The immune system includes a wide variety of cells that communicate with each other via adhesion and signaling molecules. Many of these molecules behave as 'nanomachines' that connect, sense, move, actuate, to name a few of their functions. To understand the inner workings of such nanomachines requires the characterization of kinetic properties that govern their interactions. We have used atomic force microscopy (AFM) to directly measure mechanistically the behavior of several immune system receptors at the level of single pair of interacting molecules, which extends and complement the ensemble assays commonly used in biochemistry and biophysics. These experiments are laborious and time-consuming due to the requirement for a large ensemble of stochastic data. To address the throughput bottleneck, we will develop arrays of AFM with sufficient sensitivity, bandwidth, and low noise that are suitable for single molecule experiments. This will be done by integrating several existing technologies - cantilever array, photodetector array, and functionalized tip arrays on substrates with a new technology - acoustic radiation pressure (ARP) actuator array. The ARP-driven AFM arrays - protein chips - will enable a large number of parallel single molecular experiments at the same time instead of using a single AFM to conduct them sequentially over a long period of time. The ARP actuation technology will be tested in a single AFM and subsequently in the form of 1D and 2D AFM arraysfor their ability to perform single molecule experiments. Using these AFM arrays, we will characterize the force regulation of kinetics of T cell receptor-ligand interactions. Better understanding of the properties and functions of these molecules may lead to the development of diagnostic and therapeutic strategies for immune system dysfunctions. The proposed research may also have significant commercial value because of the potential application of the AFM array technology to drug discovery and high speed drug screening. Moreover, protein chips such as this may provide means for the transduction of molecular recognition signals into mechanical, optical, and electrical signals through a natural interface to link individual biomolecular functions to microelectromechanical systems, which may have far-reaching implications to nanobiotechnology and biological/chemical warfare identification for national defense.
