Optical Tweezers for Single Molecule Biology

Molecular machinery on the micro-scale, believed to be the fundamental building blocks of life, involve forces of 1-100 pN and movements of nanometers to micrometers. Micromechanical single-molecule experiments seek to understand the physics of nucleic acids, molecular motors, and other biological systems through direct measurement of forces and displacements. Optical tweezers are a popular choice among several complementary techniques for sensitive force-spectroscopy in the field of single molecule biology. The main objective of this thesis was to design and construct an optical tweezers instrument capable of investigating the physics of molecular motors and mechanisms of protein/nucleic-acid interactions on the single-molecule level. A double-trap optical tweezers instrument incorporating acousto-optic trap-steering, two independent detection channels, and a real-time digital controller was built. A numerical simulation and a theoretical study was performed to assess the signal-to-noise ratio in a constant-force molecular motor stepping experiment. Real-time feedback control of optical tweezers was explored in three studies. Position-clamping was implemented and compared to theoretical models using both proportional and predictive control. A force-clamp was implemented and tested with a DNA-tether in presence of the enzyme lambda exonuclease. The results of the study indicate that the presented models describing signal-to-noise ratio in constant-force experiments and feedback control experiments in optical tweezers agree well with experimental data. The effective trap stiffness can be increased by an order of magnitude using the presented position-clamping method. The force-clamp can be used for constant-force experiments, and the results from a proof-of-principle experiment, in which the enzyme lambda exonuclease converts double-stranded DNA to single-stranded DNA, agree with previous research. The main objective of the thesis was thus achieved. The developed instrument and presented results on feedback control serve as a stepping stone for future contributions to the growing field of single molecule biology.Molekylära biologiska mekanismer involverar 0-100 pN krafter och rörelser från under en nanometer till flera mikrometer. Mikromekaniska experiment strävar till att förklara hur dessa mekanismer/maskiner fungerar genom direkt mätning av rörelser och krafter. Bland flera komplementära enmolekyls-tekniker är den optiska pincetten populär pga. dess höga platsresolution och lämpliga styvhet. En optisk pincett använder laserljus för att ta tag i och mäta krafter/rörelser hos dielektriska partiklar. I enmolekyls experiment binds biomolekyler kemiskt till plastbollar som hålls i pincetten och fungerar som 'handtag' till mikro-världen. Huvudmålet med denna avhandling var att utveckla och bygga ett laserpincett instrument som möjliggör enmolekyls experiment som förklarar molekylära mekanismer samt biofysiken i protein/nukleinsyra-växelverkningar. Ett laserpincett instrument med två pincetter, den ena styrbar, position/kraft mätning i båda pincetterna med hög resolution, samt en realtids kontroller byggdes. Avhandlingen består av fyra artiklar och ett sammandrag. Den första artikeln analyserar signal/brusförhållandet (SNR) i ett enmolekyls experiment. I mikro-skala påverkas det biologiska system som undersöks samt plastbollarna av värmerörelse, vilket försämrar SNR. Artikeln visar vilka förhållanden (molekylens längd, styvhet, spänning, osv.) leder till hög SNR och experiment som kan observera enskilda steg hos molekylära maskiner (steglängden varierar från 0,34 nm för t.ex. DNA-polymeras, till 36 nm för t.ex. myosin). I den andra och tredje artikeln användes realtids-kontrollern för att fixera en plastboll i pincetten, värmerörelsen till trots. Pincetten styrdes 200 000 gånger per sekund på basen av bollens mätta position. Pincettens effektiva styvhet kunde ökas ca. 10-faldigt. I den fjärde artikeln användes kontrollern för att upprätthålla en konstant 3.4 pN spänning i en DNA-molekyl. Molekylen fördes till en lösning innehållande enzymet lambda exonukleas som konverterar dubbel-strängat DNA till en-strängat. Vid 3.4 pN spänning är en-strängat DNA signifikant kortare än dubbel-strängat, och molekylens krympande från ~14 μm till ~8 μm längd observerades under ca 20 min. Detta experiment visar att huvudmålet för avhandlingen uppnåddes. Det utvecklade instrumentet lämpar sig för enmolekylsexperiment och kan användas för framtida kontributioner inom enmolekyls biofysik

In situ sensors for measurements in the global trosposphere

Current techniques available for the in situ measurement of ambient trace gas species, particulate composition, and particulate size distribution are reviewed. The operational specifications of the various techniques are described. Most of the techniques described are those that have been used in airborne applications or show promise of being adaptable to airborne applications. Some of the instruments described are specialty items that are not commercially-available. In situ measurement techniques for several meteorological parameters important in the study of the distribution and transport of ambient air pollutants are discussed. Some remote measurement techniques for meteorological parameters are also discussed. State-of-the-art measurement capabilities are compared with a list of capabilities and specifications desired by NASA for ambient measurements in the global troposphere

Image-Guided Robot-Assisted Techniques with Applications in Minimally Invasive Therapy and Cell Biology

There are several situations where tasks can be performed better robotically rather than manually. Among these are situations (a) where high accuracy and robustness are required, (b) where difficult or hazardous working conditions exist, and (c) where very large or very small motions or forces are involved. Recent advances in technology have resulted in smaller size robots with higher accuracy and reliability. As a result, robotics is fi nding more and more applications in Biomedical Engineering. Medical Robotics and Cell Micro-Manipulation are two of these applications involving interaction with delicate living organs at very di fferent scales.Availability of a wide range of imaging modalities from ultrasound and X-ray fluoroscopy to high magni cation optical microscopes, makes it possible to use imaging as a powerful means to guide and control robot manipulators. This thesis includes three parts focusing on three applications of Image-Guided Robotics in biomedical engineering, including: Vascular Catheterization: a robotic system was developed to insert a catheter through the vasculature and guide it to a desired point via visual servoing. The system provides shared control with the operator to perform a task semi-automatically or through master-slave control. The system provides control of a catheter tip with high accuracy while reducing X-ray exposure to the clinicians and providing a more ergonomic situation for the cardiologists. Cardiac Catheterization: a master-slave robotic system was developed to perform accurate control of a steerable catheter to touch and ablate faulty regions on the inner walls of a beating heart in order to treat arrhythmia. The system facilitates touching and making contact with a target point in a beating heart chamber through master-slave control with coordinated visual feedback. Live Neuron Micro-Manipulation: a microscope image-guided robotic system was developed to provide shared control over multiple micro-manipulators to touch cell membranes in order to perform patch clamp electrophysiology. Image-guided robot-assisted techniques with master-slave control were implemented for each case to provide shared control between a human operator and a robot. The results show increased accuracy and reduced operation time in all three cases

A cryogenic liquid-mirror telescope on the moon to study the early universe

We have studied the feasibility and scientific potential of zenith observing liquid mirror telescopes having 20 to 100 m diameters located on the moon. They would carry out deep infrared surveys to study the distant universe and follow up discoveries made with the 6 m James Webb Space Telescope (JWST), with more detailed images and spectroscopic studies. They could detect objects 100 times fainter than JWST, observing the first, high-red shift stars in the early universe and their assembly into galaxies. We explored the scientific opportunities, key technologies and optimum location of such telescopes. We have demonstrated critical technologies. For example, the primary mirror would necessitate a high-reflectivity liquid that does not evaporate in the lunar vacuum and remains liquid at less than 100K: We have made a crucial demonstration by successfully coating an ionic liquid that has negligible vapor pressure. We also successfully experimented with a liquid mirror spinning on a superconducting bearing, as will be needed for the cryogenic, vacuum environment of the telescope. We have investigated issues related to lunar locations, concluding that locations within a few km of a pole are ideal for deep sky cover and long integration times. We have located ridges and crater rims within 0.5 degrees of the North Pole that are illuminated for at least some sun angles during lunar winter, providing power and temperature control. We also have identified potential problems, like lunar dust. Issues raised by our preliminary study demand additional in-depth analyses. These issues must be fully examined as part of a scientific debate we hope to start with the present article.Comment: 35 pages, 11 figures. To appear in Astrophysical Journal June 20 200

GAIA: Composition, Formation and Evolution of the Galaxy

The GAIA astrometric mission has recently been approved as one of the next two `cornerstones' of ESA's science programme, with a launch date target of not later than mid-2012. GAIA will provide positional and radial velocity measurements with the accuracies needed to produce a stereoscopic and kinematic census of about one billion stars throughout our Galaxy (and into the Local Group), amounting to about 1 per cent of the Galactic stellar population. GAIA's main scientific goal is to clarify the origin and history of our Galaxy, from a quantitative census of the stellar populations. It will advance questions such as when the stars in our Galaxy formed, when and how it was assembled, and its distribution of dark matter. The survey aims for completeness to V=20 mag, with accuracies of about 10 microarcsec at 15 mag. Combined with astrophysical information for each star, provided by on-board multi-colour photometry and (limited) spectroscopy, these data will have the precision necessary to quantify the early formation, and subsequent dynamical, chemical and star formation evolution of our Galaxy. Additional products include detection and orbital classification of tens of thousands of extra-Solar planetary systems, and a comprehensive survey of some 10^5-10^6 minor bodies in our Solar System, through galaxies in the nearby Universe, to some 500,000 distant quasars. It will provide a number of stringent new tests of general relativity and cosmology. The complete satellite system was evaluated as part of a detailed technology study, including a detailed payload design, corresponding accuracy assesments, and results from a prototype data reduction development.Comment: Accepted by A&A: 25 pages, 8 figure

A Step-by-step Guide to the Realisation of Advanced Optical Tweezers

Since the pioneering work of Arthur Ashkin, optical tweezers have become an indispensable tool for contactless manipulation of micro- and nanoparticles. Nowadays optical tweezers are employed in a myriad of applications demonstrating the importance of these tools. While the basic principle of optical tweezers is the use of a strongly focused laser beam to trap and manipulate particles, ever more complex experimental set-ups are required in order to perform novel and challenging experiments. With this article, we provide a detailed step- by-step guide for the construction of advanced optical manipulation systems. First, we explain how to build a single-beam optical tweezers on a home-made microscope and how to calibrate it. Improving on this design, we realize a holographic optical tweezers, which can manipulate independently multiple particles and generate more sophisticated wavefronts such as Laguerre-Gaussian beams. Finally, we explain how to implement a speckle optical tweezers, which permit one to employ random speckle light fields for deterministic optical manipulation.Comment: 29 pages, 7 figure

Applications of airborne remote sensing in atmospheric sciences research

This paper explores the potential for airborne remote sensing for atmospheric sciences research. Passive and active techniques from the microwave to visible bands are discussed. It is concluded that technology has progressed sufficiently in several areas that the time is right to develop and operate new remote sensing instruments for use by the community of atmospheric scientists as general purpose tools. Promising candidates include Doppler radar and lidar, infrared short range radiometry, and microwave radiometry

Optical vortices generated by a PANDA ring resonator for drug trapping and delivery applications

We propose a novel drug delivery system (DDS) by using a PANDA ring resonator to form, transmit and receive the microscopic volume by controlling some suitable ring parameters. The optical vortices (gradient optical field/well) can be generated and used to form the trapping tool in the same way as the optical tweezers. The microscopic volume (drug) can be trapped and moved (transported) dynamically within the wavelength router or network. In principle, the trapping force is formed by the combination between the gradient field and scattering photons, which has been reviewed. The advantage of the proposed system is that a transmitter and receiver can be formed within the same system, which is called transceiver, in which the use of such a system for microscopic volume (drug volume) trapping and transportation (delivery) can be realized