Abstract
BackgroundIn the past two decades, methods have been developed to measure the mechanical properties of single biomolecules. One of these methods, Magnetic tweezers, is amenable to acquisition of data on many single molecules simultaneously, but to take full advantage of this "multiplexing" ability, it is necessary to simultaneously incorporate many capabilities that have been only demonstrated separately.MethodsOur custom built magnetic tweezer combines high multiplexing, precision bead tracking, and bi-directional force control into a flexible and stable platform for examining single molecule behavior. This was accomplished using electromagnets, which provide high temporal control of force while achieving force levels similar to permanent magnets via large paramagnetic beads.ResultsHere we describe the instrument and its ability to apply 2–260 pN of force on up to 120 beads simultaneously, with a maximum spatial precision of 12 nm using a variety of bead sizes and experimental techniques. We also demonstrate a novel method for increasing the precision of force estimations on heterogeneous paramagnetic beads using a combination of density separation and bi-directional force correlation which reduces the coefficient of variation of force from 27% to 6%. We then use the instrument to examine the force dependence of uncoiling and recoiling velocity of type 1 fimbriae from Eschericia coli (E. coli) bacteria, and see similar results to previous studies.ConclusionThis platform provides a simple, effective, and flexible method for efficiently gathering single molecule force spectroscopy measurements.
Highlights
In the past two decades, methods have been developed to measure the mechanical properties of single biomolecules
The magnetic tweezer (MT) is a relatively recently developed instrument that allows the examination of hundreds of single molecule measurements simultaneously [9,10,11]
This ability to acquire measurements of the same phenomena simultaneously is known as multiplexing, and can significantly reduce data acquisition time when compared to other Single molecule force spectroscopy (SMFS) instruments
Summary
In the past two decades, methods have been developed to measure the mechanical properties of single biomolecules One of these methods, Magnetic tweezers, is amenable to acquisition of data on many single molecules simultaneously, but to take full advantage of this "multiplexing" ability, it is necessary to simultaneously incorporate many capabilities that have been only demonstrated separately. Since some studies require hundreds to thousands of measurements to accurately model force dependence, The magnetic tweezer (MT) is a relatively recently developed instrument that allows the examination of hundreds of single molecule measurements simultaneously [9,10,11] This ability to acquire measurements of the same phenomena simultaneously is known as multiplexing, and can significantly reduce data acquisition time when compared to other SMFS instruments. The chamber can be as simple as two glass slides separated by Johnson et al Journal of Biological Engineering (2017) 11:47
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