Immunization against clostridium difficile disease
The invention provides active and passive immunization methods for preventing and treating Clostridium difficile infection, which involve percutaneous administration of C. difficile toxin-neutralizing polyclonal immune globulin, C. difficile toxoids, or combinations thereof. Also provided by the invention are C. difficile toxoids, C. difficile toxin-neutralizing polyclonal immune globulin, and methods of identifying subjects that produce C. difficile toxin-neutralizing polyclonal immune globulin.
1 . A method of producing a Clostridium difficile toxoid, said method comprising the steps of:
providing a Clostridium difficile bacterium;
culturing said bacterium in medium lacking animal products to generate a culture;
purifying clostridial toxin A and clostridial toxin B from said culture to generate a mixture of purified toxins A and B; and
inactivating said purified toxins A and B by incubation in formaldehyde at a temperature of 25° C. or less to generate said clostridial toxoid.
2 . The method of claim 1 , wherein said purified toxins A and B are incubated in formaldehyde at a temperature of about 15° C. or less, or a temperature of about 5° C. or less.
3 . The method of claim 1 , wherein said purified toxins A and B are present in said mixture at a ratio of about 0.1:1 to 10:1 or a ratio of about 2:1.
4 . A Clostridium difficile toxoid produced by the method of claim 1 .
5 . A vaccine composition comprising the Clostridium difficile toxoid of claim 4 .
6 . The vaccine composition of claim 5 , further comprising 0.012-0.020% formaldehyde.
7 . The vaccine composition of claim 5 , further comprising an adjuvant.
8 . The vaccine composition of claim 7 , wherein the adjuvant is an aluminum compound.
9 . A method of producing human immune globulin directed against a clostridium , said method comprising the steps of:
administering a clostridial toxin or toxoid to a human; and
isolating said immune globulin against a clostridium from said human.
10 . The method of claim 9 , wherein the clostridium is Clostridium difficile.
11 . The method of claim 9 , wherein said clostridial toxoid comprises toxin A and toxin B.
12 . Clostridium difficile immune globulin produced by administering a Clostridium difficile toxin or toxoid to a human, and isolating said Clostridium difficile immune globulin from said human
13 . A method of identifying a human producing a Clostridium difficile immune globulin, said method comprising the steps of:
obtaining a blood sample from a human vaccinated with a Clostridium difficile toxoid;
determining the level of antibodies to Clostridium difficile Toxins A and B in said blood sample by an enzyme-linked immunosorbent assay; and
determining the level of in vitro cytotoxicity neutralization activity against Clostridium difficile Toxins A and B in said blood sample, wherein detection of increased levels of antibodies to Clostridium difficile Toxins A and B in said blood sample, and
detection of in vitro cytotoxicity neutralization activity against Clostridium difficile Toxins A and B in said blood sample, indicates identification of a human producing a Clostridium difficile immune globulin useful for treatment or prevention of Clostridium difficile infection.