The goal of this proposal is to develop the instrumentation and methodology necessary to make direct mechanical measurement of inter- and intra-molecular forces, F-SPEC, a user friendly, inexpensive and broadly applicable tool for molecular analysis and high throughput molecular screening. This novel approach is based on the principles of the atomic force microscope (AFM) which is capable of measuring forces in the pico Newton range and transducing these forces into macroscopic signals. In the proposed program, a simple ultra-sensitive and cost effective F-SPEC device will be constructed and tested. This device will include a flow chamber and a mechanism for temporary immobilization of microparticles to allow rapid, sequential analysis of molecular species tethered to the microparticles. The system will be evaluated using two molecular systems previously studied by F-SPEC; avidin/biotin and antibody/antigen molecular pairs. Methods will be further developed using DNA-protein and intramolecular DNA hairpin molecular systems. These efforts will result in the production of a technology for measuring forces in and between molecules, that will have applications in a large number of biomedically important areas including basic biomolecular analysis, signal transduction, immunology and immunodiagnostics, pharmacology, drug discovery and high throughput screening of surface immobilized combinatorial molecular libraries.
