The method of immunohistochemistry assay for the detection of scrapie in Chinese Short-tailed Han sheep was established using monoclonal antibody. Genomic DNA was isolated from Chinese Short-tailed Han sheep blood. Using the polymerase chain reaction technique, PrP27-30 gene sequence was amplified from Chinese Short-tailed Han sheep genomic DNA. By recombinant DNA technology, the recombinant protein of Chinese Short-tailed Han sheep PrP27-30 was obtained. Then, using standard methodology of myeloma cell fusion, a panel of monoclonal antibodies was generated. With mAbs, scrapie in Chinese Short-tailed Han sheep was detected by immunohistochemistry assay. The recombinant protein of Chinese Short-tailed Han sheep PrP27-30 was obtained and a panel of six hybridoma cell lines secreting specific antibodies to Chinese Short-tailed Han sheep PrP27-30 related to scrapie was obtained with one fusion between myeloma Sp2/0 and spleen cells from mice immunized with the purified recombinant protein. Four hybridoma cell lines can be used in immunohistochemistry assay for the detection of scrapie in Chinese Short-tailed Han sheep. So that the special monoclonal antibody developed in author’s institute can be used to detect PrPSc of scrapie in Chinese Short-tailed Han sheep by immunohistochemistry in China.
给家兔灌服低剂量(2.5 m g/kg体质量)和中剂量(5.0 m g/kg体质量)的甲胺磷,在灌药前和灌药后1、6、12、24、48 h分别采血,检测血清胆碱酯酶(ChE)、超氧化物歧化酶(SOD)、乳酸脱氢酶(LDH)和碱性磷酸酶(AKP)活性及丙二醛(M DA)含量的动态变化。结果表明:(1)中剂量组和低剂量组分别于灌药后45 m in和60 m in出现明显的中毒症状;(2)与灌药前相比,2组各时间相血清ChE活性均显著降低(P<0.01),呈现先抑制后逐渐恢复的变化趋势,在中毒后12 h,血清ChE活性降至最低,中剂量组和低剂量组分别为灌药前的35%和42%;至48 h时,血清ChE活性仍明显低于灌药前(P<0.01);(3)血清SOD活性降低(P<0.05或P<0.01),M DA含量显著高于灌药前(P<0.05或P<0.01);(4)血清LDH和AKP活性逐渐升高,与灌药前相比差异极显著(P<0.05或P<0.01);(5)灌药后不同时间组间各项检测指标比较,均有显著差异(P<0.05或P<0.01)。提示家兔甲胺磷急性中毒的症状与ChE抑制程度不一致,影响中毒的关键是ChE活性降低的速度,且在中毒过程中机体遭受脂质过氧化损伤。
