Disclosed herein is a method of determining a concentration of a subject based on fraction bound measurement. The method of determining a concentration of a subject based on fraction bound measurement may include fixing a ligand to a surface of an optical device, measuring a fraction bound of a subject to be detected based on an optical signal when the subject reacts to the ligand fixed to the surface of the optical device, and determining a relative value of a concentration of the subject based on a ratio of measured values of the fraction bounds of the subject and a reference signal.