Production of a monoclonal antibody specific for the major outer membrane protein of Campylobacter jejuni and characterization of the epitope.
Campylobacter species are important enteric pathogens causing disease in humans and animals. There is a lack of a good immunological test that can be used routinely to separate Campylobacter jejuni from other Campylobacter species. We produced monoclonal antibodies (MAbs) directed against the major outer membrane protein (MOMP) of C. jejuni using recombinant MOMP as the antigen. One MAb, designated MAb5C4 and of the immunoglobulin G1 isotype, was found to be potentially specific for C. jejuni. Dot blots demonstrated that MAb5C4 reacted with all 29 isolates of C. jejuni tested but did not react with 2 C. jejuni isolates, 26 other Campylobacter spp. isolates, and 19 non-Campylobacter isolates. Western blotting showed that MAb5C4 bound to a single protein band approximately 43 kDa in size, corresponding to the expected size of C. jejuni MOMP. The detection limit of MAb5C4 in a dot blot assay was determined to be about 5 x 10(3) bacteria. The epitope on the MOMP was mapped to a region six amino acids in length with the sequence 216GGQFNP221, which is 97% conserved among C. jejuni strains but divergent in other Campylobacter spp.; a GenBank search indicated that 95% of C. jejuni isolates will be able to be detected from non-Campylobacter spp. based on the highly specific and conserved region of the GGQFNP polypeptide. The epitope is predicted to be located in a region that is exposed to the periplasm. MAb5C4 is a potentially specific and sensitive MAb that can be used for the specific detection and identification of C. jejuni.
Qian
Hongliang
H
Animal Health Biotechnology Group, Temasek Life Sciences Laboratory, 1 Research Link, National University of Singapore, Singapore 117604, Republic of Singapore.
Pang
Ervinna
E
Du
Qingyun
Q
Chang
Jason
J
Dong
Jin
J
Toh
Say Ling
SL
Ng
Fook Kheong
FK
Tan
Ai Ling
AL
Kwang
Jimmy
J
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000276:immunology; D000918:Antibody Specificity; D001426:Bacterial Proteins / Q000008:administration & dosage / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D016123:Campylobacter jejuni / Q000276:immunology*; D018604:Epitope Mapping; D000939:Epitopes / Q000737:chemistry / Q000276:immunology*; D006801:Humans; D006825:Hybridomas; D007114:Immunization; D015151:Immunoblotting; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D018272:Porins / Q000008:administration & dosage / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D011994:Recombinant Proteins / Q000008:administration & dosage / Q000235:genetics / Q000276:immunology; D017422:Sequence Analysis, DNA
D000911:Antibodies, Monoclonal; D001426:Bacterial Proteins; D000939:Epitopes; D018272:Porins; D011994:Recombinant Proteins; C095080:major outer membrane protein, Campylobacter jejuni