[Objective] To develop an indirect sandwich ELISA for rapid detection of duck swollen head septicemia virus （DSHSV）. [Method] DSHSV hyperimmune sara were prepared in ducks and rabbits by inoculation with DSHSV XD st...[Objective] To develop an indirect sandwich ELISA for rapid detection of duck swollen head septicemia virus （DSHSV）. [Method] DSHSV hyperimmune sara were prepared in ducks and rabbits by inoculation with DSHSV XD strain and then purified using saturated ammonium sulfate and Sephadex G-150 column chromatography to obtain anti-DSHSV IgG. An indirect sandwich ELISA was developed using the purified duck anti-DSHSV IgG and rabbit anti-DSHSV IgG after reaction conditions were optimized. Its specificity, sensitivity and repeatability were evaluated, and its accuracy was confirmed by observation with immunoelectron microscopy. Then, distribution of DSHSV in tissues of challenged ducks was also detected. [Result] Through optimizing conditions, the ELISA was developed. Only DSHSV could be detected by the developed method, but other pathogens could not be detected. Compared with agar gel diffusion test, the developed method was more sensitive. The coefficient of variation was less than 10%, and the developed method had good repeatability. In addition, the ELISA-positive samples contained DSHSV, as confirmed by im- munoelectron microscopy. All heart, liver, lung and kidney collected from the DSHSV-challenged ducks were positive when they were detected by the developed ELISA. [ Conclusion] The developed ELISA method is rapid, simple, specific and sensitive, and it is suitable for large-scale quaran- tine of DSHS. Heart, liver, lunq and kidney should be selected preferentially as specimens for diaclnosis of DSHS.展开更多
基金supported by Program for Changjiang Scholars and Innovative Research Team in University (IRT0848)
文摘[Objective] To develop an indirect sandwich ELISA for rapid detection of duck swollen head septicemia virus （DSHSV）. [Method] DSHSV hyperimmune sara were prepared in ducks and rabbits by inoculation with DSHSV XD strain and then purified using saturated ammonium sulfate and Sephadex G-150 column chromatography to obtain anti-DSHSV IgG. An indirect sandwich ELISA was developed using the purified duck anti-DSHSV IgG and rabbit anti-DSHSV IgG after reaction conditions were optimized. Its specificity, sensitivity and repeatability were evaluated, and its accuracy was confirmed by observation with immunoelectron microscopy. Then, distribution of DSHSV in tissues of challenged ducks was also detected. [Result] Through optimizing conditions, the ELISA was developed. Only DSHSV could be detected by the developed method, but other pathogens could not be detected. Compared with agar gel diffusion test, the developed method was more sensitive. The coefficient of variation was less than 10%, and the developed method had good repeatability. In addition, the ELISA-positive samples contained DSHSV, as confirmed by im- munoelectron microscopy. All heart, liver, lung and kidney collected from the DSHSV-challenged ducks were positive when they were detected by the developed ELISA. [ Conclusion] The developed ELISA method is rapid, simple, specific and sensitive, and it is suitable for large-scale quaran- tine of DSHS. Heart, liver, lunq and kidney should be selected preferentially as specimens for diaclnosis of DSHS.
