Single-step purification of pertussis toxin and its subunits by heat-treated fetuin-sepharose affinity chromatography.
A general procedure for purifying biologically active pertussis toxin from Bordetella pertussis fermentation broth using affinity chromatography on heat-treated fetuin-Sepharose CL-4B is described. Diethanolamine is used as eluent in this single-step purification to prepare endotoxin-free pertussis toxin in good yield (70%) and high purity (greater than 95%). This one-step affinity chromatography procedure can be easily applied for large-scale preparation of pertussis toxin S1 subunit and its B-component. The affinity-purified S1 subunit is devoid of any of the histamine-sensitizing activity normally associated with pertussis toxin. The chromatographically purified pertussis toxin and its subunits retained their immunogenicity and could induce high levels of anti-toxin neutralizing antibodies.
Chong
P
P
Connaught Centre for Biotechnology Research, Willowdale, Ont., Canada.
Klein
M
M
D000818:Animals; D001426:Bacterial Proteins / Q000032:analysis; D001886:Bordetella pertussis / Q000378:metabolism; D002846:Chromatography, Affinity; D003470:Culture Media; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D006632:Histamine / Q000633:toxicity; D008070:Lipopolysaccharides / Q000378:metabolism; D051379:Mice; D037342:Pertussis Toxin; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D012685:Sepharose / Q000031:analogs & derivatives; D010566:Virulence Factors, Bordetella / Q000302:isolation & purification* / Q000633:toxicity; D000509:alpha-Fetoproteins
D001426:Bacterial Proteins; D003470:Culture Media; D008070:Lipopolysaccharides; D010566:Virulence Factors, Bordetella; D000509:alpha-Fetoproteins; C035771:Sepharose CL 4B; D006632:Histamine; D012685:Sepharose; D011065:Poly(ADP-ribose) Polymerases; D037342:Pertussis Toxin