Improvement of Immunoglobulin M Capture Immunoassay Specificity: Toxoplasma Antibody Detection Method as a Model

Improvement of Immunoglobulin M Capture Immunoassay Specificity: Toxoplasma Antibody Detection Method as a Model Tamara Tuuminen * , Helena Seppänen , Eeva-Marja Pitkänen , Pekka Palomäki , and Kirsti Käpyaho Labsystems Research Laboratories, Labsystems OY, 00881 Helsinki, Finland ABSTRACT In the Toxoplasma gondii immunoglobulin M (IgM) capture fluorometric enzyme immunoassay used as a model, nonspecific responses due to the binding of human IgM to horseradish peroxidase (HRP) conjugates were observed despite the removal of the Fc portion of the immunoglobulin. This interaction may be mediated through the binding of human IgM to the HRP moiety of the conjugate. Addition of polymerized HRP into the reaction mixture reduced nonspecific signals in the majority of low false-positive serum reactions. Other plausible sites of interaction are conserved epitopes of mouse immunoglobulins presenting antigenic similarities with the allotopes of other species. Fragmentation of mouse antimicrobial IgG to Fab′ and selection of proper conjugation procedure improved assay specificity.