Abstract
BackgroundPorcine epidemic diarrhea virus (PEDV) is a highly effective pathogen that can cause death of new-born piglet, resulting in big economical loss in pig farming industry. For rapid detection of PEDV, a new immunochromatographic assay (ICA) based on monoclonal antibodies (mAbs) was developed in this study.ResultsThe mAbs were prepared by using PEDV positive hybridoma cells that were selected by using cell surface fluorescence immunosorbent assay (CSFIA). Fourteen mAbs against PEDV strain isolated from south of China were prepared. The optimal mAb 4A11 was coated on NC membrane as the capturing reagent and the mAb A11H7 was coupled to gold nanoparticles (AuNPs) as detection reagent for the new ICA. The new ICA was used to measure PEDV in phosphate buffer containing tween-20. Results indicated that the limit of detection (LOD) of the new ICA was 0.47 μg/mL (5.9 × 103 TCID50/mL) and the liner detection range of the ICA was 0.625–10 μg/mL (7.8 × 103–105 TCID50/mL). The specificity analysis results showed that this new ICA had no cross reaction in the presence of other porcine viruses. The ICA was also validated for the detection of PEDV in swine stool samples with little interference from swine stool. To compare its accuracy to other traditional detection methods, 27 swine stool samples from south of China were investigated with the new developed ICA, commercial strip and RT-PCR. Results showed that the new ICA was more comparable to RT-PCR than commercial test strip.ConclusionsA new ICA based on mAbs prepared by CSFIA was developed in this study. It was a sensitive, specific and rapid method that could be used for on-site detection of PEDV and therefore was useful for the diagnosis and prevention of PED.
Highlights
Porcine epidemic diarrhea virus (PEDV) is a highly effective pathogen that can cause death of newborn piglet, resulting in big economical loss in pig farming industry
Materials and reagents Vero cell, porcine epidemic diarrhea virus (PEDV) strain named CHYJ130330, pseudorabies virus (PRV), classical swine fever virus (CSFV), transmissible gastroenteritis virus (TGEV) and swine stool were obtained from Guangdong Haid Institute of Animal Husbandry & Veterinary (Guangdong, China)
From the hybridoma cells generated against PEDV, 14 clones that could produce monoclonal antibodies (mAbs) with strong PEDV binding ability were selected
Summary
Porcine epidemic diarrhea virus (PEDV) is a highly effective pathogen that can cause death of newborn piglet, resulting in big economical loss in pig farming industry. Separation identification means to isolate PEDV in Vero cell cultures from the small intestine of a PEDV infected piglet It is a classical method for PEDV detection, but it is time-consuming for it needs at least one week. Recently developed immunochromatographic assay (ICA) is much more applicable for on-site diagnosis of PEDV for it is simple, sensitive, specific and can be operated without training. This method could be useful for monitoring the infection of PED [18,19,20]
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