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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
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dc.contributor.advisor | 陳媺玫 | |
dc.contributor.author | Mei-Ling Chen | en |
dc.contributor.author | 陳玫伶 | zh_TW |
dc.date.accessioned | 2021-06-08T05:04:03Z | - |
dc.date.copyright | 2011-02-20 | |
dc.date.issued | 2011 | |
dc.date.submitted | 2011-02-13 | |
dc.identifier.citation | 劉馨淳。錦鯉疱疹病毒分離技術之建立及高敏感性診斷技術之開發。國立台灣大學獸醫專業學院獸醫學研究所碩士論文。2009。
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dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/23566 | - |
dc.description.abstract | 錦鯉疱疹病毒 (koi herpesvirus, KHV) 為一新興起的魚類病毒性疾病,會造成錦鯉 (Cyprinus carpio koi) 及鯉魚 (Cyprinus carpio carpio) 80~100% 的死亡率。此病毒性疾病具有季節性、在水溫18-28℃間最具感受性。KHV基因體為線性、具有雙股DNA且其大小約為295 kbp。經親源性分析結果,KHV目前已被歸類至Alloherpesviridae下之鯉科疱疹病毒第三型 (Cyprinid herpesvieus 3, CyHV-3)。KHV於1998年最初發生於以色列,我國則於2002年12月出現首例。許多文獻指出耐過KHV感染的錦鯉可能會成為帶原KHV並長時間表現潛伏性感染的帶原者。這些帶原者可能會在季節變化時,將體內潛在的KHV傳播給健康的錦鯉或新進的魚隻,導致高死亡率及大規模的疾病爆發。本實驗為了解潛在性的帶原者,將收集各養殖場錦鯉血清,先進行免疫點墨法的初步篩檢,再另外作KHV間接型ELISA的建立,最佳化後的陽性陰性比顯現可有效區分陽性及陰性的血清,並用以測試錦鯉體內是否有抗體的產生。同時再配合血清中和試驗,比較兩種抗體力價的結果,發現同一血清之ELISA及中和力價並非成正比的關係。我們依據各血清兩種抗體力價的表現可分成三組,分別為ELISA與中和力價皆高、ELISA低而中和力價高以及ELISA力價高而中和力價低的組別,我們從中選擇具代表性的血清來作西方墨點法的分析,可見不同組別的血清,其蛋白質表現皆有所不同。在純化病毒的抗原上,中和力價高的血清傾向於較大的蛋白質表現,而ELISA力價高的血清則傾向於有較小的蛋白質訊號。 | zh_TW |
dc.description.abstract | Koi herpesvirus (KHV), is a newly recognized virus associated with 80~100% mortality in common (Cyprinus carpio carpio) and koi (Cyprinus carpio koi) carp. This disease is seasonal, afflicting fish when the water temperature is 18 to 28℃. The genome of KHV comprises a linear, double-stranded DNA sequence of 295 kbp. Phylogenetic analysis of the KHV genome sequence led to its classification in the new family Alloherpesviridae under the species name Cyprinid herpesvirus 3. KHV was found firstly in Israel in 1998, and the first case in Taiwan has been identified in December 2002. Some reports indicated that koi survived from KHV infection could be the carrier harboring KHV, presenting latent infection for a long period. These suspected carriers could spread KHV to naive koi or new fish through the seasonal change, cause high mortality and large scale of illness. To understand the titer of antibody in serum of these potential carriers, dot blot assay can be used to primarily screen the koi serum collected from fisheries, optimize the reagents, then established an indirect ELISA method for the diagnosis of KHV. Our results revealed good positive-to-negative ratio (P/N ratio) that could effectively differentiate the positive from negative serum and detect the titer of antibodies in koi. Comparison of serum neutralization (SN) test and ELISA value indicated that the titers of ELISA and serum neutralization were not positive correlation. Based on the correlation of two kind of titers, sera could be seperated to three groups as high ELISA and SN titer, low ELISA but high SN titer and high ELISA but low SN titer as well. We choose one most remarkable serum from each groups to analyze by western blot assay, and the protein patterns of each groups were different. Serum with high SN titer tended to express strong signals to larger proteins while serum with high ELISA titer express most protein signals to smaller proteins on purified KHV virion. | en |
dc.description.provenance | Made available in DSpace on 2021-06-08T05:04:03Z (GMT). No. of bitstreams: 1 ntu-100-R97629024-1.pdf: 1990280 bytes, checksum: 33f3771f767343ec51576edbc264fce4 (MD5) Previous issue date: 2011 | en |
dc.description.tableofcontents | 致謝 I
摘要 II Abstract III 目錄 V 表目錄 VIII 圖目錄 IX 第一章 緒論 1 第二章 文獻回顧 4 第一節 錦鯉疱疹病毒之簡介 4 1.1 分類學及病名由來 4 1.2 病毒顆粒 (virion) 之結構與特徵 5 1.3 病毒之物理化學特性 5 1.4 錦鯉疱疹病毒的形成與複製機制 6 1.5 病毒潛伏與再發 11 第二節 魚類的免疫學 12 2.1 魚類的免疫器官 12 2.2 魚類對疾病入侵的體液性免疫反應 12 第三節 酵素連結免疫吸附法 (Enzyme-linked immunosorbent assays) 18 3.1 ELISA原理及應用 18 3.2 ELISA的分類 19 3.3 KHV於ELISA之應用 20 第四節 血清抗體中和效力 21 4.1 中和作用的原理 21 4.2 補體在疱疹病毒中和作用的角色 22 4.3 血清中和試驗於血清學診斷上之應用 22 第三章 材料及方法 24 第一節 實驗設計及流程 24 第二節 血清樣本收集 25 第三節 病毒純化及定量 25 3.1 實驗材料 25 3.2 病毒增殖 26 3.3 檢測病毒核酸 26 3.4 病毒定量 28 3.5 病毒濃縮 29 3.6 病毒純化 29 3.7 負染色 30 3.8 病毒蛋白質定量 30 第四節 免疫點墨法 31 4.1 實驗材料 31 4.2 實驗方法 31 第五節 發展塗鍍KHV全病毒之間接型ELISA 33 5.1 實驗材料 33 5.2 最佳化間接型ELISA條件 33 5.3 間接型ELISA操作流程 34 5.4 OD450值分析 35 第六節 病毒中和試驗 36 6.1 實驗方法 36 第七節 聚丙烯醯胺膠片電泳(SDS-PAGE)及西方墨點法 37 7.1 聚丙烯醯胺膠片電泳(SDS-PAGE) 37 7.2 蛋白質轉印法 41 7.3 西方墨點法 42 第四章 實驗結果 44 第一節 血清樣本收集 44 第二節 病毒純化及定量 44 2.1 病毒定量 44 2.2 病毒純化 44 2.3 純化病毒蛋白質定量 45 2.4 負染色 45 第三節 免疫點墨法 45 第四節 發展塗鍍KHV全病毒之間接型ELISA 46 4.1 最佳化間接型ELISA條件 46 4.2 間接型ELISA之cut-off值分析 46 4.3 以間接型ELISA偵測收集血清 46 第五節 病毒中和試驗 47 第六節 聚丙烯醯胺膠片電泳 (SDS-PAGE)與西方墨點法 47 第五章 討論 49 第一節 病毒純化 49 第二節 以免疫點墨法做血清之初步篩檢 50 第三節 以間接型ELISA測定抗體力價 52 第四節 血清抗體中和力價 57 第五節 血清抗體ELISA力價與中和力價之關聯性 60 第六節 不同力價血清之蛋白質分析 61 第六章 參考文獻 83 | |
dc.language.iso | zh-TW | |
dc.title | 錦鯉疱疹病毒抗體力價與中和效力之相關性分析 | zh_TW |
dc.title | Analysis the Correlation between ELISA Titer and Neutralizing Effect of Antibody of Koi Herpesvirus | en |
dc.type | Thesis | |
dc.date.schoolyear | 99-1 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 張本恆,涂堅,謝嘉裕 | |
dc.subject.keyword | 抗體,間接型酵素連結免疫吸附法,錦鯉疱,疹病毒,中和力價, | zh_TW |
dc.subject.keyword | antibody,ELISA,koi herpesvirus,neutralizing, | en |
dc.relation.page | 89 | |
dc.rights.note | 未授權 | |
dc.date.accepted | 2011-02-14 | |
dc.contributor.author-college | 獸醫專業學院 | zh_TW |
dc.contributor.author-dept | 獸醫學研究所 | zh_TW |
顯示於系所單位: | 獸醫學系 |
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