Appeal No. 1999-2634
Application No. 08/108,606
providing at least one E. coli bacterial strain expressing colonization factor
antigens on the surface of said bacteria,
growing said E. coli bacterial strain in a liquid culture medium with vigorous
agitation to a predetermined density,
harvesting said E. coli bacterial strain,
resuspending said harvested E. coli bacterial strain in saline,
adding formalin to said harvested, resuspended bacterial strain to a final
concentration of 0.2M formaldehyde,
incubating said formalin-treated E. coli bacterial strain at 37EC under conditions
of continuous agitation for about 2 hours,
further incubating said formalin-treated bacterial strain at 4EC for between about
24 hours and about 48 hours, thereby obtaining a formalin-killed E. coli bacterial strain
and,
collecting said formalin-killed E. coli bacterial strain.
9. The method according to claim 7, wherein said liquid culture medium
comprises 1% (w/v) casamino acids, 0.15% (w/v) yeast extract, 0.4 mM MgSo4,0.04mM
MgCl2, and deinozed water at pH 7.4, and said growing step is conducted with vigorous
agitation at about 37EC for at least 4-6 hours before said harvesting step.
11. The method according to claim 7 further comprising adding an acid-
neutralizing buffer.
13. A method of prevention [sic of] an enteric infection caused by
enterotoxigenic E. coli bacteria in humans comprising administrating an appropriate
amount for preventing said infection of a vaccine comprising at least one formalin-
inactivated E. coli strain expressing colonization factor antigens and further having
substantially preserved antigenic and hemagglutinating properties of said colonization
factor antigens.
15. The method of claim 13, wherein said vaccine further comprises cholera
toxin b-subunit.
The prior art references relied upon by the examiner are:
Myers 4,338,298 July 6, 1982
Evans, D.J., et al (Evans 1), AImmunoprotective oral whole cell vaccine for
enterotoxigenic Escherichia coli diarrhea prepared by in situ destruction of
chromosomal and plasmid DNA with colicin E2,@ FEMS Microbiology and Immunology,
Vol. 47, pp. 9-18 (1988)
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