The matrix effects on kinetic rate constants of antibody-antigen interactions reflect solvent viscosity.
This study describes the influence of different matrices on two model antibody-antigen interactions; that between beta2microglobulin and anti beta2microglobulin, and that of rabbit anti mouse Fc fragment (RAMFc) with mouse IgG. The matrices investigated were; phosphate-buffered saline pH 7.4 containing 0.05% Tween 20 detergent, horse serum, a 50:50 mixture of phosphate-buffered saline/Tween 20 and horse serum, and four glycerol solutions of differing concentrations. A recently developed optical biosensor, the IAsys, was used to monitor the interactions in real-time and provide precise determinations of k(ass), k(diss) and KA values. The results show that the rates of association and dissociation for the two different antibody:antigen models are significantly affected by the surrounding matrix. Glycerol of known viscosity was used as a matrix in both models to show that this effect is attributable to the viscosity as opposed to proteins present in the matrix. The viscosity of the matrix has also been shown to have an apparent influence upon the overall equilibrium/affinity constant for the interaction, with measurements of KA tending to increase with viscosity. The significant effects of matrix on kinetic rate constants for antibody-antigen interactions shown here have important implications in the use of immunoassays where non-equilibrium measurements are made in serum matrices.
Department of Clinical Biochemistry, St Bartholomew's and the Royal London School of Medicine and Dentistry, UK.
C L
CL
Morgan
D J
DJ
Newman
J M
JM
Burrin
C P
CP
Price
D000818:Animals; D000937:Antigen-Antibody Reactions / Q000187:drug effects*; D016477:Artifacts; D015374:Biosensing Techniques; D005990:Glycerol / Q000494:pharmacology*; D007141:Immunoglobulin Fc Fragments / Q000276:immunology; D007074:Immunoglobulin G / Q000276:immunology; D007700:Kinetics; D051379:Mice; D011817:Rabbits; D012997:Solvents / Q000494:pharmacology*; D014783:Viscosity; D001613:beta 2-Microglobulin / Q000276:immunology
D007141:Immunoglobulin Fc Fragments; D007074:Immunoglobulin G; D012997:Solvents; D001613:beta 2-Microglobulin; D005990:Glycerol