Genetic and immunological comparison of anti-botulinum type A antibodies from immune and non-immune human phage libraries.
Understanding the antibody response in botulinum intoxication is important for vaccine design and passive prophylaxis. To investigate this activity, we have studied the immune response to BoNT/A (botulinum neurotoxin serotype A) binding domain (HC) at the molecular level using phage display. The scFv antibodies were isolated from V-gene repertoires prepared from (a) human volunteer immunized with pentavalent botulinum toxoid and (b) non-immune human peripheral blood lymphocytes and spleenocytes. A large panel of serotype specific phage expressing botulinum binding scFv could be selected from both libraries. Epitope mapping of immune scFv binders towards BoNT/A HC revealed surprisingly a limited number of scFv recognizing conformational epitopes that corresponded to two distinct groups, clusters I and II. Only scFv from cluster I exhibited neutralizing activity in the mouse hemidiaphragm assay. Anti- BoNT/A HC clones derived from a non-immune library could be conveniently grouped into clusters III-XI and appeared to share no overlapping epitopes with cluster I or II. In addition they showed no neutralization of toxin at biologically significant concentrations. We therefore suggest that a vaccine based on the pentavalent botulinum toxoid directs the humoral immune response to a limited number of immunodominant epitopes exposed on the binding domain HC.
Amersdorfer
Peter
P
Department of Anesthesia and Pharmaceutical Chemistry, San Francisco General Hospital, University of California, San Francisco Rm 3C-38, 1001 Potrero Avenue, San Francisco, CA 94110, USA. peter_amersdorfer@corvas.com
Wong
Cindy
C
Smith
Theresa
T
Chen
Steven
S
Deshpande
Sharad
S
Sheridan
Robert
R
Marks
James D
JD
D000907:Antibodies, Bacterial / Q000235:genetics* / Q000276:immunology*; D000911:Antibodies, Monoclonal / Q000235:genetics / Q000276:immunology; D019274:Botulinum Toxins, Type A / Q000276:immunology* / Q000633:toxicity; D001906:Botulism / Q000276:immunology / Q000517:prevention & control / Q000628:therapy; D003014:Clostridium botulinum / Q000276:immunology; D018604:Epitope Mapping; D006801:Humans; D007135:Immunoglobulin Variable Region / Q000235:genetics / Q000276:immunology; D009500:Neutralization Tests; D019151:Peptide Library
D000907:Antibodies, Bacterial; D000911:Antibodies, Monoclonal; D007135:Immunoglobulin Variable Region; D019151:Peptide Library; D019274:Botulinum Toxins, Type A