TY - JOUR
T1 - Enhancement of the antigen-binding capacity of incomplete IgG antibodies to Brucella melitensis through Fc region interactions with staphylococcal protein A
AU - Jagannath, Chinnaswamy
AU - Sehgal, S.
PY - 1989/11/30
Y1 - 1989/11/30
N2 - Incomplete IgG anti-Brucella antibodies in human sera were detected using both a conventional ELISA and a modified method. In the ELISA procedure serum IgG was allowed to bind solid-phase B. melitensis antigen and, after washing, biotinylated staphylococcal protein A (BioSPA) was used as an Fc-specific tracer followed by streptavidin-HRP conjugate. In the modified method, serum IgG was co-incubated with a defined quantity of BioSPA in the presence of solid-phase antigen. BioSPA bound Fc regions of serum IgG irrespective of antigen specificity whilst antibodies which were specific for Brucella bound the solid-phase antigen through their Fab regions and were detected subsequently by streptavidin-HRP. IgG anti-Brucella antibodies were detectable with a 5-25-fold increase in sensitivity when they were thus 'activated' in situ with BioSPA. In contrast with the IgG antibodies of untreated human sera, BioSPA-activated IgG showed strong antigen binding capacity and resisted the dissociation effect of the chaotropic agent, guadinene hydrochloride. In similar experiments, BioSPA did not enhance the affinity of IgG anti-Salmonella antibodies of human sera towards S. typhi antigen. The activating effect of BioSPA on the incomplete IgG anti-Brucella antibodies from patients with brucellosis possibly involves a re-orientation of Fab sites.
AB - Incomplete IgG anti-Brucella antibodies in human sera were detected using both a conventional ELISA and a modified method. In the ELISA procedure serum IgG was allowed to bind solid-phase B. melitensis antigen and, after washing, biotinylated staphylococcal protein A (BioSPA) was used as an Fc-specific tracer followed by streptavidin-HRP conjugate. In the modified method, serum IgG was co-incubated with a defined quantity of BioSPA in the presence of solid-phase antigen. BioSPA bound Fc regions of serum IgG irrespective of antigen specificity whilst antibodies which were specific for Brucella bound the solid-phase antigen through their Fab regions and were detected subsequently by streptavidin-HRP. IgG anti-Brucella antibodies were detectable with a 5-25-fold increase in sensitivity when they were thus 'activated' in situ with BioSPA. In contrast with the IgG antibodies of untreated human sera, BioSPA-activated IgG showed strong antigen binding capacity and resisted the dissociation effect of the chaotropic agent, guadinene hydrochloride. In similar experiments, BioSPA did not enhance the affinity of IgG anti-Salmonella antibodies of human sera towards S. typhi antigen. The activating effect of BioSPA on the incomplete IgG anti-Brucella antibodies from patients with brucellosis possibly involves a re-orientation of Fab sites.
KW - Avidin-Biotin ELISA
KW - Brucellosis
KW - Incomplete IgG
KW - Protein A
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U2 - 10.1016/0022-1759(89)90361-X
DO - 10.1016/0022-1759(89)90361-X
M3 - Article
C2 - 2600428
AN - SCOPUS:0024788386
SN - 0022-1759
VL - 124
SP - 251
EP - 257
JO - Journal of Immunological Methods
JF - Journal of Immunological Methods
IS - 2
ER -