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Mapping the Binding Site of an Aptamer on ATP Using MicroScale Thermophoresis
Published on: January 7, 2017
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Thermophoresis in nanoliter droplets to quantify aptamer binding
Susanne A I Seidel1, Niklas A Markwardt, Simon A Lanzmich
1Systems Biophysics, Physics Department, NanoSystems Initiative Munich and Center for Nanoscience, Ludwig-Maximilians-University Munich, Amalienstrasse 54, 80799 Munich (Germany) http://www.biosystems.physik.lmu.de.
Angewandte Chemie (International Ed. in English)
|June 5, 2014
Summary
This study miniaturizes biomolecule interaction analysis using acoustic droplet robotics, reducing sample volume by 50x for high-throughput thermophoresis. This enables sensitive, low-volume screening for drug discovery and diagnostics.
Area of Science:
- Biophysics
- Biochemistry
- Analytical Chemistry
Background:
- Biomolecule interactions are crucial for pharmacology and diagnostics.
- Thermophoresis quantifies these interactions by measuring molecule movement in a temperature gradient.
- Traditional methods require large sample volumes (≥0.5 μL).
Purpose of the Study:
- To develop a miniaturized thermophoresis assay for biomolecule interaction analysis.
- To significantly reduce sample consumption for high-throughput screening.
- To enable novel applications in diagnostics and drug discovery.
Main Methods:
- Utilized acoustic droplet robotics to generate 10 nL droplets, a 50x reduction in sample volume.
- Stabilized droplets in an oil-surfactant mixture for analysis.
- Employed IR laser heating, fluorescence microscopy, and numerical simulation to analyze thermophoresis, Marangoni flow, and concentration distribution.
Main Results:
- Successfully quantified biomolecule interactions (AMP-aptamer affinity, cooperativity, buffer dependence) in 10 nL droplets.
- Demonstrated the method's sensitivity and accuracy at the nL scale.
- Validated the high-throughput and automation-friendly nature of the miniaturized assay.
Conclusions:
- Miniaturized thermophoresis using acoustic droplet robotics enables ultra-low volume analysis of biomolecule interactions.
- The 1536-well plate format facilitates high-throughput and automation, suitable for drug discovery and diagnostics.
- This approach opens avenues for innovative applications, including assays in human serum and label-free screening.
