Taipei Medical University Institutional Repository:Item 987654321/6061
English  |  正體中文  |  简体中文  |  Items with full text/Total items : 45073/58249 (77%)
Visitors : 2390044      Online Users : 160
RC Version 7.0 © Powered By DSPACE, MIT. Enhanced by NTU Library IR team.
Scope Tips:
  • please add "double quotation mark" for query phrases to get precise results
  • please goto advance search for comprehansive author search
    •  Adv. Search
    Home ‧ Login ‧ Upload ‧ Help ‧ About ‧ Administer Goto mobile version


    Please use this identifier to cite or link to this item: http://libir.tmu.edu.tw/handle/987654321/6061


    Title: 利用噬菌體重組基因體庫製造SARS-CoV核殼蛋白抗原之雞抗體片段
    Chicken single chain variable fragment recognizing SARS-CoV Nucleocapsid protein antigen using phage display antibody technology
    Authors: 王凱玄
    Kai-Hsuan Wang
    Contributors: 細胞及分子生物研究所
    Keywords: 嗜菌體展示
    急性呼吸道症候群
    免疫蛋黃球蛋白
    phage display
    SARS
    IgY
    Date: 2007
    Issue Date: 2009-09-11 15:51:54 (UTC+8)
    Abstract: 嚴重急性呼吸道症候群(severe acute respiratory syndrome, SARS)在2003年在世界各地造成重大傷亡，目前已知為新型冠狀病毒SARS-CoV所引起，而研究顯示病毒顆粒中的核殼蛋白(nucleocapsid; N protein)可以幫助病毒入侵宿主細胞，造成感染。所以快速檢驗出以及有效的治療方法是必須的。本篇研究目標是利用重組表現五個不同長度片斷的核殼蛋白來免疫雞隻，並藉由噬菌體展現技術 (phage display technique) 來建立重組抗體基因庫 (antibody library) 篩選對於核殼蛋白具有特異性結合力的單株抗體片段 (scFv, single-chain variable fragment)。此實驗使用pGEX質體，分別在大腸桿菌BL-21細胞中表現五個病毒核殼蛋白(nucleocapsid protein) 的DNA片段。利用GST SepharoseTM 4B做純化後，可以利用coomassie blue染色的聚丙醯氨膠體電泳分析以及西方墨點法 (western blot) 來辨認蛋白。而之後將進一步將純化後的GST- nucleocapsid片段蛋白與佐劑混合均勻，以皮下注射方式注入實驗雞隻大腿部位，每週一次、連續四週。純化雞蛋內的多株抗體 (poly-IgY)，並利用酵素連結免疫吸附分析法（enzyme-linked immunosorbent assay, ELISA）以及西方墨點法來確認免疫過後雞隻所誘發出的抗體可辨認GST-nucleocaspsid片段蛋白。並以嗜菌體展現（phage display）技術，找尋出對於核殼蛋白可辨認具有特異性結合力的單株抗體片段 (scFv, single-chain variable fragment) 。未來希望能進一步運用這些scFv抗體片段應用到臨床及實驗研究上，並成為專一性的診斷試劑。
    Severe acute respiratory syndrome (SARS) has led to serious casualties worldwide in year 2003, which can be attributed to a SARS-associated coronavirus or SARS-CoV. Research revealed that nucleocapsid protein (N protein) in the virus particles allows it to invade the host cells and cause infection. An effective and rapid diagnosis or treatment of SARS should be developed. In this study, 5 truncated SARS-CoV nucleocapsid DNA fragment were cloned into pGEX vector and expressed as GST-nucleocapsid in the E. coli BL-21 cells. We used GST Sepharose 4B to purify GST-nucleocapsid protein and then use coomassie blue stain and western blot for the detection of purified GST-nucleocapsid fusion protein. The purified GST-nucleocapsid fusion protein was then mixed with adjuvant and injected intramuscularly into Leghorn chicken. Polyclonal IgY antibodies were purified from the immunized chicken and examined by enzyme-linked immunosorbent assay (ELISA) and western blot analysis. Our study aims to use a recombinant 5 nucleocapsid protein of different length fragment to immunize chicken and constructed an antibody library by phage display technology. By screening of a scFv (single-chain variable fragment) phage library, the nucleocapsid protein specific scFv antibodies was isolated. This will generate specific binding scFv that can identify N protein by phage display. These scFv antibody molecules can be further applied clinically and experimentally for future studies.
    Data Type: thesis
    Appears in Collections:[Graduate Institute of Medical Sciences] Dissertation/Thesis

    Files in This Item:

    File Description SizeFormat
    摘要.doc28KbMicrosoft Word156View/Open
    摘要.pdf67KbAdobe PDF352View/Open
    摘要.ppt110KbMicrosoft Powerpoint223View/Open
    摘要.ps403KbPostscript85View/Open


    All items in TMUIR are protected by copyright, with all rights reserved.

    著作權聲明 Copyright Notice

    • 本平台之數位內容為臺北醫學大學所收錄之機構典藏，包含體系內各式學術著作及學術產出。秉持開放取用的精神，提供使用者進行資料檢索、下載與取用，惟仍請適度、合理地於合法範圍內使用本平台之內容，以尊重著作權人之權益。商業上之利用，請先取得著作權人之授權。

      The digital content on this platform is part of the Taipei Medical University Institutional Repository, featuring various academic works and outputs from the institution. It offers free access to academic research and public education for non-commercial use. Please use the content appropriately and within legal boundaries to respect copyright owners' rights. For commercial use, please obtain prior authorization from the copyright owner.

    • 瀏覽或使用本平台，視同使用者已完全接受並瞭解聲明中所有規範、中華民國相關法規、一切國際網路規定及使用慣例，並不得為任何不法目的使用TMUIR。

      By utilising the platform, users are deemed to have fully accepted and understood all the regulations set out in the statement, relevant laws of the Republic of China, all international internet regulations, and usage conventions. Furthermore, users must not use TMUIR for any illegal purposes.

    • 本平台盡力防止侵害著作權人之權益。若發現本平台之數位內容有侵害著作權人權益情事者，煩請權利人通知本平台維護人員([email protected])，將立即採取移除該數位著作等補救措施。

      TMUIR is made to protect the interests of copyright owners. If you believe that any material on the website infringes copyright, please contact our staff([email protected]). We will remove the work from the repository.

    Back to Top
    DSpace Software Copyright © 2002-2004  MIT &  Hewlett-Packard  /   Enhanced by   NTU Library IR team Copyright ©   - Feedback