Microscale Thermophoresis: Immobilization-free Binding Assays

摘要

Microscale thermophoresis is an immobilization-free, low volume method for the analysis of biomolecules. It uses the directed motion of molecules in microscopic temperature gradients. The method is not only sensitive to changes in mass, size or charge of a molecule, but also measures changes in the hydration-shell of a molecule. The presented experiments highlight the simplicity and broad scope of the method. Methods to detect biomolecules or to analyze biomolecule interactions are of great importance to molecular biology and the development of pharmaceuticals. In this work, we use microscale thermophoresis (MST), an all-optical, immobilization-free and low volume method (<1 μl) to characterize biomolecules and study their interaction in a simple Mix-and-Read fashion. We demonstrate the feasibility of the method by detecting DNA sequences and measuring the stoichiometry of an high affinity interaction. As the name already implies MST is based on thermophoresis, the directed movement of particles in a temperature gradient. A temperature difference ΔT in space leads to a depletion of the solvated biomolecules in the region of elevated temperature, quantified by the Soret coefficient ST: c_(hot)/c_(cold) = exp(-S_TΔT). This thermophoretic depletion depends on the interface between molecule and solvent. Under constant buffer conditions, thermophoresis probes the size, charge and solvation entropy of the molecules.