Free-Solution Comparison of DNA Hybridization using Isothermal Titration Calorimetry and Backscattering Interferometry
The characterization of biomolecular binding interactions is crucial to the understanding of biological processes and comparison of therapeutic efficacies. This study uses the novel technique of backscattering interferometry with an ITC benchmark to examine the impact of surface immobilization and the presence of a labeled probe on binding affinity. Specifically, the binding affinity of DNA hybridization with and without a fluorescent probe will be compared using both free-solution and surface-bound methodologies. The results from these experiments show a significant increase in the binding affinity for free-solution, label-free DNA hybridization compared to fluorescently labeled, surface-bound DNA hybridization. Additionally, the utility of backscattering interferometry will be compared to ITC as an analytical technique for studying various intermolecular interactions.