Kinetic analysis of monoclonal antibody-antigen interactions with a new biosensor based analytical system.
An automated biosensor system for measuring molecular interactions has been used to study the kinetics of monoclonal antibody-antigen reactions. The system combines a microfluidic unit in contact with a sensor surface for surface plasmon resonance detection. The specificity of the surface is determined by the operator. Antibody or antigen is immobilised in a dextran matrix attached to the sensor surface. The interaction of matrix bound antibody or antigen with the corresponding partner in solution is monitored in real time. None of the interacting molecules needs to be labelled and it is not necessary to determine the concentration of the the matrix bound component in advance. Two systems were studied: matrix bound monoclonal antibodies (MAbs) interacting with HIV-1 core protein p24 and immobilised aminotheophylline reacting with MAbs. Control of the amount of immobilised ligand and reusable sensor surfaces permits the comparison of different MAbs reacting with antigen under almost identical conditions. Differences in affinity and reaction rates are immediately apparent. The calculated association rate constants for p24 MAbs ranged from 3 x 10(4) - 7.4 x 10(5) M-1 s-1 and for theophylline MAbs association rate constants as high as 1 x 10(6) M-1 s-1 were encountered. The calculated dissociation rate constants were in the region 2 x 10(-4) s-1 to 2 x 10(-2) s-1.
Karlsson
R
R
Pharmacia Biosensor AB, Uppsala, Sweden.
Michaelsson
A
A
Mattsson
L
L
D000911:Antibodies, Monoclonal / Q000276:immunology*; D000915:Antibody Affinity; D000937:Antigen-Antibody Reactions; D015374:Biosensing Techniques; D015483:HIV Antibodies / Q000276:immunology; D016655:HIV Core Protein p24 / Q000276:immunology; D007163:Immunosorbent Techniques; D066298:In Vitro Techniques; D007700:Kinetics; D012031:Refractometry; D013806:Theophylline / Q000276:immunology
D000911:Antibodies, Monoclonal; D015483:HIV Antibodies; D016655:HIV Core Protein p24; D013806:Theophylline