布鲁氏菌病荧光偏振抗体检测方法的建立
发布时间：2018-11-12 13:46
【摘要】：【目的】布鲁氏菌病(Brucellosis)简称布病,是由布鲁氏菌引起的以感染家畜为主的人畜共患传染病,造成严重的公共卫生问题。目前全世界范围内消除该病的主要方法是扑杀与免疫相结合,所以建立快速准确的诊断方法对防治和清除布病非常必要。本文建立布鲁氏菌病荧光偏振(FPA)抗体检测方法,为布鲁氏菌病(布病)的快速高效诊断提供技术手段。【方法】提纯猪种布鲁氏菌S2株脂多糖O链(OPS),经异硫氰酸荧光素(FITC)标记后作为诊断抗原。通过对样品稀释液、抗原稀释度、反应时间等条件的优化,初步建立了布鲁氏菌荧光偏振诊断方法。用该方法对148份布病阳性血清(其中牛血清70份,羊血清78份)和155份布病阴性血清(其中牛血清82份,羊血清73份)进行检测,确定其敏感性和特异性。按确定的技术参数,制备3批布鲁氏菌FPA抗体检测试剂盒,使用质控阴、阳性血清分别评价试剂盒的批内和批间重复性。用400份临床样本比较本研究开发试剂盒与商品化进口FPA试剂盒的符合率。【结果】使用0.5%蔗糖磷酸缓冲液作为血清样品稀释液;标记抗原的使用浓度为90μg/m L;最佳反应时间为3-5 min。本检测方法的判定标准为:δm P值20时为阴性,δm P值≥20时为阳性。按上述条件建立的FPA检测148份布病阳性血清和155份布病阴性血清,结果敏感性为98.6%,特异性为98.7%。对400份临床样本的比对检测显示,研究建立的FPA方法与进口商品化试剂盒的总符合率为94.0%。【结论】研究建立的布鲁氏菌PFA抗体检测方法具有良好的特异性和敏感性,可作为一种重要的布病诊断快速诊断方法。
[Abstract]:[objective] brucellosis (Brucellosis) is a zoonotic infectious disease caused by brucellosis, which causes serious public health problems. At present, the main method to eliminate the disease worldwide is the combination of culling and immunity, so it is necessary to establish a rapid and accurate diagnostic method for preventing and eliminating brucellosis. In this paper, a fluorescence polarization (FPA) antibody detection method for brucellosis was established, which provides a technical means for rapid and efficient diagnosis of brucellosis (brucellosis). [methods] purification of lipopolysaccharide O chain (OPS), from Brucella spp S2 strain Fluorescein isothiocyanate (FITC) was used as diagnostic antigen. Based on the optimization of sample dilution, antigen dilution and reaction time, a fluorescence polarization diagnostic method for brucella was established. This method was used to detect 148 brucellosis positive serum (70 bovine serum, 78 sheep serum) and 155 brucellosis negative serum (82 bovine serum and 73 sheep serum). The sensitivity and specificity of the method were determined. According to the determined technical parameters, three batches of brucella FPA antibody detection kits were prepared, and the intra- and inter-batch repeatability of the kits were evaluated by using quality control negative and positive serum respectively. The coincidence rate between the developed kit and the commercial FPA kit was compared with 400 clinical samples. [results] 0.5% sucrose phosphate buffer solution was used as the diluent of serum sample, and the concentration of labeled antigen was 90 渭 g / mL. The optimum reaction time is 3-5 min.. The criterion of this method is that 未 MP is negative at 20:00 and 未 MP 鈮，
本文编号：2327286
[Abstract]:[objective] brucellosis (Brucellosis) is a zoonotic infectious disease caused by brucellosis, which causes serious public health problems. At present, the main method to eliminate the disease worldwide is the combination of culling and immunity, so it is necessary to establish a rapid and accurate diagnostic method for preventing and eliminating brucellosis. In this paper, a fluorescence polarization (FPA) antibody detection method for brucellosis was established, which provides a technical means for rapid and efficient diagnosis of brucellosis (brucellosis). [methods] purification of lipopolysaccharide O chain (OPS), from Brucella spp S2 strain Fluorescein isothiocyanate (FITC) was used as diagnostic antigen. Based on the optimization of sample dilution, antigen dilution and reaction time, a fluorescence polarization diagnostic method for brucella was established. This method was used to detect 148 brucellosis positive serum (70 bovine serum, 78 sheep serum) and 155 brucellosis negative serum (82 bovine serum and 73 sheep serum). The sensitivity and specificity of the method were determined. According to the determined technical parameters, three batches of brucella FPA antibody detection kits were prepared, and the intra- and inter-batch repeatability of the kits were evaluated by using quality control negative and positive serum respectively. The coincidence rate between the developed kit and the commercial FPA kit was compared with 400 clinical samples. [results] 0.5% sucrose phosphate buffer solution was used as the diluent of serum sample, and the concentration of labeled antigen was 90 渭 g / mL. The optimum reaction time is 3-5 min.. The criterion of this method is that 未 MP is negative at 20:00 and 未 MP 鈮，
本文编号：2327286
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