Disclosed are a method for detecting a biomolecule including: immobilizing a nucleic acid aptamer capable of specifically binding to a biomolecule to be detected on the surface of a bead on which fluorophores are arranged; hybridizing the nucleic acid aptamer with a guard nucleic acid (g-nucleic acid) labeled with a quencher to quench fluorescence; and reacting a sample including the biomolecule to be detected with the nucleic acid aptamer and detecting a fluorescence signal emitted as the biomolecule binds with the nucleic acid aptamer and the g-nucleic acid labeled with the quencher is separated, and a device for detecting a biomolecule for conducting the detection method. The present disclosure allows for effective, convenient and fast detection of the biomolecule to be detected, enables quantitative analysis, and enables detection of even a trace amount of sample.