Monoclonal anti-peroxidase isotype switch variants. Applications in studies of protein A binding and characterization of rat monoclonal antibodies.
A series of heavy chain isotype switch variants was derived from a hybridoma cell line secreting monoclonal antibodies specific for horseradish peroxidase. By the combined use of sensitive isotype-specific ELISAs and sequential sublining IgG2b, IgG2a, IgE and Iga anti-peroxidase-producing variants were successively isolated out of IgG1-secreting parental cells. The anti-peroxidase isotype variant antibodies are particularly appropriate for use in studies of the influence of heavy chain isotype in the effector functions of immunoglobulins. The use of variant antibodies with specificity for an enzyme favors their application in immunoassays because an enzyme-conjugated second antibody is not needed. Here we describe two applications of the anti-peroxidase switch variants. First, the variants are compared with respect to their affinity for Staphylococcus protein A. While IgG1 anti-peroxidase showed weak binding, both IgG2 variants strongly bound to protein A, whereas IgE and IgA variants had no affinity for protein A. Next, the switch variants were used to determine the isotype specificity of rat monoclonal antibodies generated to murine IgE.
D000818:Animals; D000888:Antibodies, Anti-Idiotypic / Q000276:immunology; D000911:Antibodies, Monoclonal / Q000235:genetics / Q000276:immunology*; D000915:Antibody Affinity; D004797:Enzyme-Linked Immunosorbent Assay; D006735:Horseradish Peroxidase / Q000276:immunology*; D006825:Hybridomas / Q000276:immunology; D007073:Immunoglobulin E / Q000276:immunology; D007143:Immunoglobulin Heavy Chains / Q000235:genetics / Q000276:immunology; D007132:Immunoglobulin Isotypes / Q000235:genetics / Q000276:immunology*; D007134:Immunoglobulin Switch Region; D008297:Male; D051379:Mice; D009154:Mutation; D010544:Peroxidases / Q000276:immunology*; D051381:Rats; D011919:Rats, Inbred Strains; D013205:Staphylococcal Protein A / Q000276:immunology
D000888:Antibodies, Anti-Idiotypic; D000911:Antibodies, Monoclonal; D007143:Immunoglobulin Heavy Chains; D007132:Immunoglobulin Isotypes; D013205:Staphylococcal Protein A; C026408:anti-IgE antibodies; D007073:Immunoglobulin E; D006735:Horseradish Peroxidase; D010544:Peroxidases