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Kowalski A.♦, Piros T.♦, Błoński S., Kurniawan T., Korczyk P., Hołyst R.♦, Microfluidic-based platform for the semi-automated study of concentration, diffusion, and equilibrium binding of biomolecules,
Chemical Engineering Journal, ISSN: 1385-8947, DOI: 10.1016/j.cej.2025.169512, Vol.524, pp.169512-1-10, 2025 Abstract:
We introduce a novel microfluidic-based platform that automates single-molecule-sensitive measurements of biomolecular interactions using Fluorescence Correlation Spectroscopy (FCS) combined with FRET and Molecular Brightness Analysis (MBA). The developed system allows for the simultaneous analysis of biomolecule concentration, diffusion coefficient, and equilibrium constant in nanoliter-volume droplets. At the same time, it speeds up the measuring procedure by 5 times compared to the standard one, reduces reactant consumption by 2000 times compared to conventional cuvette-based FCS measurements, and limits the researcher’s intervention to setting up the experiment and calibrating the microscope. We show the performance of the platform by studying DNA–DNA complex formation in response to ionic strength changes, as well as interactions of idarubicin with various nucleic acid complexes. The proposed solution paves the way for the automated optimization of non-covalent reaction conditions, leveraging non-invasive, high-precision statistical methods. Keywords:
Fluorescence correlation spectroscopy, Microfluidics, Biomolecular interactions, Automation, Diffusion coefficient, Equilibrium constant, DNA hybridization Affiliations:
| Kowalski A. | - | other affiliation | | Piros T. | - | other affiliation | | Błoński S. | - | IPPT PAN | | Kurniawan T. | - | IPPT PAN | | Korczyk P. | - | IPPT PAN | | Hołyst R. | - | other affiliation |
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