Preparation of antibody-lectin conjugate using reversibly inactivated concanavalin A.
We have tried to modify the lectin-immunotest first described by Guesdon and Avrameas by use of inactivated ConA for the preparation of the conjugates. The inactivation of ConA took place by removal of the bivalent cations Mn++ and Ca++ by means of dialysis against oxalic acid at pH 4.0. The inactivated ConA was coupled to an anti-rabbit IgG antibody using glutaraldehyde. Following the antigen-antibody reaction the lectin could be reactivated. The Con A was again able to react with horseradish peroxidase. The reactivation and the binding of the enzyme can be performed as a one-step procedure. In comparison to the conjugate of the same antibody and active ConA the conjugate of inactivated ConA showed the same sensitivity in the determination of the corresponding antigen but the concentration of the antibody in the working dilution had to be increased.
D000818:Animals; D000906:Antibodies; D000937:Antigen-Antibody Reactions; D002118:Calcium / Q000378:metabolism; D003208:Concanavalin A / Q000378:metabolism*; D007074:Immunoglobulin G / Q000378:metabolism*; D007158:Immunologic Techniques; D008345:Manganese / Q000378:metabolism; D011489:Protein Denaturation; D011817:Rabbits; D013997:Time Factors
D000906:Antibodies; D007074:Immunoglobulin G; D003208:Concanavalin A; D008345:Manganese; D002118:Calcium