Fermentation, Recovery, and Purification of the HC Fragments of the Botulinum Neurotoxin from Pichia Pastoris
Abstract
The report documents the research and development activities on BoNTC Hc, BoNTF Hc, and BoNTE Hc. BoNTC Hc research focused on characterizing the capture of the BoNTC Hc from P. pastoris lysate and the interference of DNA-type material on the binding of BoNTC Hc to a anion exchange column. BoNTF Hc process development was concerned with confirmation of a process developed by USARMIID transferred to Covance and tested at UN-L. UN-L found two steps needed further development, i.e. cell breakage conditions and one chromatographic condition. This process was scaled-up to the pilot scale and transferred back to USAMRIID. Fermentation research at the UN-L focused on developing standard fermentation protocols that can be easily transferred to Covance. The BoNTC sub 1 Hc P. pastoris fermentation process was successfully developed and optimized using a quantifiable Western blot procedure. Also, a fermentation process for BoNTE Hc express in E. coil was developed and conditions were determined for expression of soluble material. UN-L also provided fermentation support for BoNT D, B, and E Hc for both USAMRIID and UN-L purification activities. The Quality Assurance/Quality Control group at UN-L established multiple pre-seeds for clones developed at USAMRIID and established batch records for the BoTNF Hc process.
Document Details
- Document Type
- Technical Report
- Publication Date
- Jun 01, 2000
- Accession Number
- ADA384855
Entities
People
- Michael M. Meagher
Organizations
- University of Nebraska–Lincoln