It is an object of the present invention to provide a method for measuring the interaction between a physiologically active substance immobilized on the surface of a substrate and a test substance, wherein the influence of negative signals intermittently generated due to desorption of the physiologically active substance as a ligand from the surface of the substrate upon the measured value that indicates the binding amount of a test substance as an analyte can be eliminated. The present invention provides a method for measuring the interaction between a physiologically active substance immobilized on the surface of a substrate and a test substance, wherein a calibration curve is produced by using baseline values obtained by repeatedly measuring a single type of solution, and the measurement value of said test substance is calibrated by said calibration curve, so as to obtain the interaction signals of said test substance.