Enhancement of the antigen-binding capacity of incomplete IgG antibodies to Brucella melitensis through Fc region interactions with staphylococcal protein A.
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, guanidine 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.
Jagannath
C
C
Division of Zoonoses, National Institute of Communicable Diseases, New Delhi, India.
Sehgal
S
S
D000907:Antibodies, Bacterial / Q000276:immunology*; D000915:Antibody Affinity; D000942:Antigens, Bacterial / Q000276:immunology; D001710:Biotin; D002002:Brucella / Q000276:immunology; D002006:Brucellosis / Q000175:diagnosis* / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D006386:Hemagglutination Tests; D006801:Humans; D007141:Immunoglobulin Fc Fragments / Q000378:metabolism*; D007074:Immunoglobulin G / Q000276:immunology*; D008024:Ligands; D012698:Serologic Tests; D013205:Staphylococcal Protein A / Q000378:metabolism / Q000494:pharmacology*
D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D007141:Immunoglobulin Fc Fragments; D007074:Immunoglobulin G; D008024:Ligands; D013205:Staphylococcal Protein A; D001710:Biotin