Premium
Scaled‐up production and purification of Clostridium perfringens type A enterotoxin
Author(s) -
Reynolds D.,
Tranter H. S.,
HAMBLETON P.
Publication year - 1986
Publication title -
journal of applied bacteriology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.889
H-Index - 156
eISSN - 1365-2672
pISSN - 0021-8847
DOI - 10.1111/j.1365-2672.1986.tb01091.x
Subject(s) - enterotoxin , clostridium perfringens , toxin , chemistry , microbiology and biotechnology , sephadex , chromatography , biochemistry , biology , bacteria , enzyme , escherichia coli , genetics , gene
Methods for small‐scale production of Clostridium perfringens type A enterotoxin were unsuitable for large‐scale culture of this organism. Rapid, efficient harvesting of 40 1 batch culture of Cl. perfringens was achieved by tangential flow micro‐filtration with the Millipore Pellicon cassette system. Enterotoxin‐containing extracts were prepared by passing concentrated suspensions of the harvested cells through a French pressure cell. The overall yield of purified enterotoxin was 38·8%. The toxin gave a single band on native polyacrylamide gels but formed high molecular weight aggregates in the presence of sodium dodecyl sulphate. These aggregates frequently occurred during storage of non‐sterile enterotoxin preparations but could be separated from the monomer toxin by gel filtration on Sephadex G‐100. Purified monomer enterotoxin had biological activities of 119·3 μ g/kg mouse lethal dose when injected intraperitoneally and 3333 capillary permeability increasing units/mg protein in guinea pig skin. Thirty μg of the enterotoxin caused fluid accumulation in ligated rabbit ileal loops. Aggregated enterotoxin had no demonstrable biological or immunological activity.