Production of serotype C specific and serotype C/D generic monoclonal antibodies using recombinant H(C) and H(N) fragments from Clostridium botulinum neurotoxin types C(1) and D.
Sequence variability of Clostridium botulinum serotypes C and D is particularly complex. Some serotype C and D strains have unique gene structures that encode mosaic isoforms of botulinum neurotoxin (BoNT) containing components of both BoNT type C(1) (BoNT/C(1)) and BoNT type D (BoNT/D). Such sequence variability and the potential for cross neutralisation must be taken into consideration when developing serotype C and D detection and identification assays. Three fusion proteins containing either a fragment from the carboxyl-terminal domain of the heavy chain (H(C)) of BoNT/C(1) (strain 573), a fragment from the H(C) of BoNT/D (strain BVD/-3) or a fragment from the amino-terminal domain of the heavy chain (H(N)) of BoNT/C(1) (strain 573) were expressed in Escherichia coli, and administered as immunogens to mice. Monoclonal antibodies (mAbs) against the recombinant BoNT fragments were prepared by three fusions. MAbs recognising native BoNT/C(1) and BoNT/D were detected by enzyme-linked immunosorbent assay (ELISA). Nine monoclonal antibodies (mAbs) were produced, six of which recognised a BoNT fragment that is highly conserved across all serotype C and D producing strains. We conclude that these mAbs and this approach to mAb production may facilitate the development of immunological diagnostic techniques that are not constrained by the existence of mosaic isoforms for the detection and identification of serotypes C and D.
Curran
R M
RM
Veterinary Sciences Division/School of Biological Sciences, Queens University of Belfast, Stoney Rd., Stormont, Belfast BT43SD, United Kingdom. rcurran05@qub.ac.uk
Fringuelli
E
E
Graham
D
D
Elliott
C T
CT
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000235:genetics; D001905:Botulinum Toxins / Q000235:genetics / Q000276:immunology*; D001906:Botulism / Q000175:diagnosis / Q000382:microbiology / Q000662:veterinary*; D047968:Clostridium botulinum type C / Q000235:genetics / Q000276:immunology*; D047969:Clostridium botulinum type D / Q000235:genetics / Q000276:immunology*; D003433:Crosses, Genetic; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay / Q000662:veterinary; D005260:Female; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008813:Mice, Inbred ICR; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000662:veterinary; D020033:Protein Isoforms; D011994:Recombinant Proteins / Q000235:genetics
D000911:Antibodies, Monoclonal; D004269:DNA, Bacterial; D020033:Protein Isoforms; D011994:Recombinant Proteins; C096325:botulinum toxin type D; D001905:Botulinum Toxins; C096324:botulinum toxin type C