This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Byrne, M. P.
Right arrow Articles by Smith, L. A.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Byrne, M. P.
Right arrow Articles by Smith, L. A.

 Previous Article  |  Next Article 

Infection and Immunity, October 1998, p. 4817-4822, Vol. 66, No. 10
0019-9567/98/$00.00+0

Purification, Potency, and Efficacy of the Botulinum Neurotoxin Type A Binding Domain from Pichia pastoris as a Recombinant Vaccine Candidate

Michael P. Byrne, Theresa J. Smith, Vicki A. Montgomery, and Leonard A. Smith*

Division of Toxinology, United States Army Research Institute for Infectious Diseases, Frederick, Maryland 21702-5011

Received 30 April 1998/Returned for modification 9 June 1998/Accepted 7 July 1998

Recombinant botulinum neurotoxin serotype A binding domain [BoNT/A(Hc)], expressed in Pichia pastoris, was developed as a vaccine candidate for preventing botulinum neurotoxin type A (BoNT/A) intoxication. After fermentation and cell disruption, BoNT/A(Hc) was purified by using a three-step chromatographic process consisting of expanded-bed chromatography, Mono S cation-exchange chromatography, and hydrophobic interaction chromatography. Two pools of immunogenic product were separated on the Mono S column and processed individually. Both products were more than 95% pure and indistinguishable by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, Western blot analysis, and enzyme-linked immunosorbent assay (ELISA). Each protein was assayed for potency in mice at immunogen doses ranging from 2.4 ng to 10 µg, followed by challenge with 1,000 mouse intraperitoneal 50% lethal doses (i.p. LD50) of BoNT/A. The calculated 50% effective dose for both peaks was approximately 0.1 µg/mouse. Peak 1 was evaluated further in a mouse efficacy assay. Mice were injected either once, twice, or three times at five different doses and subsequently challenged with 100,000 mouse i.p. LD50 of BoNT/A. In general, multiple injections protected better than one, with complete or nearly complete protection realized at doses of >= 0.5 µg/mouse. Serum neutralization and ELISA titers were also determined. Tellingly, 82 of 83 mice with antibody titers of >= 1,600, as measured by ELISA, survived, but only 6 of 42 mice with titers of <= 100 survived. This work shows that the purified BoNT/A(Hc) produced was a highly effective immunogen, able to protect against a high challenge dose of neurotoxin.

* Corresponding author. Mailing address: Toxinology Division, US AMRIID, Building 1425, Fort Detrick, MD 21702-5011. Phone: (301) 619-4238. Fax: (301) 619-2348. E-mail: Dr._Leonard_Smith{at}detrick.army.mil.

Infection and Immunity, October 1998, p. 4817-4822, Vol. 66, No. 10
0019-9567/98/$00.00+0


This article has been cited by other articles:

  • Shone, C., Agostini, H., Clancy, J., Gu, M., Yang, H.-H., Chu, Y., Johnson, V., Taal, M., McGlashan, J., Brehm, J., Tong, X. (2009). Bivalent Recombinant Vaccine for Botulinum Neurotoxin Types A and B Based on a Polypeptide Comprising Their Effector and Translocation Domains That Is Protective against the Predominant A and B Subtypes. Infect. Immun. 77: 2795-2801 [Abstract] [Full Text]  
  • Yu, Y.-Z., Li, N., Wang, R.-L., Zhu, H.-Q., Wang, S., Yu, W.-Y., Sun, Z.-W. (2008). Evaluation of a Recombinant Hc of Clostridium botulinum Neurotoxin Serotype F as an Effective Subunit Vaccine. CVI 15: 1819-1823 [Abstract] [Full Text]  
  • Keller, J. E. (2008). Characterization of New Formalin-Detoxified Botulinum Neurotoxin Toxoids. CVI 15: 1374-1379 [Abstract] [Full Text]  
  • Baldwin, M. R., Tepp, W. H., Przedpelski, A., Pier, C. L., Bradshaw, M., Johnson, E. A., Barbieri, J. T. (2008). Subunit Vaccine against the Seven Serotypes of Botulism. Infect. Immun. 76: 1314-1318 [Abstract] [Full Text]  
  • Dembek, Z. F., Smith, L. A., Rusnak, J. M. (2007). Botulism: Cause, Effects, Diagnosis, Clinical and Laboratory Identification, and Treatment Modalities. dmphp 1: 122-134 [Abstract] [Full Text]  
  • Ravichandran, E., Al-Saleem, F. H., Ancharski, D. M., Elias, M. D., Singh, A. K., Shamim, M., Gong, Y., Simpson, L. L. (2007). Trivalent Vaccine against Botulinum Toxin Serotypes A, B, and E That Can Be Administered by the Mucosal Route. Infect. Immun. 75: 3043-3054 [Abstract] [Full Text]  
  • Maddaloni, M., Staats, H. F., Mierzejewska, D., Hoyt, T., Robinson, A., Callis, G., Kozaki, S., Kiyono, H., McGhee, J. R., Fujihashi, K., Pascual, D. W. (2006). Mucosal Vaccine Targeting Improves Onset of Mucosal and Systemic Immunity to Botulinum Neurotoxin A. J. Immunol. 177: 5524-5532 [Abstract] [Full Text]  
  • Baldwin, M. R., Tepp, W. H., Pier, C. L., Bradshaw, M., Ho, M., Wilson, B. A., Fritz, R. B., Johnson, E. A., Barbieri, J. T. (2005). Characterization of the Antibody Response to the Receptor Binding Domain of Botulinum Neurotoxin Serotypes A and E. Infect. Immun. 73: 6998-7005 [Abstract] [Full Text]  
  • Lee, J. S., Pushko, P., Parker, M. D., Dertzbaugh, M. T., Smith, L. A., Smith, J. F. (2001). Candidate Vaccine against Botulinum Neurotoxin Serotype A Derived from a Venezuelan Equine Encephalitis Virus Vector System. Infect. Immun. 69: 5709-5715 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»