Kinetic microscale thermophoresis

Stein, Julian A. C.; Ianeselli, Alan und Braun, Dieter (2021): Kinetic microscale thermophoresis. In: Angewandte Chemie

Abstract

We established an extension of Microscale Thermophoresis (MST) to measure binding kinetics together with binding affinity in a single experimental run, by increasing the thermal dissipation of the sample. After the switch-off of an IR laser, that locally heated the sample, the temperature re-equilibrated within 250 ms. The kinetic relaxation fingerprints were extracted from the fluorescence changes back to thermodynamic equilibrium. We measured DNA hybridization on-rates and off-rates in the range between 104-106 M-1s-1 and 10-4-10-1 s‑1, respectively. We observed the expected exponential dependence of the DNA hybridization off-rates on salt concentration, strand length and inverse temperature. The measured on-rates showed a linear dependence on salt and weak if no dependence at all on length and temperature. For biological binding reactions with sufficient enthalpic contributions, Kinetic MST offers a robust and immobilization-free determination of kinetic rates and binding affinity and also in crowded solutions.

• Dokumententyp: Zeitschriftenartikel
• EU Funded Grant Agreement Number: 787356
• EU-Projekte: Horizon 2020 > ERC Grants > ERC Advanced Grant > ERC Grant 787356: EvoTrap - Mechanisms to emerge and replicate the first sequence information of life in geothermal microfluidics of early Earth
• Fakultät: Physik
• Fakultätsübergreifende Einrichtungen: Center for NanoScience (CENS)
• Themengebiete: 500 Naturwissenschaften und Mathematik > 530 Physik
• Sprache: Englisch