Monoclonal antibodies recognizing the surface autolysin IspC of Listeria monocytogenes Serotype 4b: epitope localization, kinetic characterization, and cross-reaction studies

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DOIResolve DOI: http://doi.org/10.1371/journal.pone.0055098
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TypeArticle
Journal titlePLoS ONE
ISSN1932-6203
Volume8
Issue2
Article numbere55098
Subjectautolysin; epitope; immunogenic surface protein C; monoclonal antibody; monoclonal antibody M2773; monoclonal antibody M2774; monoclonal antibody M2775; monoclonal antibody M2777; monoclonal antibody M2778; monoclonal antibody M2779; monoclonal antibody M2780; monoclonal antibody M2781; monoclonal antibody M2785; monoclonal antibody M2787; monoclonal antibody M2790; monoclonal antibody M2791; monoclonal antibody M2792; monoclonal antibody M2795; monoclonal antibody M2797; monoclonal antibody M2799; monoclonal antibody M2800; unclassified drug; amino terminal sequence; antibody affinity; antibody specificity; article; bacterium isolate; C terminal cell wall binding domain; controlled study; cross reaction; dissociation constant; enzyme linked immunosorbent assay; epitope mapping; kinetics; Listeria monocytogenes; mass spectrometry; molecular weight; nonhuman; nucleotide sequence; protein domain; protein localization; serotype; surface plasmon resonance; Amino Acid Sequence; Antibodies, Monoclonal; Antibody Specificity; Antigens, Bacterial; Bacterial Proteins; Binding Sites; Cell Wall; Cross Reactions; Enzyme-Linked Immunosorbent Assay; Epitopes; Kinetics; Listeria monocytogenes; Mass Spectrometry; Molecular Sequence Data; N-Acetylmuramoyl-L-alanine Amidase; Protein Binding; Sequence Alignment; Serotyping; Surface Plasmon Resonance
AbstractListeria monocytogenes serotype 4b is responsible for a high percentage of fatal cases of food-borne infection. In a previous study, we created 15 monoclonal antibodies (MAbs) against a ~77 kDa antigen that is associated with the cell surface of live L. monocytogenes serotype 4b cells. Here we report an extensive characterization of these MAbs to further their development as diagnostic reagents. The ~77 kDa target antigen was identified by mass spectrometry and N-terminal sequencing to be IspC, a novel surface associated autolysin. Epitope localization experiments revealed that each of the 15 MAbs recognized the C-terminal cell-wall binding domain of IspC. The presence of IspC was shown to be highly conserved within L. monocytogenes serotype 4b, as evidenced by a strong reaction between anti-IspC MAbs and all 4b isolates. To determine the range of cross-reactivity with other L. monocytogenes serotypes ELISA was used to test each MAb against multiple isolates from each of the L. monocytogenes serotypes. Of the 15 MAbs, five: M2774, M2775, M2780, M2790 and M2797, showed specificity for L. monocytogenes serotype 4b and only cross reacted with serotype 4ab isolates. The kinetics of the interaction between each of the MAbs and IspC was measured using surface plasmon resonance. The MAbs M2773, M2792, M2775, M2797 and M2781 each had very low dissociation constants (4.5 × 10-9 to 1.2 × 10-8 M). While several of these antibodies have properties which could be useful in diagnostic tests, the combined high fidelity and affinity of M2775 for the IspC protein and serotype 4b isolates, makes it a particularly promising candidate for use in the development of a specific L. monocytogenes serotype 4b diagnostic test. © 2013 Ronholm et al.
Publication date
LanguageEnglish
AffiliationNational Research Council Canada (NRC-CNRC); NRC Institute for Biological Sciences (IBS-ISB)
Peer reviewedYes
NPARC number21269638
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Record identifier31baa45a-cbfd-455b-aac4-36bf566de3cd
Record created2013-12-13
Record modified2016-05-09
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