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Iranian Journal of Biotechnology، جلد ۱۲، شماره ۲، صفحات ۹-۱۴
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عنوان انگلیسی Cloning and Expression of the Variable Regions of Anti-EGFR Monoclonal Antibody in E. coli for Production of a Single Chain Antibody
چکیده انگلیسی مقاله Background:Epidermal growth factor receptor (EGFR) overexpression is a characteristic of several malignancies and could be considered as an excellent target for designing specific inhibitors such as anti-EGFR monoclonal antibodies for cancer therapy. Drawbacks exerted by large sizes of full-length antibodies have lead to the development of single chain antibodies, which benefit from having smaller sizes and short circulation half-lives. Objectives:The aim of this study was cloning, expression and purification of variable regions of anti-EGFR monoclonal antibody in E. coli for production of single chain antibodies. Materials and Methods:The RNA, extracted from the C225 hybridoma cells, was reverse transcribed into cDNA and used for PCR amplification of genes encoding light and heavy chains from the variable regions. The PCR products were cloned and expressed in E. coli BL21 for production of a single chain antibody. The expressed protein was analyzed by SDS-PAGE and purified by Ni-NTA affinity chromatography. The reactivity of purified C225-scFv with EGFR-expressing A431 tumor cell line was tested by western blotting and enzyme-linked immunosorbent assays. Results:The results indicated that C225-scFv was highly expressed in E. coli and appeared as a protein with a mass of 27 kDa in the sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) analysis of the induced cell lysate. Reactivity analysis of the purified C225-scFv with A431 tumor cell line by western blotting and enzyme linked immune sorbant assay (ELISA) revealed high binding affinity of the recombinant C225-scFv to the target cells. Conclusions:The results of this study indicated that C225-scFv is capable of binding to EGFR and could be considered as a useful tool for diagnosis and treatment of EGFR-overexpressing tumor cells.
کلیدواژه‌های انگلیسی مقاله Single-Chain Antibody,EGFR,C225
نویسندگان مقاله فرزانه jalalypour |
biotechnology research center, tabriz university of medical sciences, tabriz, i.r. iran drug applied research center, tabriz university of medical sciences, tabriz, i.r. iran
سازمان اصلی تایید شده: دانشگاه علوم پزشکی تبریز (Tabriz university of medical sciences)

صفر فرج نیا |
immunology research center, tabriz university of medical sciences, tabriz, i.r. iran
سازمان اصلی تایید شده: دانشگاه علوم پزشکی تبریز (Tabriz university of medical sciences)

فاطمه محمودی |
department of cellular and molecular biology, azarbaijan shahid madani university, tabriz, i.r. iran
سازمان اصلی تایید شده: دانشگاه شهید مدنی (Azarbaijan shahid madani university)

بهزاد برادران |
immunology research center, tabriz university of medical sciences, tabriz, i.r. iran
سازمان اصلی تایید شده: دانشگاه علوم پزشکی تبریز (Tabriz university of medical sciences)

داود فرج زاده |
department of cellular and molecular biology, azarbaijan shahid madani university, tabriz, i.r. iran
سازمان اصلی تایید شده: دانشگاه شهید مدنی (Azarbaijan shahid madani university)

لیلا رهبرنیا |
drug applied research center, tabriz university of medical sciences, tabriz, i.r. iran
سازمان اصلی تایید شده: دانشگاه علوم پزشکی تبریز (Tabriz university of medical sciences)

جعفر مجیدی |
immunology research center, tabriz university of medical sciences, tabriz, i.r. iran
سازمان اصلی تایید شده: دانشگاه علوم پزشکی تبریز (Tabriz university of medical sciences)

نشانی اینترنتی http://www.ijbiotech.com/article_7251_a16219ecffaaa4edb07a22d0174b84be.pdf
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