Multifrequency evaluation of different immunosorbents on acoustic plate mode sensors

Previous studies of acoustic plate modes on ZX-LiNbO3 have indicated that practical mass-sensitive immunosensors can be implemented by using devices with higher frequencies of operation and/or by improving techniques for the immobilization of antibodies. However, it is also known from these studies that the viscoelastic properties of aminosilane films, used for the covalent immobilization of antibodies on the crystal surface, cannot be ignored in the sensor response. In the present work, in an attempt to study the effect of viscoelasticity of the binding film, three different films with different viscoelasticity and binding capacities, an aminosilane, a dextran, and a poly-(etherurethane)-based immunosorbent (XP-5), were prepared on the sensor surface for the immobilization of antibodies. Immunochemical reactions were monitored by the acoustic plate mode sensor at three different frequencies, thus allowing the direct observation of the frequency dependence of mass sensitivity with different films. Depending on the type of immunosorbent, the sensitivity at the third harmonic was enhanced by a factor of 2-5 with respect to the fundamental response. A third acoustic mode at a closely spaced frequency to the third harmonic yielded lower sensitivity values, which indicates that sensitivity depends not only on the frequency of device operation but also on particle displacement amplitude and components of the selected wave. Since antigen binding capacities of the different immunosorbents were determined independently by a modified ELISA test, sensor responses can also be correlated to the immunosorbent structure, and hence the viscoelastic properties. A dual delay line configuration was used which compensates for second-order effects such as temperature variations and nonspecific adsorption.