【摘要】：布魯氏菌病嚴重危害著人類的健康和畜牧業(yè)的快速發(fā)展,它是一種動物源性、自然疫源性人獸共患病。萊姆病是一種人獸共患傳染性疾病,是由蜱傳播的若干個不同基因種的伯氏疏螺旋體引起的疾病。目的:(1)本研究基于血清學(xué)診斷方法,建立一種快速、便捷的適用于基層診斷布魯氏菌的免疫膠體金試紙條檢測方法。(2)本研究應(yīng)用免疫滲濾技術(shù),建立一種早期鑒別萊姆病的快速診斷方法。方法:(1)提取純化羊種布魯氏菌強毒株16M LPS,對其純度、濃度進行鑒定后,以LPS作為包被抗原,優(yōu)化完善膠體金免疫層析技術(shù)以及試紙條的制作條件,完成試紙條的組裝后,對不同動物、人類血清樣品進行檢測,并與虎紅平板凝集試驗、試管凝集試驗結(jié)果進行對比,分析以16M LPS作為包被抗原的試紙條的可行性。(2)使用石河子大學(xué)人獸共患病實驗室構(gòu)建的萊姆重組蛋白Bmp A,經(jīng)過純化、濃縮,達到最佳包被濃度后,優(yōu)化免疫滲濾技術(shù)的條件,使用免疫滲濾技術(shù)對疑似萊姆患者血清及未知綿羊血清進行檢測,通過檢測結(jié)果,探究萊姆膜脂蛋白Bmp A作為包被抗原應(yīng)用免疫滲濾技術(shù),對萊姆病診斷的可行性。結(jié)果:(1)免疫膠體金試紙條LPS最佳包被濃度為2.0 mg/m L。440份血清樣品檢測結(jié)果為:試紙條與試管凝集試驗的符合率為95.45%,明顯高于虎紅平板凝集試驗與試管凝集試驗的符合率(79.55%);且試紙條的陽性檢出率為84.32%,試管凝集試驗陽性檢出率為86.59%,相差較小。(2)重組蛋白Bmp A以2.0 mg/m L為制作免疫滲濾卡盒的最適包被量,用免疫滲濾卡對8份確定感染萊姆病患者的血清進行檢測,檢測出3份陽性血清;對30份的綿羊血清進行檢測,檢測出2份陽性血清,經(jīng)過WB驗證結(jié)果一致。結(jié)論:本研究建立了以LPS作為包被抗原的布魯氏菌病膠體金試紙條診斷方法,可用于基層對布魯氏菌病的初步篩查;優(yōu)化免疫滲濾技術(shù),建立了以純化的Bmp A萊姆病伯氏疏螺旋體膜脂蛋白作為包被抗原的萊姆病免疫滲濾卡診斷方法。為布魯氏菌病和萊姆病的臨床診斷提供了技術(shù)支持。
[Abstract]:Brucellosis seriously endangers human health and the rapid development of animal husbandry, it is an animal-derived, natural epidemic zoonosis. Lyme disease is a zoonotic infectious disease caused by several tick-borne spirochetes. Objective: (1) based on the method of serological diagnosis, this study established a rapid and convenient method for the detection of immuno-colloidal gold strip for the diagnosis of brucella. To establish a rapid diagnostic method for early differential diagnosis of Lyme disease. Methods: (1) the purity and concentration of 16m LPS, were extracted and purified from Brucella virulent strain of sheep. LPS was used as the coating antigen to optimize the preparation conditions of colloidal gold immunochromatography and test strip. After the assembly of the test strip, different animal and human serum samples were detected, and the results were compared with the results of the Tiger Red plate agglutination test and the test tube agglutination test. The feasibility of using 16m LPS as the test strip of coated antigen was analyzed. (2) the recombinant protein Bmp A was constructed by the laboratory of zoonoses of Shihezi University. After purification and concentration, the recombinant protein Bmp A was obtained to the best coating concentration. The condition of immune filtration technique was optimized, and the serum of suspected Lyme patients and unknown sheep serum were detected by using immunofiltration technique. Through the test results, the application of immunofiltration technique was studied as the coating antigen of Lim membrane lipoprotein Bmp A. The feasibility of diagnosis of Lyme disease. Results: (1) the best coating concentration of LPS was 2.0 mg/m L. 440 serum samples. The results showed that the coincidence rate between the test strip and the test tube agglutination test was 95.45%. The coincidence rate of Tiger red plate agglutination test and test tube agglutination test (79.55%) was higher than that of Tiger red plate agglutination test (79.55%). The positive rate of the test strip was 84.32 and the positive rate of the test tube agglutination test was 86.59, the difference was small. (2) the recombinant protein Bmp A used 2.0 mg/m L as the optimal envelope for making the immune leachate card box. Eight sera from patients with Lyme disease were detected with immune leachate and 3 positive sera were detected. Two positive sera were detected from 30 sheep sera, and the results were consistent by WB. Conclusion: in this study, a method for the diagnosis of brucellosis colloidal gold strip with LPS as the coating antigen was established, which can be used for the primary screening of brucellosis at the basic level. A method for the diagnosis of Bmp A Lyme disease by using purified membrane lipoprotein of Borrelia burgdorferi as antigen was established by optimizing immunofiltration technique. It provides technical support for the clinical diagnosis of brucellosis and Lyme disease.
【學(xué)位授予單位】：石河子大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2015
【分類號】：S855

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本文編號：2395136