TWENTY-THIRD ANNUAL EASTERN FISH HEALTH WORKSHOP JOHN CARVER INN, PLYMOUTH, MA 30 MARCH - 2 APRIL, 1998 Immunopurification Of Salmonid Antibodies Maintaining Intact Affinity Profiles H. L. Zhang and S. L. Kaattari Department of Environmental Sciences, School of Marine Science, Virginia Institute of Marine Science, College of William & Mary, Gloucester Point, VA 23062 Antibody affinity determines the strength of antibody-antigen interaction. The greater the affinity, the tighter the antibody-antigen bonding, and thus, in the case of neutralizing antibody, the higher the neutralization efficiency. Therefore, affinity analysis is essential for understanding the nature of humoral immunity elicited by infection or vaccination and should be an important criteria to evaluate the quality of natural or engineered antibodies used in immunotherapy. It is demonstrated here that a commonly employed antibody purification procedure masks higher affinity subpopulations by retention of antigen in their binding sites. Ion exchange-mediated antigen removal is required to free the blocked binding sites. Specifically, these studies demonstrate that even after extensive dialysis, the TNP hapten used to elute anti-TNP antibodies from an immunoabsorbent column was still bound to the highest affinity antibodies. Consequently, these antibodies were blocked from reacting in subsequent affinity analysis, thereby inaccurately portraying a lower affinity profile. However, the retained hapten can be removed by passing the eluants through an anion exchange matrix and the original affinity profile was correspondingly restored.   Return to 23rd Annual Eastern Fish Health Workshop Return to Leetown Science Center Home Page