Development of amperometric and microgravimetric immunosensors and reversible immunosensors using antigen and photoisomerizable antigen monolayer electrodes

Antigen monolayers assembled onto Au-electrodes or Au-electrodes associated with quartz crystals act as active interfaces for the amperometric or microgravimetric analysis of the complementary antibody and provide the grounds for the development of electrochemical and piezoelectric immunosensors. The antigen monolayer of Nε-2,4-dinitrophenyl-L-lysine is assembled on an electrode. The anti-dinitrophenyl antibody, anti-DNP-Ab, is sensed by the antigen monolayer, and the formation of the antigen-antibody complex at the monolayer interface is probed by the insulation of the electrode toward a redox probe in the electrolyte solution. The formation of the antibody-antigen complex is amplified by the application of the anti-antibody or the use of a ferrocene-functionalized redox-enzyme, glucose oxidase, as redox probe. A 3,5-dinitrosalicylic acid antigen monolayer bound to Au-electrodes associated with a quartz crystal is used as active interface for the microgravimetric, quartz-crystal-microbalance analysis of the anti-DNP-Ab. Photoisomerizable antigen monolayer electrodes provide the basis for tailoring reversible immunosensors. The dinitrospiropyran monolayer, SP-state, is assembled on Au-electrodes or quartz crystals. The monolayer exhibits reversible photoisomerizable features, and irradiation of the SP-monolayer, 360 < λ < 380 nm, yields the protonated merocyanine monolayer, MRH⁺-state. Further irradiation of the MRH⁺-monolayer electrode, λ >495 nm, restores the SP-monolayer electrode. The SP-monolayer acts as antigen for anti-DNP-Ab, whereas the MRH⁺-monolayer lacks antigen properties for anti-DNP-Ab. This enables the amperometric or piezoelectric transduction of the formation of the antigen-anti-DNP-Ab complex at the SP-monolayer interface. By photoisomerization of the monolayer to the MRH⁺-state, the Ab is washed-off from the sensing interface. Subsequent light-induced isomerization of the monolayer to the SP-state regenerates the electrochemically or piezoelectrically active sensing interfaces.