The humoral response to in vitro generated parasite antigens is enhanced by the removal of a defined media component prior to immunization.

Concentrated culture supernatants containing the extracellular products (ECP) of the protozoan oyster parasite Perkinsus marinus were used to immunize mice. This preparation, produced by ultrafiltration, was found to be both poorly immunogenic and toxic to experimental animals. The possibility that these effects were due to toxic parasite products and/or medium constituents was examined. Co-administration of this material with highly immunogenic oyster hemolymph caused a substantive suppression of the specific antibody response to hemolymph, as well as a decrease in the number of epitopes recognized. Potential protein/protease toxin-mediated causes of the immunosuppression were addressed by heat denaturation and proteolytic inhibition of the concentrate; neither substantially enhanced immunogenicity. Analysis of media constituents revealed that the known immunomodulatory surfactant, Pluronic F-68 (PF68), used in the defined lipid concentrate supplement, was capable of eliciting significant immunosuppression. Although isolated protein antigens from P. marinus ECP remain highly immunosuppressive, separation of the protein antigens from the PF68 has enabled production of polyclonal antisera with a broader recognition of antigens.