A new method for the rapid purification of FanC, the major subunit of K99
- نوع فایل : کتاب
- زبان : انگلیسی
- مؤلف : Mehdi Golchin & Fatameh Mohammadi
- چاپ و سال / کشور: 2011
Description
Several strategies and methods have been attempted to purify K99 fimbriae from enterotoxigenic Escherichia coli strains. Most of these methods are technically complex and time consuming. In current study a single-step ion-exchange chromatography method that takes just a few hours for purification of this important virulence factor was developed. K99 fimbriae were stripped from E. coli B41 strain by heat treatment and phosphate– urea buffer. The crude extracts were then equilibrated with Tris buffer and were loaded to a HiTrap SP XL column. By stepwise elevation of sodium chloride concentration, the FanC was eluted as a single species. The purified protein had high activity with anti-K99 monoclonal antibody in ELISA. Also, preincubation of red blood cells with the purified protein blocked adhesion of B41 bacteria. LPS contamination of purified FanC was in acceptable range using limulus amebocyte lysate method. In summary, this simple, inexpensive method yields purified FanC that has good purity, stability, and biological activity, making it suitable for many purposes including vaccination.
Comp Clin Pathol DOI 10.1007/s00580-011-1289-1 Received: 14 March 2011 / Accepted: 8 July 2011
