SummaryThis paper is the first to describe characterization of monoclonal antibodies (MAbs) against a South African Territories 2 (SAT 2) foot-and-mouth disease virus (isolate Rho 1/48). Twelve MAbs which neutralized homologous virus were characterized in indirect and sandwich ELISA using purified Rho 1/48 virus particles, subunits, trypsin-treated, and chemically denatured virus. All the Mabs inhibited haemagglutination by parental virus. Binding of the MAbs to 73 SAT 2 field isolates was measured in a sandwich ELISA and defined four distinct antigenic regions. Preliminary characterization of escape mutants selected with some of the MAbs using virus neutralization tests, ELISA, and amino acid sequencing is included. MAbs 2, 25, 40, 48 and 64, reacted with a linear epitope on the VP1 loop region. An amino acid change at position 149 (valine to glutamic acid) was detected in mutants selected by MAb 2 and 40 and this eliminated binding and neutralization by all the other MAb. This epitope was conformation-dependent and was conserved in all 73 isolates of SAT 2 examined. Escape mutants isolated with MAb 41 and 44, had changes at positions 156 (glycine to aspartic acid), or 158 (serine to leucine) respectively. These MAbs bound with Rho 1/48 only out of 73 field strain viruses studies and the reactions of MAbs from the other groups was unaltered. MAb 27, 28 and 37 reacted with a conformation-dependent epitope on VP1 which was not conserved in field isolates. All mutants selected by these MAbs had a single amino acid substitution at position 149 (valine to alanine). The same change was always found in field isolates which did not bind MAbs from this group. MAb 11 reacted with a linear epitope associated with amino acids 147 or 148 on VP1 and showed similar binding characteristics to a conformation dependent MAb 7, no amino acid residue changes were found within VP1 for monoclonal antibody 7 mutants.
Eight neutralizing and two non-neutralizing antifoot-and-mouth disease virus (FMDV) bovine IgG1 and IgG2 monoclonal antibodies (BMAbs) recognize conformationally dependent epitopes. The majority of those shown to neutralize virus passively protected mice from virus challenge, regardless of isotype. Well-characterized anti-FMDV mouse MAbs, representing five independent neutralizing epitopes on O 1 serotype virus, were examined with each of the ten BMAbs in a competition-based ELISA. Five of the neutralizing BMAbs (C48, C65, C74, C83 and MH6) were shown to be directed against epitopes on, or in close proximity to, that previously defined as neutralizing antigenic site 2. Another neutralizing BMAb, MH5, bound to an epitope on, or in close proximity to antigenic site 3. Two neutralizing BMAbs (C2 and C96) simultaneously
We have developed technology to create monoclonal antibodies (MAbs) using lymphocytes from immunized sheep. The affinities of these sheep monoclonal antibodies (SMA) can be several orders of magnitude higher than mouse MAbs. This paper reports the development and validation of a modified enzyme-linked immunoadsorbent assay (ELISA) method to select high-affinity antibodies of the desired specificity at the first screen. Using this method, we have isolated high affinity SMA to carcinoembryonic antigen (CEA), a marker of colon cancer. Comparisons of our novel SMA with mouse MAbs using the new ELISA and BIAcore technology (BIAcore AB, Stevenage, Herts, UK) have confirmed that we have made super-high affinity antibodies to CEA. One of these has a t(1/2) for dissociation of 8 days, which could provide a longer therapeutic window than is available with murine monoclonals currently being used in the clinic. This antibody has a specific tissue staining profile; it thus appears to be an excellent candidate for use in the clinic.
This paper is the first description of monoclonal antibodies specific for foot-and-mouth disease virus (FMDV). Using these antibodies, it was possible to distinguish similar and unique antigenic sites on complete (146S) or subunit (12S) virus particles. Some of these monoclonal antibodies could also distinguish between 12S subunits made by acid degradation of 146S virus (12SA) and those viral components found in infected cell lysates which sediment in sucrose density gradients as "12S subunits" (12SN).
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