INTELLECTUAL MERIT: Analysis of nucleic acids is becoming increasingly integrated into health care and clinical diagnostics. A technology that especially excels in combining throughput with affordability is multiplexed surface hybridization (SH). Clinical SH tests are becoming available for applications that include cancer diagnostics, cystic fibrosis prescreening, pathogen detection, and assessment of drug metabolism. These tests function by monitoring the extent of hybridization, or association, between two nucleic acid strands, one a "probe" immobilized on a solid material surface and the other a sample "target" sequence present in solution. Arrival of SH technologies in clinical diagnostics greatly intensifies the need for fundamental understanding, so that diagnostic performance can be optimized. This project addresses two outstanding challenges about the material surfaces used in SH applications: (1) the link between hybridization thermodynamics at the surface with those in solution, and (2) the timescales for approaching equilibrium when many target sequences compete for the probes. The first topic is essential for enabling decades of solution hybridization research to be applied to SH applications, for example, to the design of optimized probe sequences and the development of corrective algorithms for cross-hybridization. The second topic centers on identification of rate-limiting bottlenecks in competitive SH, when many target sequences compete for the probes, with the goal to advance strategies for minimizing kinetic biases and for enhancing sensitivity to lower copy target sequences. As part of these studies, benchmark thermodynamic data will be obtained on molecular interactions, including between probes, that affect hybridization reactions on densely modified material surfaces. By elucidating the thermodynamic connection between surface and solution hybridization, and identifying kinetic bottlenecks to equilibration, this project will formulate design principles central to material surfaces used in research and emergent clinical technologies based on surface hybridization.
BROADER IMPACTS: The complex molecular phenomena underlying SH have long hindered development of universal guidelines for diagnostic applications. Results from this project will therefore be immediately useful for design of probe-modified material surfaces as well as for interpretation of experiments performed in hundreds of research and, increasingly, clinical laboratories. The research effort will be integrated with an educational initiative on development of educational gaming software that will introduce general concepts from science and engineering through direct integration into game mechanics. The pedagogical strategy of the software is to develop qualitative familiarity with STEM concepts through an enjoyable gaming experience that can heighten interest in students before they encounter related concepts in formal coursework. The software will be developed in collaboration with the Game Innovation Laboratory at the Polytechnic Institute of NYU, at a level accessible to middle school students. The initial concept for the gaming software is to use fairy tale characters to solve problems through relying on molecular concepts. The software will introduce notions of molecules, of selective interactions between molecules (e.g. assembly of supramolecular structures), and of simple renditions of thermodynamic concepts through use of "molecular bricks" to build fortifications with different energy scores, all as an integral part of game mechanics.
