_(4)/Ag magnetic substrates.Trace PST and SSD could be actually detected and quantified as 55.9μg/kg and 64.0μg/kg in aquatic products,respectively.Good recoveries of 83.9%-116%with relative standard deviations(RSDs)of 0.5%-3.2%for PST and 80.2%-102%with RSDs of 1.3%-5.8%for SSD were obtained.This work proposed an efficient and reliable method for rapid quantification of trace multiple sulfonamides in complex aquatic samples during food security inspection.
基金supported by Key-Area Research and Development Program of Guangdong Province,China(2022B1111010004,2021B1212040017)Basic Research Program of Guangzhou Municipal Science and Technology Bureau(SL2023A03J01074)+1 种基金Guangdong Provincial Highly Pathogenic Microorganism Science Data Center(2024B1212070013)Shenzhen Science and Technology Program(ZDSYS20230626091203007).
文摘The increase in emerging and reemerging infectious diseases has underscored the need for the prompt monitoring of intact infectious viruses and the quick assessment of their infectivity.However,molecular techniques cannot distinguish between intact infectious and noninfectious viruses.Here,two distinct methodologies have been developed for the expeditious and dependable quantification of intact infectious H1N1 virus,and several experiments have been conducted to substantiate their efficacy.One is an integrated cell absorption quantitative polymerase chain reaction(qPCR)method(ICA-qPCR),and the other is a combined propidium monoazide qPCR method(PMA-qPCR).The quantification limit is 100 cell culture infective dose 50%(CCID50)/mL in ICA-qPCR following a 1.5-hour cell absorption or 126 CCID50/mL after a 15-minute incubation.For PMA-qPCR,the limit was 2,512 CCID50/mL.The number of genome copies quantified by the ICA-qPCR and PMA-qPCR methods was strongly correlated with the infectious titer determined by the CCID50 assay,thereby enabling the estimation of virus infectivity.The ICA-qPCR and PMA-qPCR methods are both suitable for the identification and quantification of intact infectious H1N1 virus in inactivated samples,wastewater,and biological materials.In conclusion,the ICA-qPCR and PMA-qPCR methods have distinct advantages and disadvantages,and can be used to quantify intact infectious viruses rapidly.These methodologies can facilitate the identification of the presence of intact infectious viruses in wastewater or on pathogen-related physical surfaces in high-level biosafety laboratories and medical facilities.Furthermore,these methodologies can also be utilized to detect other highly pathogenic pathogens.
