A Biophysics Toolbox for Reliable Data Acquisition and Processing in Integrated Force-Confocal Fluorescence Microscopy

Journal article (2024)

Authors

Z. Liu Kavli institute of nanoscience Delft, BN/Nynke Dekker Lab - AS
E.N.W. van Veen BN/Nynke Dekker Lab - AS , Kavli institute of nanoscience Delft
Humberto Sánchez Kavli institute of nanoscience Delft, BN/Nynke Dekker Lab - AS
H. Sánchez Kavli institute of nanoscience Delft, BN/Nynke Dekker Lab - AS
Humberto Sanchez González Kavli institute of nanoscience Delft, BN/Nynke Dekker Lab - AS
H. Sanchez González Kavli institute of nanoscience Delft, BN/Nynke Dekker Lab - AS
B.P. Solano Hermosilla Kavli institute of nanoscience Delft, BN/Nynke Dekker Lab - AS
F.J.P.M. Palmero Moya BN/Nynke Dekker Lab - AS , Kavli institute of nanoscience Delft
K.A. Mc Cluskey Kavli institute of nanoscience Delft, BN/Nynke Dekker Lab - AS
D.F. Ramirez Montero BN/Nynke Dekker Lab - AS , Kavli institute of nanoscience Delft
T. van Laar Kavli institute of nanoscience Delft, BN/Nynke Dekker Lab - AS
N.H. Dekker BN/Nynke Dekker Lab - AS , Kavli Institute of Nanoscience Discovery, University of Oxford
Research Group
BN/Nynke Dekker Lab (AS) (TU Delft)
More Info
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Published Date

2024

Language

English

Faculty

Applied Sciences

Department

BN/Bionanoscience

Research Group

BN/Nynke Dekker Lab

Abstract

Integrated single-molecule force-fluorescence spectroscopy setups allow for simultaneous fluorescence imaging and mechanical force manipulation and measurements on individual molecules, providing comprehensive dynamic and spatiotemporal information. Dual-beam optical tweezers (OT) combined with a confocal scanning microscope form a force-fluorescence spectroscopy apparatus broadly used to investigate various biological processes, in particular, protein:DNA interactions. Such experiments typically involve imaging of fluorescently labeled proteins bound to DNA and force spectroscopy measurements of trapped individual DNA molecules. Here, we present a versatile state-of-the-art toolbox including the preparation of protein:DNA complex samples, design of a microfluidic flow cell incorporated with OT, automation of OT-confocal scanning measurements, and the development and implementation of a streamlined data analysis package for force and fluorescence spectroscopy data processing. Its components can be adapted to any commercialized or home-built dual-beam OT setup equipped with a confocal scanning microscope, which will facilitate single-molecule force-fluorescence spectroscopy studies on a large variety of biological systems.