China Animal Husbandry and Veterinary Medicine ›› 2022, Vol. 49 ›› Issue (10): 3953-3962.doi: 10.16431/j.cnki.1671-7236.2022.10.028

• Preventive Veterinary Medicine • Previous Articles     Next Articles

Optimization of Dual Strip for FMDV Immunized Antibody Evaluating and Discriminating Vaccinated Animals from Infected Animals

YANG Suzhen1, SUN Yaning1, LIU Yunchao1, CHAI Shujun1, ZHANG Gaiping1,2,3   

  1. 1. Key Laboratory of Animal Immunology, Henan Academy of Agricultural Sciences, Zhengzhou 450002, China;
    2. College of Animal Science and Veterinary Medicine, Henan Agricultural University, Zhengzhou 450002, China;
    3. Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou 225009, China
  • Received:2022-04-22 Online:2022-10-05 Published:2022-09-30

Abstract: 【Objective】 The purpose of this study was to establish a stable production process of Foot-and-mouth disease virus(FMDV) infection and immune differential diagnosis and immune evaluation test paper,and promote its industrial production and clinical application.【Method】 In this study,the colloidal gold immunoprobe was combined with the epitope polypeptide coupling carrier protein of porcine IgG,FMDV structural protein (SP) and non structural protein (NSP) to prepare artificial antigen,and two detection lines were set to accurately intercept the detection mode of antibody. Taking the specificity and sensitivity of the test strip as the evaluation index,the parameters such as protein for colloidal gold immunoprobe,polypeptide antigen for test strip interception,detection line position,spray film buffer,gold standard protein preservation solution,sample pad buffer and sample diluent were optimized.The optimized test paper was used to detect 266 field pig serum samples,and the results were compared with the results of foot-and-mouth disease O antibody liquid phase blocking ELISA (LPB ELISA) and 3ABC blocking ELISA antibody detection kit (3ABC ELISA).The coincidence rate between the test paper and commercial ELISA kit was calculated.【Result】 After optimization,the parameters of the dual test strip for differential diagnosis of FMDV infection and immunity and immune evaluation were as follows:Staphylococcus aureus A protein (SPA) labeled with colloidal gold was used as immunoprobe,the mixed BSA-peptides were used as test lines.BSA-NSPs and BSA-SPs were dispensed on NC membrane as the test line 1 and test line 2,respectively.0.1 mol/L Tris-HCl was used as membrane spraying buffer.ddH2O containing 15 mg/mL BSA,13.25 mg/mL Na2HPO4·12H2O,15.9 mg/mL NaH2PO4·2H2O,10% Trition X-100 and 0.3 mg/mL NaN3 were used as gold standard protein preservation solution.0.02 mol/L Na2B4O7·10H2O containing 10 mg/mL casein,5% TritionX-100,0.3 mg/mL NaN3 were used as sample pad buffer.Normal saline containing 1.0% Tween-20 were used as sample dilution.After optimization,the coincidence rate of the optimized strip with FMDV 3ABC ELISA and FMDV LPB ELISA was 96.20% and 94.36%,respectively.【Conclusion】 By optimizing the production process,a stable production technology was developed.The color of the test lines of the optimized strip were clearly visible with high recognition by the naked eyes.The specificity and sensitivity of the strip were markedly improved.This study laid the basis for the mass production and industrial application of the strip and provided a stable,specific,sensitive and accurate method for FMDV detection.

Key words: Foot-and-mouth disease virus (FMDV); antibody; strip; optimization

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