Methods for producing enhanced antigenic campylobacter bacteria and vaccines
AssigneeAntex Biologics Inc.
InventorsJohn Lee Pace, Richard Ives Walker, Steven Michael Frey
Life of this patent
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Abstract
Methods using in vitro processes are disclosed for inducing or enhancing expression of enteric bacterial antigens or virulence factors. The methods, therefore, produce antigenically enhanced enteric bacteria. Also disclosed are methods for using the antigenically enhanced bacteria, as well as vaccines containing the enteric bacteria. Specifically, a whole enteric bacteria or components thereof are provided by Campylobacter species. In addition to this species there are other enteric bacteria, such as Helicobacter pylori, which are useful for inducing or enhancing expression of enteric bacterial antigens and/or virulence factors.
Claims12
A method of growing Campylobacter bacteria having an enhanced antigenic property which comprises growing a culture of a Campylobacter species in vitro in a culture medium with a combination of conditions comprising:
- a) about 0.05% to about 3% bile or about 0.025% to about 0.6% of one or more bile acids or salts thereof;
- b) at a temperature between about 30.degree. C. and about 42.degree. C.;
- c) in air or a gas mixture, wherein the gas mixture comprises i) about 5% to about 20% CO.sub.2 with about 80% to about 95% air;
- or ii), about 5% to about 10% O.sub.2 with about 10% to about 20% CO.sub.2 with about 70% to about 85% N.sub.2 ;
- and
- d) a divalent cation chelator selected from the group consisting of 0 to about 100 .mu.M of 1,2-bis(2-aminophenoxy)ethane-N,N,N', N'-tetraacetic acid/acetoxymethyl ester, 0 to about 10 mM of ethylene-bis(oxyethylenenitrilo)-tetraacetic acid and 0 to about 100 .mu.M of ethylene-bis(oxyethylenenitrilo)-tetraacetic acid/acetoxymethyl ester,
- for a sufficient time so that the culture is in a growth phase at about early log phase, between early log phase and stationary phase, or at about stationary phase, wherein the enhanced antigenic property is a higher level of an immunogenic antigen or a new immunogenic antigen when compared to the antigenic property of bacteria from a culture of the Campylobacter species grown in brain heart infusion broth.
A method of growing Campylobacter bacteria having an enhanced antigenic property which comprises growing a culture of a Campylobacter species in vitro in a culture medium with a combination of conditions comprising:
- a) a divalent cation chelator selected from the group cosisitng of about 1.0 to about 25 .mu.M of 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid/acetoxymethyl ester, about 0.5 to about 10 mM of ethylene-bis(oxyethylenenitrilo)-tetraacetic acid, and about 1 to about 100 .mu.M of ethylene-bis(oxyethylenenitrilo)-tetraacetic acid/acetoxymethyl ester;
- b) at a temperature between about 30.degree. C. and about 42.degree. C.;
- and
- c) in air or a gas mixture, wherein the gas mixture comprises i) about 5% to about 20% CO.sub.2 with about 80% to about 95% air;
- or ii) about 5% to about 10% O.sub.2 with about 10% to about 20% CO.sub.2 with about 70% to about 85%
- for a sufficient time so that the culture is in a growth phase at about early log phase, between early log phase and stationary phase, or at about stationary phase, wherein the enhanced antigenic property is a higher level of an immunogenic antigen or a new immunogenic antigen when compared to the antigenic property of bacteria from a culture of the Campylobacter species grown in brain heart infusion broth.
A Campylobacter bacterium having an enhanced antigenic property, which is harvested from a culture of a Campylobacter species grown in vitro in a culture medium with a combination of conditions comprising:
- a) about 0.05% to about 3% bile or about 0.025% to about 0.6% of one or more bile acids or salts thereof;
- b) at a temperature between about 30.degree. C. and about 42.degree. C.;
- c) in air or a gas mixture, wherein the gas mixture comprises i) about 5% to about 20% CO.sub.2 with about 80% to about 95% air;
- or ii) about 5% to about 10% O.sub.2 with about 10% to about 20% CO.sub.2 with about 70% to about 85% N.sub.2 ;
- and
- d) a divalent cation chelator selected from the group consisting of 0 to about 100 .mu.M of 1,2-bis(2-aminophenoxy)ethane-N,N,N', N'-tetraacetic acid/acetoxymethyl ester, 0 to about 10 mM of ethylene-bis(oxyethylenenitrilo)-tetraacetic acid, and 0 to about 100 .mu.M of ethylene-bis(oxyethylenenitrilo)-tetraacetic acid/acetoxymethyl ester,
- wherein said Campylobacter culture is in a growth phase at about early log phase, between early log phase and stationary phase, or at about stationary phase, and the enhanced antigenic property is a higher level of an immunogenic antigen or a new immunogenic antigen when compared to the antigenic property of bacteria from a culture of the Campylobacter species grown in brain heart infusion broth.
A Campylobacter bacterium having an enhanced antigenic property, which is harvested from a culture of a Campylobacter species grown in vitro in a culture medium with a combination of conditions comprising:
- a) a divalent cation chelator selected from the group consisting of about 1.0 to about 25 .mu.M of 1,2-bis(2-aminophenoxy)ethane-N, N, N', N'-tetraacetic acid/acetoxymethyl ester,0.5 about 10 mM of ethylene-bis(oxyethylenenitrilo)-tetraacetic acid, and about 1.0 to about 100 .mu.M of ethylene-bis(oxyethyleneitrilo)-tetraacetic acid/acetoxymethyl ester;
- b) at a temperature between about 30.degree. C. and about 42.degree. C.;
- and
- c) in air or a gas mixture, wherein the gas mixture comprises i) about 5% to about 20% CO.sub.2 with about 80% to about 95% air;
- or ii) about 5% to about 10% O.sub.2 with about 10% to about 20% CO.sub.2 with about 70% to about 85% N.sub.2,
- wherein said Campylobacter culture is at about early log phase, between early log phase and stationary phase, or at about stationary phase, and the enhanced antigenic property is a higher level of an immunogenic antigen or a new immunogenic antigen when compared to the antigenic property of bacteria from a culture of the Campylobacter species grown in brain heart infusion broth.
Dependencies are inferred from the claim text.