Production of Methotrexate antiserum in rabbits: the significance of immunogen solubility, hapten content, and mode of administration on the antibody response

A method for raising specific antibodies of high avidity against Methotrexate in rabbits is described. The antibody response was tested after intramuscular and after multi-site intradermal adminstration of three different immunogens. Antisera of high specificity and avidity were obtained by multi-site intradermal administration of a water-soluble immunogen with high hapten content (40 mol hapten/mol protein). A water-soluble immunogen with low hapten content (15-18 mol hapten/mol protein) resulted in antisera of high specificity but low avidity. No antibody response was obtained with a water-insoluble immunogen with high hapten content (40 mol hapten/mol protein).