Generation of HER2 monoclonal antibodies using epitopes of a rabbit polyclonal antibody
2014 (English)In: New Biotechnology, ISSN 1871-6784, Vol. 31, no 1, 35-43 p.Article in journal (Refereed) Published
One of the issues in using polyclonal antibodies is the limited amount of reagent available from an immunisation, leading to batch-to-batch variation and difficulties in obtaining the same antibody performance when the same antigen is re-immunised into several separate animals. This led to the development of hybridoma technology allowing, at least theoretically, for an unlimited production of a specific binder. Nevertheless, polyclonal antibodies are widely used in research and diagnostics and there exists a need for robust methods to convert a polyclonal antibody with good binding performance into a renewable monoclonal with identical or similar binding specificity. Here we have used precise information regarding the functional recognition sequence (epitope) of a rabbit polyclonal antibody with attractive binding characteristics as the basis for generation of a renewable mouse monoclonal antibody. First, the original protein fragment antigen was used for immunisation and generation of mouse hybridoma, without obtaining binders to the same epitope region. Instead a peptide designed using the functional epitope and structural information was synthesised and used for hybridoma production. Several of the monoclonal antibodies generated were found to have similar binding characteristics to those of the original polyclonal antibody. These monoclonal antibodies detected native HER2 on cell lines and were also able to stain HER2 in immunohistochemistry using xenografted mice, as well as human normal and cancer tissues.
Place, publisher, year, edition, pages
2014. Vol. 31, no 1, 35-43 p.
Batch-to-batch variations, Binding characteristics, Binding specificities, Hybridoma technology, Immunohistochemistry, Polyclonal antibody, Recognition sequence, Structural information
Biochemistry and Molecular Biology
IdentifiersURN: urn:nbn:se:kth:diva-139254DOI: 10.1016/j.nbt.2013.10.002ISI: 000328131200004ScopusID: 2-s2.0-84889568450OAI: oai:DiVA.org:kth-139254DiVA: diva2:685945
FunderEU, FP7, Seventh Framework Programme, 241481VinnovaKnut and Alice Wallenberg FoundationScience for Life Laboratory - a national resource center for high-throughput molecular bioscience
QC 201401102014-01-102014-01-082014-01-10Bibliographically approved