Development of a Competitive Chemiluminescent Enzyme Immunoassay Based on Recombinant FMDV VP2 Protein for Rapid and Simultaneous Detection of Antibodies Against Foot-and-Mouth Disease Virus Serotypes O and A

Figures

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  Figure 1. Optimization of FMDV-VP2 cCLEIA. (A) Screening of the optimal concentration of rVP2 protein. (B) Optimization of blocking conditions. (C) Determination of the optimal concentration of HRP-labeled FMDV monoclonal antibody (FMDV MAb-HRP). (D) Determination of incubation temperature and time. The data points represent mean values, and error bars represent standard deviation (SD). HRP = horseradish peroxidase.

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  Figure 2. Schematic illustration of the FMDV-VP2 cCLEIA for detection of FMDV antibody. Arrows indicate the sequential steps of the reaction. Step 1: FMDV VP2 antigen is immobilized on the solid phase. Step 2: FMDV-MAb-HRP and testing antibody compete for binding to VP2. Step 3: Luminol substrate is added, and the HRP-catalyzed reaction produces a chemiluminescent signal. MAb = monoclonal antibody; HRP = horseradish peroxidase.

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  Figure 3. Distribution of PI % for FMDV antibody-negative sera.

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  Figure 4. Analytical sensitivity and cross-reactivity of the FMDV-VP2 cCLEIA. (A) Analytical sensitivity of the cCLEIA. Positive when PI > 35. (B) Analytical sensitivity of the FMDV O LPBE. According to the manufacturer’s instructions, serum samples with optical density at 450 nm (OD450) equal to or greater than the cut-off value of 0.5685 are interpreted as negative for the antibody. (C) Analytical sensitivity of the FMDV A LPBE. According to the manufacturer’s instructions, serum samples with OD450 equal to or greater than the cut-off value of 0.728 are interpreted as negative for the antibody. (D) Cross-reactivity detection.