Regensburg 2016 – wissenschaftliches Programm
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CPP: Fachverband Chemische Physik und Polymerphysik
CPP 23: Charged Soft Matter
CPP 23.5: Vortrag
Dienstag, 8. März 2016, 15:15–15:30, H40
Binding quantification by photochemically triggered microscale electrophoresis — •Friederike M. Möller, Michael Kieß, and Dieter Braun — Systems Biophysics, Nanosystems Initiative and Center for NanoScience, LMU Munich, Germany
Intricate feedback situations arise when chemical reactions are coupled to physical transport mechanisms. Self-organized patterns are well known for spatio-temporally controlled reaction-diffusion scenarios mostly in biology, but also in geology and chemistry. Here we use the light triggered dissociation of one reaction to create local electrical fields which enable the measurement of the association strength of another reaction.
In more detail, the photoactive compound o-nitrobenzaldehyde (o-NBA) dissociates upon UV irradiation and releases a proton. Differential diffusion of the charged photoproducts perturbs electroneutrality and generates a local electrical potential Φ on the low mV scale. Biomolecules move within the electrical field by electrophoresis. Due to backdiffusion on the microscale, concentration profiles settle within seconds proportional to exp(−µ/DΔΦ). The diffusive character of transport makes the local, aqueous electrophoresis size selective.
We exploit the size selectivity for the quantification of aptamer-protein affinity in aqueous solution. Using the well characterized thrombin aptamer TBA15 as a proof of principle, we succeed in determining the KD of binding in the nanomolar regime. Our results are in good agreement with literature and thermophoretic binding assays both with and without o-NBA.