A hydrogel-based interdigitated microelectrode biosensor is disclosed. The hydrogel-based interdigitated microelectrode biosensor includes: a first interdigitated microelectrode having a plurality of first protrusion electrodes arranged in a comb-like shape on a substrate; a second interdigitated microelectrode facing the first interdigitated microelectrode and having a plurality of second protrusion electrodes arranged in a comb-like shape on the substrate, the second protrusion electrodes being arranged alternately with the first protrusion electrodes of the first interdigitated microelectrode; and a hydrogel filled in a space between the first and second interdigitated microelectrodes arranged alternately with each other. The hydrogel is provided between the interdigitated microelectrodes such that the presence and concentration of a biological substance, such as a protein, are detected by measuring the impedance between the interdigitated microelectrodes.