Development Of An Immune Tuberculosis Antibody Library: Monoclonal Antibody Generation For a-Crystalline Antigen
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Date
2017-05
Authors
Hamidon, Nurul Hamizah
Journal Title
Journal ISSN
Volume Title
Publisher
Universiti Sains Malaysia
Abstract
Antibodies are very important for therapeutic and diagnostic applications for
infectious diseases. Immune libraries are useful for infectious disease applications
because it would allow for better and selective enrichment of antibodies against
disease antigens. The generations of antibody libraries are necessary to produce the
monoclonal antibodies. Immune scFv antibody library was generated from six
donors (tuberculosis patients). Conventional PCR method was used to amplify all
possible antibody genes, which were then cloned into the pLABEL, phagemid
vector. A two-step cloning procedure was done to generate the antibody library. The
library size of the mini HC library and the scFv library constructed was
approximately around 107 and 109 individual clones respectively. The insert rate for
two-step cloning was only 63% and -crystalline antigen was used for verification of
the antibody library and monoclonal isolation. Conventional plate panning technique
was used for selection of monoclonal antibodies. Three monoclonal scFv antibodies
were successfully obtained for the Mycobacterium tuberculosis, -crystalline
(Rv2331c) antigen and the antibody sequence was analyzed using the software,
VBASE2. The clones were IGKV3 and IGHV2 for clone 1H, IGKV4 and IGHV2 for
clone 1B and IGLV1 and LGHV1 for clone 10G. The antibodies enriched was also
evaluated in soluble form for binding activities. In conclusion, a diverse immune
scFv library was constructed from which soluble monoclonal scFv antibodies against
a-crystalline antigen was obtained.
Description
Keywords
Development of an immune tuberculosis antibody , monoclonal antibody generation