Microrheology With Optical Tweezers

Thanks to the pioneering works of Ashkin and coworkers, optical tweezers (OTs) have become an invaluable tool for myriad studies throughout the natural sciences. Their success relies on the fact that they can be considered as exceptionally sensitive transducers that are able to resolve pN forces and nm displacements, with high temporal resolution, down to μs. Hence their application to study a wide range of biological phenomena such as measuring the compliance of bacterial tails, the forces exerted by a single motor protein, and the mechanical properties of human red blood cells and of individual biological molecules. The number of articles related to them totals to a whopping 58,000 (source Google Scholar)! Microrheology is a branch of rheology, but it works at micrometer length scales and with microliter sample volumes. Therefore, microrheology techniques have been revealed to be very useful tools for all those rheological/mechanical studies where rare or precious materials are employed, such as in biological and biomedical studies. The aim of this book is to provide a pedagogical introduction to the physics principles governing both the optical tweezers and their application in the field of microrheology of complex materials. This is achieved by following a linear path that starts from a narrative introduction of the "nature of light," followed by a rigorous description of the fundamental equations governing the propagation of light through matter. Moreover, some of the many possible instrumental configurations are presented, especially those that better adapt to perform microrheology measurements. In order to better appreciate the microrheological methods with optical tweezers explored in this book, informative introductions to the basic concepts of linear rheology, statistical mechanics, and the most popular microrheology techniques are also given. Furthermore, an enlightening prologue to the general applications of optical tweezers different from rheological purposes is provided at the end of the book.

Thanks to the pioneering works of Ashkin and coworkers, optical tweezers (OTs) have become an invaluable tool for myriad studies throughout the natural sciences. Their success relies on the fact that they can be considered as exceptionally sensitive transducers that are able to resolve pN forces and nm displacements, with high temporal resolution, down to μs. Hence their application to study a wide range of biological phenomena such as measuring the compliance of bacterial tails, the forces exerted by a single motor protein, and the mechanical properties of human red blood cells and of individual biological molecules. The number of articles related to them totals to a whopping 58,000 (source Google Scholar)! Microrheology is a branch of rheology, but it works at micrometer length scales and with microliter sample volumes. Therefore, microrheology techniques have been revealed to be very useful tools for all those rheological/mechanical studies where rare or precious materials are employed, such as in biological and biomedical studies. The aim of this book is to provide a pedagogical introduction to the physics principles governing both the optical tweezers and their application in the field of microrheology of complex materials. This is achieved by following a linear path that starts from a narrative introduction of the "nature of light," followed by a rigorous description of the fundamental equations governing the propagation of light through matter. Moreover, some of the many possible instrumental configurations are presented, especially those that better adapt to perform microrheology measurements. In order to better appreciate the microrheological methods with optical tweezers explored in this book, informative introductions to the basic concepts of linear rheology, statistical mechanics, and the most popular microrheology techniques are also given. Furthermore, an enlightening prologue to the general applications of optical tweezers different from rheological purposes is provided at the end of the book.

This thesis presents a method to measure the linear viscoelastic response of fluids by tracking and analyzing the thermal, Brownian motion of suspended tracer particles, known as passive microrheology. The particle is confined in a harmonic optical trap and its one dimensional trajectory is obtained by a home-built split beam detection system, which works similar but responds faster than position detection with commercial quadrant photodiodes. The theory which is necessary to convert the particle trajectory into the complex shear modulus is derived in detail, pointing out that the commonly used Mason-Weitz method needs to be modified in order to obtain correct results at high frequencies due to hydrodynamic effects of the fluid. It follows a detailed explanation of the data analysis procedure which is verified for water up to angular frequencies of 107 rad/s in very good agreement with the theory. Finally, there is an outlook how to apply the method to actual complex fluids.

This book presents a comprehensive overview of microrheology, emphasizing the underlying theory, practical aspects of its implementation, and current applications to rheological studies in academic and industrial laboratories. The field of microrheology continues to evolve rapidly, and applications are expanding at an accelerating pace. Readers will learn about the key methods and techniques, including important considerations to be made with respect to the materials most amenable to microrheological characterization and pitfalls to avoid in measurements and analysis. Microrheological measurements can be as straightforward as video microscopy recordings of colloidal particle Brownian motion; these simple experiments can yield rich rheological information. Microrheology covers topics ranging from active microrheology using laser or magnetic tweezers to passive microrheology, such as multiple particle tracking and tracer particle microrheology with diffusing wave spectroscopy. Overall, this introduction to microrheology informs those seeking to incorporate these methods into their own research, or simply survey and understand the growing body of microrheology literature. Many sources of archival literature are consolidated into an accessible volume for rheologist and non-specialist alike. The small sample sizes of many microrheology experiments have made it an important method for studying emerging and scarce biological materials, making this characterization method suitable for application in a variety of fields.

Microscale Diagnostic Techniques highlights the most innovative and powerful developments in microscale diagnostics. It provides a resource for scientists and researchers interested in learning about the techniques themselves, including their capabilities and limitations. The fields of Micro- and Nanotechnology have emerged over the past decade as a major focus of modern scientific and engineering research and technology. Driven by advances in microfabrication, the investigation, manipulation and engineering of systems characterized by micrometer and, more recently, nanometer scales have become commonplace throughout all technical disciplines. With these developments, an entirely new collection of experimental techniques has been developed to explore and characterize such systems.

Collagen, the most abundant protein in the body, assembles into an extra-cellular fibrillar gel, which has both viscous and elastic properties. These properties can be determined by using optical tweezers to hold a micron-sized bead within the sample. Measurement of the bead's thermally induced motion enables the determination of the frequency-dependent viscoelasticity. Rather than only probing response at a single location, holographic optical tweezers create multiple, independent traps, permitting simultaneous tracking of multiple embedded beads and characterization of their correlated motion. By using this technique in a collagen gel, we will be able to determine local and cross-correlated viscoelastic properties, which vary at different locations during its formation. Implications of this research lie in the fields of health and biomaterials. The aim of this work is to devise and validate protocols for using holographic optical tweezers to measure local and through-space viscoelasticity. Rather than using laser deflection to track particle motion, I use a high-speed camera and image analysis to track the simultaneous motion of multiple beads. This approach provides nanometer-scale resolution of particle position at sampling rates up to 2.5 kHz. I compare tracking data collected from the high-speed camera to those collected by the laser deflection method and find a discrepancy in the perceived motion of the bead. I perform many experimental tests to assess the root of this problem. Additionally, I numerically represent bead motion measurements if collected using both methods (laser-deflection method and high-speed camera method) and compare them to the idealized measurement results. In doing so, I learn about the limitations of each method, and how the viscous and elastic properties inferred from the data are affected by each measurement device. Finally, based on my numerical representations, I suggest a simple procedure to gain more accuracy in the viscous and elastic properties for both simple fluids (such as water) and complex fluids (such as collagen solutions) when using each method. This procedure can be used in future holographic optical tweezers-based experiments to obtain an accurate representation of the local and correlated properties of collagen.

The aim of this volume is to provide a comprehensive overview of optical tweezers setups, both in practical and theoretical terms, to help biophysicists, biochemists, and cell biologists to build and calibrate their own instruments and to perform force measurements on mechanoenzymes both in isolation in vitro and in living cells. Chapters have been divided in three parts focusing on theory and practical design of optical tweezers, detailed protocols for performing force measurements on single DNA- and microtubule/actin-associated mechanoenzymes in isolation, and describing recent advances that have opened up quantitative force measurements in living cells. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and cutting-edge, Optical Tweezers: Methods and Protocols aims help to further expand the accessibility and use of optical traps by scientists of diverse disciplines. .