基于大片吸虫分泌排泄产物层析组分的牛片形吸虫病间接ELISA诊断方法的建立

doi:10.16431/j.cnki.1671-7236.2021.08.034

Abstract

Abstract: To establish a more sensitivity, specificity diagnostic method that could be used for the early diagnosis of bovine fascioliasis, Fasciola gigantica excretory-secretory products (FgESP) was chromatographed to screen UV280 chromatographic peak.While UV280 chromatographic peak showing the best detection effect was screened out, the best chromatographic component under corresponding chromatographic peak was further screened as diagnostic antigen.The conditions of coating antigen, antibody dilution, dilution of enzyme-labeled secondary antibody and reaction time were optimized by checkerboard titration test, and the indirect ELISA diagnostic method for detection of bovine fascioliasis was established.Sensitive, specific, clinical tests and 0-14 weeks serum IgG antibody of buffalos experimentally infected Fasciola gigantica were performed on the established indirect ELISA assay.Meanwhile, serum IgG antibody of 160 buffalos were also detected, then result was compared with that of the existing diagnostic antigen (FgESP, the cut-off value was 0.320).The results showed that chromatographic component F22 worked well.The optimal reaction conditions of the indirect ELISA were as follows:The coating antigen of F22 was 0.157 μg/mL, the testing serum dilution and the HRP conjugated anti-bovine IgG dilution were 1:400 and 1:40 000 respectively, the incubation time was 25 min.The established method was used to detect 15 buffalo negative serums, while the cut-off value of D_{450 nm} was 0.408.Comparision of F22 and FgESP indicated that they were comparable in specificity, while F22 showed higher sensitivity.Upon established indirect ELISA, it was found that from the second week anti-F22-IgG was significantly higher than that of anti-FgESP-IgG in the period of 0-14 weeks infected buffalo serum (P<0.01).Therefore, F22 showed better diagnostic effect than FgESP.The positive rate of F22 was 71.25% and FgESP was 63.75% in 160 serum samples detection of buffalos.All above suggested that indirect ELISA based on F22 showed better diagnostic effect than FgESP, which could apply to early diagnosis of fascioliasis in the future.

Key words: Fasciola gigantica; diagnostic antigen; ELISA

CLC Number:

S855.9

JIN Weikun, YUAN Xiangxiang, WANG Jinhui, HOU Linjing, ZHENG Mengwei, WU Zhengjiao, ZHANG Weiyu, DI Wenda. Establishment of Indirect ELISA for Bovine Fasciolasis Diagnosis Based on Chromatographic Component of Fasciola gigantica Excretory-secretory Products[J]. China Animal Husbandry and Veterinary Medicine, 2021, 48(8): 3010-3018.

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Establishment of Indirect ELISA for Bovine Fasciolasis Diagnosis Based on Chromatographic Component of Fasciola gigantica Excretory-secretory Products

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