Chemistry: analytical and immunological testing – Involving kinetic measurement of antigen-antibody reaction
Patent
1998-10-19
2000-11-07
Housel, James C.
Chemistry: analytical and immunological testing
Involving kinetic measurement of antigen-antibody reaction
436518, 436512, 436513, 436517, 436169, 436170, 436501, 436527, 436805, 436164, 435 4, 435 6, 435 71, 435 75, G01N 33557, G01N 33543, G01N 33566, G01N 2100, C12Q 100
Patent
active
061435740
DESCRIPTION:
BRIEF SUMMARY
TECHNICAL FIELD
The present invention relates to a method of determining solution affinity and kinetic properties for the formation of a reaction complex between an analyte and a binding partner therefor.
BACKGROUND OF THE INVENTION
Apparatus and methods for investigating the binding of analytes in solution to a receptor are known. Recently, surface sensitive measuring techniques using so-called label-free techniques have been developed for measuring and quantifying biomolecular interactions. In these techniques, a receptor capable of binding to an analyte of interest is immobilised to a sensor surface, and binding of the analyte to the receptor is detected as a resulting change of a property of the sensor surface.
One type of such apparatus (with associated computer control and data-processing means), including the commercial instruments BIAcore and BIAlite (BIAcore and BIAlite are trademarks of Pharmacia Biosensor AB, Uppsala, Sweden; BIA stands for biospecific interaction analysis) has been devised, which uses the phenomenon of surface plasmon resonance (SPR) to study the binding of analytes to receptors immobilized on a sensor chip. The apparatus and theoretical background are fully described in the literature (see e.g. Jonsson, U., et al., BioTechniques 11: 620-627 (1991)). Essentially, the technique involves the immobilisation of a receptor to the special surface of a sensor chip, contacting the sensor chip with a flow of sample containing the analyte of interest, and then measuring the change in the surface optical characteristics of the sensor chip arising form the binding of interest.
With such instrumentation, for example, affinity and kinetic analysis of interactions between soluble analytes and their immobilised binding partners may readily be performed. However, in many cases, it would also be interesting to know the true solution affinity and kinetics of the interaction between two species interacting in solution. So far, such analyses have not been done with the above described type of apparatus.
Friguet, B., et al., Anal. Biochem. 210, 344-350 (1993) discloses the determination of the true affinity constant of a monoclonal antibody for its antigen. Aliquots of radiolabeled antigen at a constant concentration are incubated with the monoclonal antibody at various known concentrations of the antibody in large excess over the antigen. When equilibrium has been reached, the concentration of free antigen is determined by the binding to dextrane beads to which the same monoclonal antibody has been covalently attached. However, this approach only gives information on the equilibrium constant and no kinetic information is provided.
SUMMARY OF THE INVENTION
The object of the present invention is therefore to to provide a method of not only determining true affinity properties but also true kinetic properties for the solution interaction between an analyte and a binding partner therefor to thereby among other things be permitted a wider choice of reaction partners than in solid phase interactions and avoid immobilisation artefacts.
According to the invention, this object is basically achieved by determining the variation of the reaction component concentrations over time in contrast to measuring at equilibrium conditions as in the prior art.
The present invention therefore provides a method of determining affinity and kinetic properites for the solution interaction between an analyte and a binding partner therefor, which method comprises the steps of: partner, contacting the resulting reaction solution with (i) immobilized binding partner, or analogue, and/or (ii) immobilised analyte, or analogue, and monitoring the binding of said analyte and/or binding partner in said reaction solution to the respective immobilised species to determine the variation with time of the concentration of free analyte and/or binding partner in said solution; and/or binding partner therefor with (i) immobilized binding partner, or analogue, for said analyte, and/or (ii) immobilised analyte, or analogue, and monitori
REFERENCES:
patent: 5770462 (1998-06-01), Molloy
Giesen et al., Monitoring of unbound protein in vesicle suspensions with off-null ellipsometry, Biochimica et Biophysica Acta 1147:125-131 (1993).
Karlsson et al., Kinetic Analysis of monoclonal antigen-antibody interactions with a new biosensor based analytical system, Journal of Immunological Methods, 145:229-240 (1991).
Karlsson et al., Real-Time Compepitive Kinetic Analysis of Interactions between Low-Molecular Weight Ligands in Solution and Surface-Immobilized Receptors, Analytical Biochemistry 221:142-151 (1994).
Paek et al., Modeling of Immunosensors Under Nonequilibrium Conditions, Analytical Biochemistry 196:319-325 (1991).
Azimzadeh et al., "Operational Aspects of Antibody Affinity Constants Measured by Liquid-phase and Solid-phase Assays," Journal Of Molecular Recognition 5:9-18, 1992.
National Library of Medicine, file Medline, Medline Accession No. 95397936, Morton et al., "Interpreting complex binding kinetics from optical biosensors: a comparsion of analysis by linearization, the integrated rate equation, and numerical integration," Anal. Biochem. 227(1):176-185, 1995. Abstract Only.
National Library of Medicine, file Medline, Medline Accession No. 93289960, Friquet et al., "A radioimmunoassay-based method for measuring the true affinity of a monoclonal antibody with trace amounts of radioactive antigen: illustration with the products of a cell-free protein synthesis system," Anal. Biochem. 210(2):344-350, 1993. Abstract Only.
Nelson and Long, "Solution-Phase Equilibrium Binding Interaction of Human Protein S with C4b-Binding Protein," Biochemistry 30:2384-2390, 1991.
Glaser Ralf W.
Karlsson Robert
Lof.ang.s Stefan
Biacore AB
Gabel Gailene R.
Housel James C.
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