High performance photonic probes and applications of optical tweezers to molecular motors

Optical tweezers are a sensitive position and force transducer widely employed in physics and biology. In a focussed laser, forces due to radiation pressure enable to trap and manipulate small dielectric particles used as probes for various experiments. For sensitive biophysical measurements, microspheres are often used as a handle for the molecule of interest. The force range of optical traps well covers the piconewton forces generated by individual biomolecules such as kinesin molecular motors. However, cellular processes are often driven by ensembles of molecular machines generating forces exceeding a nanonewton and thus the capabilities of optical tweezers. In this thesis I focused, fifirst, on extending the force range of optical tweezers by improving the trapping e fficiency of the probes and, second, on applying the optical tweezers technology to understand the mechanics of molecular motors. I designed and fabricated photonically-structured probes: Anti-reflection-coated, high-refractive-index, core-shell particles composed of titania. With these probes, I significantly increased the maximum optical force beyond a nanonewton. These particles open up new research possibilities in both biology and physics, for example, to measure hydrodynamic resonances associated with the colored nature of the noise of Brownian motion. With respect to biophysical applications, I used the optical tweezers to study the mechanics of single kinesin-8. Kinesin-8 has been shown to be a very processive, plus-end directed microtubule depolymerase. The underlying mechanism for the high processivity and how stepping is affected by force is unclear. Therefore, I tracked the motion of yeast (Kip3) and human (Kif18A) kinesin-8s with high precision under varying loads. We found that kinesin-8 is a low-force motor protein, which stalled at loads of only 1 pN. In addition, we discovered a force-induced stick-slip motion, which may be an adaptation for the high processivity. Further improvement in optical tweezers probes and the instrument will broaden the scope of feasible optical trapping experiments in the future

An optically actuated surface scanning probe

We demonstrate the use of an extended, optically trapped probe that is capable of imaging surface topography with nanometre precision, whilst applying ultra-low, femto-Newton sized forces. This degree of precision and sensitivity is acquired through three distinct strategies. First, the probe itself is shaped in such a way as to soften the trap along the sensing axis and stiffen it in transverse directions. Next, these characteristics are enhanced by selectively position clamping independent motions of the probe. Finally, force clamping is used to refine the surface contact response. Detailed analyses are presented for each of these mechanisms. To test our sensor, we scan it laterally over a calibration sample consisting of a series of graduated steps, and demonstrate a height resolution of ∼ 11 nm. Using equipartition theory, we estimate that an average force of only ∼ 140 fN is exerted on the sample during the scan, making this technique ideal for the investigation of delicate biological samples

Roadmap for Optical Tweezers 2023

Optical tweezers are tools made of light that enable contactless pushing, trapping, and manipulation of objects ranging from atoms to space light sails. Since the pioneering work by Arthur Ashkin in the 1970s, optical tweezers have evolved into sophisticated instruments and have been employed in a broad range of applications in life sciences, physics, and engineering. These include accurate force and torque measurement at the femtonewton level, microrheology of complex fluids, single micro- and nanoparticle spectroscopy, single-cell analysis, and statistical-physics experiments. This roadmap provides insights into current investigations involving optical forces and optical tweezers from their theoretical foundations to designs and setups. It also offers perspectives for applications to a wide range of research fields, from biophysics to space exploration

Roadmap for optical tweezers

Optical tweezers are tools made of light that enable contactless pushing, trapping, and manipulation of objects, ranging from atoms to space light sails. Since the pioneering work by Arthur Ashkin in the 1970s, optical tweezers have evolved into sophisticated instruments and have been employed in a broad range of applications in the life sciences, physics, and engineering. These include accurate force and torque measurement at the femtonewton level, microrheology of complex fluids, single micro- and nano-particle spectroscopy, single-cell analysis, and statistical-physics experiments. This roadmap provides insights into current investigations involving optical forces and optical tweezers from their theoretical foundations to designs and setups. It also offers perspectives for applications to a wide range of research fields, from biophysics to space exploration.journal articl

Roadmap for optical tweezers

Artículo escrito por un elevado número de autores, solo se referencian el que aparece en primer lugar, el nombre del grupo de colaboración, si le hubiere, y los autores pertenecientes a la UAMOptical tweezers are tools made of light that enable contactless pushing, trapping, and manipulation of objects, ranging from atoms to space light sails. Since the pioneering work by Arthur Ashkin in the 1970s, optical tweezers have evolved into sophisticated instruments and have been employed in a broad range of applications in the life sciences, physics, and engineering. These include accurate force and torque measurement at the femtonewton level, microrheology of complex fluids, single micro- and nano-particle spectroscopy, single-cell analysis, and statistical-physics experiments. This roadmap provides insights into current investigations involving optical forces and optical tweezers from their theoretical foundations to designs and setups. It also offers perspectives for applications to a wide range of research fields, from biophysics to space explorationEuropean Commission (Horizon 2020, Project No. 812780

Shape-induced force fields in optical trapping

Advances in optical tweezers, coupled with the proliferation of two-photon polymerization systems, mean that it is now becoming routine to fabricate and trap non-spherical particles. The shaping of both light beams and particles allows fine control over the flow of momentum from the optical to mechanical regimes. However, understanding and predicting the behaviour of such systems is highly complex in comparison with the traditional optically trapped microsphere. In this Article, we present a conceptually new and simple approach based on the nature of the optical force density. We illustrate the method through the design and fabrication of a shaped particle capable of acting as a passive force clamp, and we demonstrate its use as an optically trapped probe for imaging surface topography. Further applications of the design rules highlighted here may lead to new sensors for probing biomolecule mechanics, as well as to the development of optically actuated micromachines

Utilising high-bandwidth-sampled Brownian trajectories in optical tweezers: novel techniques and future directions.

In this thesis, I outline the development, implementation and evaluation of new sensor-side techniques in Optical tweezers that improve current capabilities in hydrodynamics, colloid-colloid studies and tweezers-based field scanning, by focussing on the high-fidelity Brownian tracking possible with photodiode sensing. Utilising a Field Programmable Gate Array (FPGA) to perform ultra-fast sensor interrogation, pre-processing and trap control, I am able to achieve multi-colloidal confinement with arrangement capacity, spatial and temporal measurement exceeding present standards, while also enabling and exploring new post-processing methods that stand to substantially improve the performance of Optically Trapped Probe Microscopy (OTPM). This thesis begins with a review of the present state-of-the-art in optical tweezers-based tracking and control, and an outline of current and emerging statistical analyses of Brownian motion. It continues with an investigation into the effects of trap-beam aberrations on measurements of Brownian motion, necessary to show that individual positional measurements in an imperfect system still possess enough information/certainty to be used in future chapters. Following this, a rapid trap-multiplexing method of multi-trap control is introduced, characterised and evaluated, whereby the FPGA is utilised as a genericised trap positioning and sensing platform, allowing arbitrary, fast-moving configurations of optical tweezers and simultaneous high-fidelity measurement. Finally, the utility of this high-bandwidth sensing is further explored with the development of a novel technique to correct for Brownian-based broadening in optical tweezer-based scanning images. The correction technique is demonstrated experimentally and using a Brownian dynamics simulation, providing both proof of concept results and promising directions for further development

Accurate position tracking of optically trapped live cells

Optical trapping is a powerful tool in Life Science research and is becoming common place in many microscopy laboratories and facilities. There is a growing need to directly trap the cells of interest rather than introduce beads to the sample that can affect the fundamental biological functions of the sample and impact on the very properties the user wishes to observe and measure. However, instabilities while tracking large inhomogeneous objects, such as cells, can make tracking position, calibrating trap strength and making reliable measurements challenging. These instabilities often manifest themselves as cell roll or re-orientation and can occur as a result of viscous drag forces and thermal convection, as well as spontaneously due to Brownian forces. In this paper we discuss and mathematically model the cause of this roll and present several experimental approaches for tackling these issues, including using a novel beam profile consisting of three closely spaced traps and tracking a trapped object by analysing fluorescence images. The approaches presented here trap T cells which form part of the adaptive immune response system, but in principle can be applied to a wide range of samples where the size and inhomogeneous nature of the trapped object can hinder particle tracking experiments