Evidence that the Fc region of autologous rabbit IgG isolated before and after hyperimmunization is structurally different: recognition by rheumatoid factor and monoclonal antibodies

Abstract

Autologous hyperimmune (HI) and pre-inoculation (PI) rabbit Fc gamma populations were found to be conformationally different by spectroscopic measurements, and antigenically different by measurements which examined rheumatoid factor (RF) and monoclonal antibody (MoAb) binding specificity for both populations in ELISA. Circular dichroism spectra of HI rabbit Fc gamma (prepared from animals after hyperimmunization with streptococcal vaccine) were both qualitatively and quantitatively different, particularly in the 225-228 nm range, in comparison to both homologous normal and autologous PI Fc gamma. Binding studies in ELISA showed that affinity constants obtained for reactions of both rabbit RF and various murine MoAb with HI IgG and Fc were approximately 10-fold higher relative to those observed for PI IgG and Fc. Enzymatic deglycosylation of HI and PI Fc gamma led to elimination of CD spectral differences. Further, association constants obtained for RF and MoAb reactions with deglycosylated (sialic acid and galactose removed) PI Fc gamma were equivalent to those obtained in the presence of untreated HI Fc gamma. Together, these results suggest the complex oligosaccharide structure of rabbit IgG may play a significant role in the expression of Fc gamma determinants, and alteration of this structure under hyperimmune or other conditions may be related to induction of an RF response.