This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The goal of this project is to advance the global effort toward developing effective therapeutic approaches for the prevention of Chlamydia trachomatis genital infections. The major outer membrane protein (MOMP) of C. trachomatis has been the target of extensive efforts to develop efficacious vaccines against chlamydial infection. Our hypothesis is that recombinant (r) MOMP coupled with a mucosal adjuvant will elicit immune responses that are comparable with the native MOMP antigen. In this study, MOMP of C. trachomatis strain MoPn Nigg II was cloned, expressed and then genetically fused with modified cholera toxin as a mucosal adjuvant (cholera toxin subunit A2B). The rMOMP gene was amplified and cloned into the plasmid vector pUAB084, which contained the ctxA2B gene. The plasmids were used to transform E. coli BL21 (DE3) and rMOMP-CTB fusion protein was expressed by isopropyl-?-D-thiogalactoside (IPTG) induction. Fractions of the expression host cells were analyzed for the presence of the MOMP-CTB fusion protein by SDS-PAGE. Polyclonal antibodies to cholera toxin and the MOMP protein were used in western blot and ELISA to confirm the presence of the MOMP-CTB fusion protein. GM1-ELISA was also performed to confirm the proper association of MOMP-CTB fusion protein. The rMOMP-CTB fusion protein was used to immunize six groups of BALB/c mice. Groups of mice were immunized intranasally or intravaginally with recombinant MOMP-CTB, live C. trachomatis, or PBS. Serum/vaginal washes were collected and the antibody titers were determined by ELISA. rMOMP-CTB fusion protein administered intranasally provided the highest antibody titer. The IgA, IgG2a, IgG2b isotypes were most prevalent in serum collected from mice immunized intranasally with rMOMP. IgA isotype titers were significantly higher in these groups compared to other immunizations including live C. trachomatis. These early studies support the hypothesis that rMOMP protein coupled to a mucosal adjuvant can stimulate mucosal immune response
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