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目的建立快速、灵敏、特异的检测新孢子虫感染牛血清抗体的免疫胶体金试纸条。方法采用柠檬酸三钠还原法制备胶体金颗粒。分别用兔抗SPA和新孢子虫P40蛋白包被硝酸纤维素膜作为质控线和检测线进行反应条件优化,组装胶体金试纸条。对试纸条的灵敏度、特异性、重复性和保存期进行检测;利用该试纸条检测120份吉林省某养牛场牛血清中的抗新孢子虫抗体。结果兔抗SPA抗体的最佳包被浓度为0.5mg/ml,新孢子虫P40蛋白的最佳包被浓度为1mg/ml。该试纸条检测牛新孢子虫抗体的灵敏性可达到1∶320(血清稀释度);且不与牛瑟氏泰勒虫感染血清发生交叉阳性反应。用3个不同批次的试纸条进行试验,已知阴性血清及阳性血清显色情况一致。该试纸条置4℃条件下可保存60d。用该试纸条检测牛血清120份,阳性率为17.5%。结论建立的牛新孢子虫感染胶体金试纸具有检测快速、灵敏、特异性强等优点,可用于牛新孢子虫感染监测。
Objective To establish a rapid, sensitive and specific immuno-gold strip for the detection of Neospora-infected bovine serum antibody. Methods The trisodium citrate reduction method was used to prepare colloidal gold particles. Respectively with rabbit anti-SPA and Ps. Neospora P40 protein coated nitrocellulose membrane as the control line and the test line to optimize the reaction conditions, the assembly of gold colloidal gold test strips. The sensitivity, specificity, repeatability and shelf-life of test strip were tested. Anti-Neospora antibody in bovine serum of 120 dairy farms in Jilin Province was detected using the test strip. Results The optimal coating concentration of rabbit anti-SPA antibody was 0.5 mg / ml and the optimum coating concentration of P40 protein of Neospora was 1 mg / ml. The test strip detection of bovine Neospora antibody sensitivity up to 1: 320 (serum dilution); and does not cross-positive reaction with the sera of T. eutropha. With three different batches of test strips, known negative serum and positive serum color consistent. The test strip can be stored at 4 ℃ for 60d. With the test strips of bovine serum 120, the positive rate was 17.5%. Conclusion The established Neisseria bassiana infection colloidal gold test strip has the advantages of rapid, sensitive and specific detection, which can be used for the monitoring of Neospora bovine infection.