Microscale Thermophoresis: Immobilization-free Binding Assays

  • Fig. 1: Microscale thermophoresis. (a) Fluorescently labeled molecules are initially distributed evenly and diffuse freely in solution. By switching on the heating with a focussed IR-Laser, the molecules experience the thermophoretic force in the temperature gradient and move out of the heated spot. In the steady state, this movement is counterbalanced by ordinary mass diffusion. After turning off the laser, the particles diffuse back towards a homogeneous distribution. (b) The IR-Laser that generates the heat in the sample is focussed with the same objective used for epi-fluorescence observation.Fig. 1: Microscale thermophoresis. (a) Fluorescently labeled molecules are initially distributed evenly and diffuse freely in solution. By switching on the heating with a focussed IR-Laser, the molecules experience the thermophoretic force in the temperature gradient and move out of the heated spot. In the steady state, this movement is counterbalanced by ordinary mass diffusion. After turning off the laser, the particles diffuse back towards a homogeneous distribution. (b) The IR-Laser that generates the heat in the sample is focussed with the same objective used for epi-fluorescence observation.

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.

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