In this study,two novel haptens of oxamichydrazide(OXZ),the metabolite of nifuraldezone,were designed and synthesized by derivatization with ethyl-4-bromobutanoate and ethyl-2-bromoacetate for the first time.The ratio...In this study,two novel haptens of oxamichydrazide(OXZ),the metabolite of nifuraldezone,were designed and synthesized by derivatization with ethyl-4-bromobutanoate and ethyl-2-bromoacetate for the first time.The rationality of the haptens was enlightened by conformational alignment and electronic evaluations based on computional chemistry.The synthesized haptens were efficient in generating antibody response to OXZ.A monoclonal antibody(mAb)named 2D9 with high affinity was obtained and used to establish an indirect competitive enzyme-linked immunosorbent assay(icELISA)for the direct determination of OXZ in chicken samples without the need for derivatization step.Under optimum conditions,the developed icELISA showed an IC50 value of 0.66 ng mL^(−1),and limit of detection of 0.11μg kg^(−1)in chicken.The average recoveries were found to be 80.4-91.3%with a coefficient of variation less than 9.0%.In conclusion,we have successfully desigened haptens,prepared mAbs and developed an immunoassay for the direct determination of OXZ in chicken samples without the need for a derivatization step for the first time.This provides a potentially rapid,accurate,and sensitive screening tool for nifuraldezone residue in animal-derived food.展开更多
A facile procedure was described for the hapten design of the N-methylcarbamate insecticide propoxur.Two new haptens of propoxur(hapten 1a and hapten 1b) were synthesized by introducing appropriate spacers in the pest...A facile procedure was described for the hapten design of the N-methylcarbamate insecticide propoxur.Two new haptens of propoxur(hapten 1a and hapten 1b) were synthesized by introducing appropriate spacers in the pesticide aromatic moiety of the analyte molecular structure.First,the propoxur reacted with nitric acid to yield the intermediate product.Then hapten 1a was prepared via the reduction of the intermediate product,and hapten 1b was formed by the acylation of hapten 1a with succinic anhydride.In addition,we have also developed a simple method to prepare hapten 2 for the pyrethroid insecticide cyhalothrin,which only requires two steps including the reactions of nitration and reduction.Three haptens were coupled to carrier proteins to prepare the corresponding artificial antigens.The molecular structures of these three haptens were identified by 1H NMR and MS,and those of the artificial antigens were confirmed by UV-vis.展开更多
To reduce the false positive results caused by cross reactivity of the antibodies with other structural analogues,it is crucial to prepare a high specificity and sensitivity antibody against target for developing an a...To reduce the false positive results caused by cross reactivity of the antibodies with other structural analogues,it is crucial to prepare a high specificity and sensitivity antibody against target for developing an accurate immunoassay.In this study,tilmicosin(TM)was selected as a model molecule.Firstly,two-dimensional similarity,electrostatic potential energy,mulliken atomic charges and overlapping of different haptens with TM were calculated using Gaussian 09W and Discovery studio,and the newly designed TM-HS was selected as the optimal hapten.Furthermore,a monoclonal antibody(mAb 12C8)was produced with the half maximal inhibitory concentration(IC50)of 0.36 ng/mL,and negligible cross-reactivity(CR)with other antibiotics.Finally,a lateral flow immunoassay(LFA)for the detection of TM based on amorphous carbon nanoparticles(ACNPs)labeled mAb 12C8 was developed by the reflectance value under natural light.The recoveries of TM ranged from 83.18%to 103.25%with a coefficient of variation(CV)<12.47%.The results showed that the cut-off value of TM in milk samples was 1 ng/mL,and the limits of detection(LODs)for chicken muscle,bovine muscle,porcine muscle and porcine liver samples were 5.23,5.98,6.85 and 7.31μg/kg,respectively.In addition,40 real samples were tested by the LFA,and the detection results were consisted with that of high-performance liquid chromatography-UV detector(HPLC-UV).Those results indicated that the developed LFA is an accurate and useful tool for on-site screening of TM in milk and animal tissues.展开更多
基金supported by H2020 EU-China-Safe(727864)Chinese Ministry of Science and Technology for the National Key R&D Program of China(2017YFE0110800).
文摘In this study,two novel haptens of oxamichydrazide(OXZ),the metabolite of nifuraldezone,were designed and synthesized by derivatization with ethyl-4-bromobutanoate and ethyl-2-bromoacetate for the first time.The rationality of the haptens was enlightened by conformational alignment and electronic evaluations based on computional chemistry.The synthesized haptens were efficient in generating antibody response to OXZ.A monoclonal antibody(mAb)named 2D9 with high affinity was obtained and used to establish an indirect competitive enzyme-linked immunosorbent assay(icELISA)for the direct determination of OXZ in chicken samples without the need for derivatization step.Under optimum conditions,the developed icELISA showed an IC50 value of 0.66 ng mL^(−1),and limit of detection of 0.11μg kg^(−1)in chicken.The average recoveries were found to be 80.4-91.3%with a coefficient of variation less than 9.0%.In conclusion,we have successfully desigened haptens,prepared mAbs and developed an immunoassay for the direct determination of OXZ in chicken samples without the need for a derivatization step for the first time.This provides a potentially rapid,accurate,and sensitive screening tool for nifuraldezone residue in animal-derived food.
基金supported by the National High Technology Research and Development Program of China(2006AA10Z449)the National Natural Science Foundation of China(20801024)Wu Jieping Medical Foundation(32067500615)
文摘A facile procedure was described for the hapten design of the N-methylcarbamate insecticide propoxur.Two new haptens of propoxur(hapten 1a and hapten 1b) were synthesized by introducing appropriate spacers in the pesticide aromatic moiety of the analyte molecular structure.First,the propoxur reacted with nitric acid to yield the intermediate product.Then hapten 1a was prepared via the reduction of the intermediate product,and hapten 1b was formed by the acylation of hapten 1a with succinic anhydride.In addition,we have also developed a simple method to prepare hapten 2 for the pyrethroid insecticide cyhalothrin,which only requires two steps including the reactions of nitration and reduction.Three haptens were coupled to carrier proteins to prepare the corresponding artificial antigens.The molecular structures of these three haptens were identified by 1H NMR and MS,and those of the artificial antigens were confirmed by UV-vis.
基金supported by the Key Scientific and Technological Project of Henan Provincial Department of China(222102310162)the National Natural Science Foundation of China(32172298)+2 种基金the Young Talents Project of Henan Agricultural University(30501305)the Henan Postgraduate Joint Training Base Project(YJS2022JD16)the Program for Innovative Research Team(in Science and Technology),University of Henan Province(NO.23IRTSTHN023).
文摘To reduce the false positive results caused by cross reactivity of the antibodies with other structural analogues,it is crucial to prepare a high specificity and sensitivity antibody against target for developing an accurate immunoassay.In this study,tilmicosin(TM)was selected as a model molecule.Firstly,two-dimensional similarity,electrostatic potential energy,mulliken atomic charges and overlapping of different haptens with TM were calculated using Gaussian 09W and Discovery studio,and the newly designed TM-HS was selected as the optimal hapten.Furthermore,a monoclonal antibody(mAb 12C8)was produced with the half maximal inhibitory concentration(IC50)of 0.36 ng/mL,and negligible cross-reactivity(CR)with other antibiotics.Finally,a lateral flow immunoassay(LFA)for the detection of TM based on amorphous carbon nanoparticles(ACNPs)labeled mAb 12C8 was developed by the reflectance value under natural light.The recoveries of TM ranged from 83.18%to 103.25%with a coefficient of variation(CV)<12.47%.The results showed that the cut-off value of TM in milk samples was 1 ng/mL,and the limits of detection(LODs)for chicken muscle,bovine muscle,porcine muscle and porcine liver samples were 5.23,5.98,6.85 and 7.31μg/kg,respectively.In addition,40 real samples were tested by the LFA,and the detection results were consisted with that of high-performance liquid chromatography-UV detector(HPLC-UV).Those results indicated that the developed LFA is an accurate and useful tool for on-site screening of TM in milk and animal tissues.
