Preparation of a highly specific and sensitive monoclonal antibody against tilmicosin and its application in lateral flow immunoassay  

作　　者：Youyi Wang Qingyue Li Guanghua Liang Huayun Li Zizhe Li Tian Gao Lianjun Song Xianqing Huang Dapeng Peng Xiya Zhang 

机构地区：[1]Henan Engineering Technology Research Center of Food Processing and Circulation Safety Control,College of Food Science and Technology,Henan Agricultural University,63 Nongye Road,Zhengzhou 450002,Henan,China [2]Development Center of Livestock and Veterinary,Fuling District,No.26 Liming North Road,Chongqing 408000,China [3]Henan Province Measurement Standard and Product Quality Inspection and Testing Center,Guancheng District,10 White South Road,Zhengzhou 450000,Henan,China [4]National Reference Laboratory of Veterinary Drug Residues,Huazhong Agricultural University,Wuhan 430070,Hubei,China

出　　处：《One Health Advances》2023年第1期1-9,共9页全健康进展（英文）

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);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).

摘　　要：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.

关 键 词：Hapten design Monoclonal antibody High sensitivity TILMICOSIN Lateral flow immunoassay 

分 类 号：Q78[生物学—分子生物学]