AbstractThere are a growing variety of research fields requiring non-contact micro- manipulation. An increasing number of these fields are turning to optical tweezers as a solution, owing to their high spatial and temporal resolution. Optical tweezers have the ability to quantitively exert and measure forces on the piconewton scale, a convenient force scale for soft biological materials, and are hugely versatile due to the wide assortment of beam shaping techniques that can be employed.
The work in this thesis can be broadly divided into two main themes: that quantifying the optical trapping forces in shaped beams; and bringing control and simplification of complex systems to non-expert users who may utilise optical tweezers as part of interdisciplinary collaborations.
Static beam shaping is used to generate a conically refracted optical trap and the trapping properties are characterised. It is shown that trapping in the lower Raman spot gives full, 3D gradient trapping, while the upper Raman spot allows for particle guiding due to its levitation properties. Particles in the Lloyd/Poggendorff rings experience a lower trap stiffness than particles in the lower Raman spot but benefit from rotational control.
Dynamic beam shaping techniques are exploited for the simplification of complex systems through the development and testing of the HoloHands program. This software allows a holographic optical tweezers experiment to be controlled by gestures that are detected by a Microsoft Kinect. Multiple particle manipulation is demonstrated, as well as a calibration of the tweezers system.
Application of trapping forces is demonstrated through an examination of integrin – ligand bond strength. Both wild type effector T cells and those with a kindlin-3 binding site mutation similar to that found in neutrophils from Leukocyte Adhesion Deficiency sufferers are investigated. Through the use of back focal plane interferometry, a bond rupture force of (17.9 ± 0.6) pN at a force loading rate of (30 ± 4) pN/s, was measured for single integrins expressed on wild type cells. As expected, a significant drop in rupture force of bonds was found for mutated cells, with a measured rupture force of (10.1 ± 0.9) pN at the same pulling rate. Therefore, kindlin-3 binding to the cytoplasmic tail of the β2-tail directly affects bond strength of single integrin-ligand bonds. An experimental system for studying these cells under more physiologically relevant conditions is also presented.
Additionally, a low-cost optical micromanipulation system that makes use of simple microfabricated components coupled to a smartphone camera for imaging is proposed and demonstrated. Through the layering of hanging droplets of polydimethylsiloxane (PDMS) on microscope coverslips, lenses capable of optical trapping are created. Combination of PDMS with Sudan II dye led to the fabrication of long pass filters. An extension of this low-cost system into the life sciences is proposed through the adaptive use of bubble wrap, which allows for the culturing of cells in a chamber compatible with optical trapping.
|Date of Award||2017|
|Supervisor||David McGloin (Supervisor) & Susanna Fagerholm (Supervisor)|
- Optical tweezers
- Optical trapping
- Beam shaping