The present invention discloses an electrode of a biosensor, a manufacturing method thereof, and a biosensor thereof. The electrode of the biosensor comprises a flexible insulation layer, a resin layer, a first metal layer, a second metal layer, a third metal layer and a fourth metal layer. The resin layer is disposed between the flexible insulating layer and the first metal layer. The second metal layer is disposed between the first metal layer and the third metal layer, and the fourth metal layer is disposed on the third metal layer. The material of the first metal layer comprises copper foil. The material of the second metal layer comprises palladium. The material of the third metal layer comprises nickel, and the material of the fourth metal layer comprises gold or palladium. The electrode further comprises a biological active substance immobilized on the surface of the plurality of metal layers.