Production and characterization of mouse monoclonal antibodies recognizing human Pan-IgG specific conformational or linear epitopes

ABSTRACT

Pan-IgG specific monoclonal antibodies [MAbs] are essential tools for assessment of humoral immunity, immune deficiency and gammopathy. In this study, four hybridoma clones producing MAbs with different specificities for human IgG isotypes were established. Splenocytes from Balb/c mice immunized with Fc fractions of human IgG were fused with SP2/0 myeloma cells. Hybridoma cells were selected in HAT selective medium and cloned by limiting dilution assay. Antibody-secreting cells were screened by enzyme-linked immunosorbent assay [ELISA] and the specificity of secreted MAbs was further analyzed using a panel of purified myeloma IgG proteins by ELISA and immunoblotting. Cross-reactivity to immunoglobulins [Igs] of other species was studied by indirect ELISA using serum samples collected from 9 animals. Immunoblotting studies revealed recognition of either linear or conformational epitopes by these MAbs. The most abundant cross-reactivity [100%] was observed with monkey Igs, while no cross-reactivity was detected with rabbit, guinea pig, sheep, goat, cat and hen sera. Two of the MAbs crossreacted with either dog or horse sera. The affinity constant of two MAbs was measured by ELISA and found to be 0.74×10[8] M[-1] and 0.96×10[7] M[-1]. Our results indicate that these pan-IgG specific MAbs recognize restricted linear or conformational epitopes located on all human IgG subclasses with no cross-reactivity to IgG from most species studied. These MAbs are potentially useful tools for quantification of total or antigen-specific IgG levels