Competitive enzyme-linked immunosorbent assay using monoclonal antibodies to the Brucella melitensis BP26 protein to evaluate antibody responses in infected and B. melitensis Rev.1 vaccinated sheep
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Abstract
Competitive enzyme-linked immunosorbent assay (C-ELISA) was performed using 15 monoclonal
antibodies (MAbs), specific for Brucella BP26 (previously also called CP28), a periplasmic
protein antigen, to investigate antibody responses in naturally and B. melitensis
H38 experimentally infected and B. melitensis Rev.1 vaccinated sheep. The antigen
preparation consisted of cytosoluble protein extract (CPE) of B. melitensis B115.
By combining the C-ELISA results of several MAbs, a high percentage of naturally infected
animals were detected which showed different status in the current conventional diagnostic
tests. Indeed, 90% of sheep which were positive in the conventional bacteriological
and serological tests were positive in C-ELISA. 72% of the bacteriologically negative
but serologically and delayed type hypersensitivity positive sheep were also positive
in the C-ELISA. Moreover, 79% of the bacteriologically and serologically negative
sheep but delayed type hypersensitivity positive were also detected by C-ELISA. Thus,
these results confirmed the importance of BP26 as a frequently recognized target of
the humoral immune response of infected sheep. The 8 B. melitensis H38 experimentally
infected sheep showed various degrees of antibody responses at the 90th day after
infection, which was delayed in comparison to that against O-polysaccharide (O-PS).
Of the 15 MAbs tested, only one MAb was weakly inhibited (20 to 35% inhibition) by
56% of negative control sera. Furthermore, no antibody response against BP26 was detected
in B. melitensis Rev.1 vaccinated sheep. Results of the C-ELISA with the 15 MAbs showed
individual variability of the antibody responses against BP26. Thus, it is suggested
that several epitopes of BP26 are of interest for diagnosis of B. melitensis infection
in sheep.