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Published on: October 15, 2016
Construction of scFv library of human antibodies against tetanus toxin
Hamideh Rouhani Nejad1, Jalil Fallah Mehrabadi2, Jamile Nourozi1
1Department of Biology, Science and Research Branch, Islamic Azad University, Tehran, Iran.
Background:
Over the last three decades, monoclonal antibodies have become powerful therapeutics and diagnostics tools. Progress in the antibody engineering, and the appearance of great selection technology such as phage display has made human antibodies production possible against antigens with high affinities.
Objective:
The purpose of this study was to construct an immune antibody library from a vaccinated donor against tetanus toxin.
Methods:
A blood sample was drawn from the donor who was vaccinated with tetanus toxoid. PBMC were isolated by using ficoll. After RNA extraction and cDNA synthesis. In order to amplify VH and VL regions, two uniplex PCRs were performed and put considering NcoI & HindIII, MluI & NotI restriction sites respectively for their cloning. These amplicons were cloned to pSEX81 vector and transformed to E. coli XL1blue strain. Eventually, recombinant plasmids were extracted and sequenced.
Results:
The result showed acceptable similarity between antibody gene library nucleotide sequences and the antibody genes were deposited in this database.
Conclusion:
In this study, the VH and VL genes human antibody library was constructed and confirmed by using DNA sequencing and alignment of sequences with blast database.
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