Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/13326
標題: 製備抗家禽流行性感冒病毒H6亞型血球凝集素之單源抗體
Produce Monoclonal Antibodies of the Hemagglutinin of Avian Influenza Virus H6
作者: 施雨華
Shih, Yu-Hau
關鍵字: Avian Influenza Virus;家禽流行性感冒病毒;Hemagglutinin;Monoclonal Antibodies;血球凝集素;單源抗體
出版社: 獸醫學系暨研究所
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摘要: 
近年來陸續有高病原性H5N1血清亞型之家禽流行性感冒病毒感染人類的病例,由2003年至2007年6月12日止,已造成190人不幸死亡,經由分析後推斷病毒是由已感染的禽鳥類直接傳染給人類。家禽流行性感冒病毒除了造成嚴重之經濟損失外,更有人畜共通疾病之隱憂。臺灣於1972年第一次爆發家禽流行性感冒病毒H6N1後,1998年開始監控水禽和家禽疫情,於家禽血清監控中發現蛋雞有50%而土雞有30%呈現H6N1血清陽性。雖然臺灣目前是高病原性家禽流行性感冒病毒非疫區,但是低病原性的H6N1仍然會造成養禽業的經濟損失,而且也由於長期於蛋雞和肉雞循環感染,因此也增加了抗原變異的可能。為求有效的監控此病毒,本研究利用H6N1血清亞型的純化病毒免疫BALB/c小鼠,以E.coli原核表現的H6 Hemagglutinin(HA)蛋白來篩選抗H6血清亞型HA的單源抗體。單源抗體經純化後,分析其特異性、敏感度以及中和能力,在特異性除了可以辨識H6 HA表現蛋白以及病毒尿囊液,其他1-14型血清亞型之HA 表現蛋白以及其他RNA病毒均無法辨識,證明具有良好的特異性。而以dot-enzyme-linked immunosorbent assay來評估單源抗體7G9的敏感度,對於表現的HA蛋白敏感度為0.24ng,對於病毒尿囊液的敏感度為0.5HAU,敏感度較HA試驗為敏感。利用中和試驗來檢驗單源抗體的中和能力,結果此單源抗體不具有效的中和能力,推測其辨識點並非在病毒的抗原決定位上。因此本實驗所製備的單源抗體可以用來開發診斷試劑,用於臨床樣本的抗原、抗體檢測,有利於家禽流行性感冒的監控。

In recent years, a highly pathogenic avian influenza virus H5N1 caused disease in human; from 2003 to 2007 June 12 led 190 people die unfortunately. Nucleotide sequence analysis showed that this virus was transmitted directly from infected avian species to humans. Avian influenza viruses not only cause serious economic losses, there is a worry of the disease becoming zoonoses. The first H6N1 avian influenza viruses outbreak in Taiwan was reported in 1972. Since 1998 began surveillance in waterfowl and poultry, and a recent serologic surveillance in poultry showed that about 50% of the layers and 30% of the native broilers had antibodies against H6N1 viruses. Although no highly pathogenic avian influenza virus has been isolated in Taiwan, low pathogenic H6N1 AIV was still made economic loss. H6N1 viruses have long period of circulation time in layers and boilers may increase chances to antigenic drift. In order to surveillance this virus, the study used purified H6N1 virus to immune BALB/c mice and screened by hemagglutinin(HA) protein of H6 expression from E. coli. After purification, we analyzed specificity, sensitivity and neutralizing efficiency. The specificity result is that the monoclonal antibodies (MAb)could recognized the HA protein expression from E. coli and homogeneous H6N1 virus. In sensitivity, we evaluated by dot-enzyme-linked immunosorbent assay. The monoclonal antibodies can detect 0.24 ng HA protein expression from E. coli and 0.5HAU virus. It is more sensitivity than HA test. Using neutralization test to evaluate the MAb's neutralizing efficiency, and the result is that MAb don't have effective neutralize efficiency. May the MAb's epitop can not recognize the antigenic site of H6N1 virus. In this study, the MAb we produced can be used to develop new detection tool to examine antigen or antibody of clinical samples for AIV surveillance.
URI: http://hdl.handle.net/11455/13326
其他識別: U0005-0208200714243100
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