Reactivity and isotype profiling of monoclonal antibodies using multiple antigenic peptides.
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Authors
Waldron, E.E.Murray, Paul G.
Kolar, Zdenek
Young, Lawrence S.
Brown, C.
Reynolds, Gary
Baumforth, Karl R. N.
Toomey, S.
Astley, S.J.
Perera, Shantha
Nelson, Paul N.
Issue Date
2002
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Show full item recordAbstract
The characterisation of monoclonal antibodies (MAbs) is essential for the development of assay systems particularly where antigens have been developed using synthetic peptides. Indeed some peptide-carrier conjugates fail to induce immune responses and may not generate antibodies that bind to native protein. As an alternative to peptide-carrier conjugates, multiple antigenic peptides (MAPs) have been used for immunization strategies, but with little regard to the characteristics of the MAbs produced. In this study, we used 3 MAPs of Epstein-Barr virus (EBV) latent membrane protein 1 (LMP1) to immunise BALB/c mice. Overall, the polyclonal antibody responses from tail bleeds showed that MAPs evoked B-cell responses. However, on screening 144 hybridomas, 24 MAb supernatants exhibited weak to moderate reactivity in enzyme-linked immunosorbant assay (ELISA) and against cell cytospin preparations (B95.8 and AG876 LCL), respectively. Isotype profiling of hybridoma supernatants also showed that 11 out of 24 were IgM. Further characterization of 6 MAbs in Western blotting showed reactivity to recombinant LMP1 and only one MAb (B28D) showed weak reactivity to the malignant cells (Hodgkin/Reed-Sternberg; HRS cells) of an EBV+ Hodgkin's lymphoma using paraffin-embedded tissue. It is probable that these MAPs failed to augment T-cell help and contributed to the production of low affinity (IgM) antibodies. These observations may be of importance to future immunization strategies, where MAPs are used in the production of monoclonal reagents.Citation
Hybridoma and Hybridomics, 21(5): 393-398Journal
Hybridoma and HybridomicsPubMed ID
12470483Additional Links
http://www.liebertonline.com/doi/abs/10.1089%2F153685902761022751Type
Journal articleLanguage
enISSN
1536-8599ae974a485f413a2113503eed53cd6c53
10.1089/153685902761022751
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