Determination of epitope specificities and affinities of monoclonal antibodies in solution phase using biotin-labeled carcinoembryonic antigen and avidin as precipitating agent. 1989

U Krüger, and L Wickert, and C Wagener
Institute of Clinical Chemistry and Pathobiochemistry, University of Technology, Aachen, F.R.G.

A generally applicable method for the selective precipitation of antigens from solution phase is presented. The method is based on the rapid and quantitative binding of avidin to biotin-labeled antigen in the presence of biotinylated carrier protein. Using carcinoembryonic antigen (CEA) as a model antigen biotin-labeled CEA was precipitated to greater than 97% at amounts of up to 5000 ng/tube. The method was applied to the determination of epitope specificities and affinities of monoclonal anti-CEA antibodies in solution phase. Non-specific precipitation did not exceed 2% for radiolabeled Fab fragments and 6% for radiolabeled IgG. Because antigen-antibody binding takes place in solution, artifacts introduced by the immobilization of reactants to solid supports are omitted.

UI MeSH Term Description Entries
D007140 Immunoglobulin Fab Fragments Univalent antigen-binding fragments composed of one entire IMMUNOGLOBULIN LIGHT CHAIN and the amino terminal end of one of the IMMUNOGLOBULIN HEAVY CHAINS from the hinge region, linked to each other by disulfide bonds. Fab contains the IMMUNOGLOBULIN VARIABLE REGIONS, which are part of the antigen-binding site, and the first IMMUNOGLOBULIN CONSTANT REGIONS. This fragment can be obtained by digestion of immunoglobulins with the proteolytic enzyme PAPAIN. Fab Fragment,Fab Fragments,Ig Fab Fragments,Immunoglobulins, Fab Fragment,Fab Immunoglobulin Fragments,Immunoglobulin Fab Fragment,Immunoglobulins, Fab,Fab Fragment Immunoglobulins,Fab Fragment, Immunoglobulin,Fab Fragments, Immunoglobulin,Fragment Immunoglobulins, Fab,Fragment, Fab,Immunoglobulin Fragments, Fab
D007700 Kinetics The rate dynamics in chemical or physical systems.
D011233 Precipitin Tests Serologic tests in which a positive reaction manifested by visible CHEMICAL PRECIPITATION occurs when a soluble ANTIGEN reacts with its precipitins, i.e., ANTIBODIES that can form a precipitate. Precipitin Test,Test, Precipitin,Tests, Precipitin
D011863 Radioimmunoassay Classic quantitative assay for detection of antigen-antibody reactions using a radioactively labeled substance (radioligand) either directly or indirectly to measure the binding of the unlabeled substance to a specific antibody or other receptor system. Non-immunogenic substances (e.g., haptens) can be measured if coupled to larger carrier proteins (e.g., bovine gamma-globulin or human serum albumin) capable of inducing antibody formation. Radioimmunoassays
D002272 Carcinoembryonic Antigen A glycoprotein that is secreted into the luminal surface of the epithelia in the gastrointestinal tract. It is found in the feces and pancreaticobiliary secretions and is used to monitor the response to colon cancer treatment. Antigens, CD66e,CD66e Antigen,Antigen, CD66e,Antigen, Carcinoembryonic,CD66e Antigens
D000818 Animals Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA. Animal,Metazoa,Animalia
D000911 Antibodies, Monoclonal Antibodies produced by a single clone of cells. Monoclonal Antibodies,Monoclonal Antibody,Antibody, Monoclonal
D000915 Antibody Affinity A measure of the binding strength between antibody and a simple hapten or antigen determinant. It depends on the closeness of stereochemical fit between antibody combining sites and antigen determinants, on the size of the area of contact between them, and on the distribution of charged and hydrophobic groups. It includes the concept of "avidity," which refers to the strength of the antigen-antibody bond after formation of reversible complexes. Affinity, Antibody,Antibody Avidity,Avidity, Antibody,Affinities, Antibody,Antibody Affinities,Antibody Avidities,Avidities, Antibody
D000939 Epitopes Sites on an antigen that interact with specific antibodies. Antigenic Determinant,Antigenic Determinants,Antigenic Specificity,Epitope,Determinant, Antigenic,Determinants, Antigenic,Specificity, Antigenic
D001360 Avidin A specific protein in egg albumin that interacts with BIOTIN to render it unavailable to mammals, thereby producing biotin deficiency.

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