Poster abstracts
Poster number 3 submitted by Janan Alfehaid
Reusable Microfluidic Chambers for Single-Molecule Microscopy
Janan Alfehaid (Department of Physics, Kent State University), Sineth G. Kodikara (Department of Physics, Kent State University), Tuqa Alhajri (Department of Physics, Kent State University), Mohammad Lutful Kabir (Department of Chemistry and Biochemistry, Kent State University), Hamza Balci (Department of Physics, Kent State University)
Abstract:
Maintaining a stable environment in single-molecule microfluidic chambers with surface-bound molecules involves time-consuming cleaning and surface passivation. Even with such measures, variations in non-specific binding and background signals are often seen across different chambers. Reusing these chambers without surface degradation offers both practical and fundamental benefits, but it requires removing surface-bound molecules like DNA, proteins, lipids, or nanoparticles. Biotin-streptavidin is a common method for attaching these molecules since biotin can easily be incorporated. In this study, we demonstrate through single-molecule fluorescence experiments that chambers can be effectively reset and reused at least 10 times by using photocleavable biotin (PC-biotin) and UV light. Unlike other methods, this approach avoids harsh chemical treatments. We show that a 5-minute exposure to UV light at a specific wavelength successfully removes all bound molecules using various PC-biotin attachment chemistries. Non-optimal wavelengths and light sources showed varying degrees of success. Importantly, our method causes no detectable surface degradation, as confirmed by the low non-specific binding of fluorescently labeled DNA and proteins, as well as the recovery of DNA secondary structure and protein activity. This resetting process is fast, efficient, cost-effective, and broadly applicable to a wide range of single-molecule experiments due to the common use of biotin-streptavidin attachments.
Keywords: single molecule, chamber recycling, UV-exposure