Optical tweezers and scissors are two common applications of laser micro-beams in biological research. Previously in our lab, there have been experiments that utilized both optical trapping and cutting, but in these experiments, optical trapping and cutting were performed at different time-points as the sample would have to be transferred from an optical cutting system to an optical trapping system or vice versa. In this project, we designed and built a combined laser scissors and tweezers microscope that (1) has two trapping beams and (2) uses a short pulsed tunable 200 fs 76 MHz 710-990 nm Ti:Sapphire laser for laser microsurgery. Both the position and power of all three beams can be independently controlled. Moreover, experiments were performed to test the ability of the system to trap and move whole chromosome or chromosome fragments inside living PTK-2 cells as well as chromosomes in suspension. The optical scissors laser was used at 730 nm to successfully cut chromosomes inside live cells. The optical traps were able to move whole chromosomes or chromosome fragments (after cutting) inside live Nocodazole-treated cells and also chromosomes isolated in suspension. The optical traps were not able to move intact or cut chromosomes inside un-treated cells, cold-treated cells or cells where the kinetochore or microtubule spindle was damaged using the optical scissors. Our experiments show that chromosome's geometry and index of refraction allow for easy trapping. However, when inside cells, the microenvironment around them, especially the microtubule spindle cage, can prevent any trapping movement.

Volume 55 in Methods in Cell Biology is a concise laboratory book that emphasizes the methods and technologies needed to use single polarized laser light source that functions simultaneously as an optical trap and a dual-beam interferometer.* * Provides a practical laboratory guide for methods and technologies used with laser tweezers* Includes comprehensive and easy-to-follow protocols

This detailed volume explores a wide variety of techniques involving optical tweezers, a technology that has become increasingly more accessible to a broad range of researchers. Beginning with recent technical advances, the book continues by covering the application of optical tweezers to study DNA-protein interactions and DNA motors, protocols to perform protein (un)folding experiments, the application of optical tweezers to study actin- and microtubule-associated motor proteins, and well as protocols for investigating the function and mechanical properties of microtubules and intermediate filaments, and more. Written for the highly successful Methods in Molecular Biology series, 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 practical, Optical Tweezers: Methods and Protocols, Second Edition serves as an ideal resource for expanding the accessibility and use of optical traps by scientists of diverse disciplines.

Optical tweezers and laser scissors are invaluable tools to manipulate cells on the micro and nano scale level. By integrating laser technologies with imaging and biochemical techniques, robust systems can be created to study various biological and biochemical processes. The purpose of this thesis is to describe two different optical methods used to study two cell-based problems. In the first experiment, optical tweezers are used to examine the effects of viscosity on sperm motility and energetics using a 1064nm Nd:YVO4 continuous laser. In the second experiment, laser scissors are used to study growth cone response from laser induced damage using a Vanguard Nd: YVO4 second harmonic generator (SHG) 532 nm picosecond green laser. The proceeding sections describe the optical systems for each experiment as well as the methods used. Results of both studies are presented and discussed in detail.

In many situations there is a need to physically manipulate microscopic objects with as much dexterity as our own hands provide in the macroscopic world. An example would be applying stresses onto cells in order to determine their mechanical properties. Our existing microscope-laser experimental system is capable of manipulating microscopic objects using two optical tweezers and one optical scissors to move and cut these objects, respectively. Despite these capabilities, however, the point-and-click user interface for optical trapping and cutting was cumbersome and hard to use, limiting the system's potential for micro-manipulation. In order to resolve this limitation, a new, more intuitive and hands-on user interface using two joysticks was designed and developed from the ground up in order to provide responsive, real-time control of the optical tweezers and scissors for microscopic manipulation. This new joystick user interface was then used to verify whether or not forces other than those due to microtubule dynamics act upon chromosomes during mitosis, tested in mitotic PtK2 and Indian Muntjac cells by 1) depolymerizing microtubules using nocodazole, and 2) disrupting microtubules using laser ablation with optical scissors, and subsequently attempting to freely manipulate chromosomes using joystick-controlled optical trapping.

There are probably few people who do not dream of the good old times, when do ing science often meant fascination, excitement, even adventure. In our time, do ing science involves often technology and, perhaps, even business. But there are still niches where curiosity and fascination have their place. The subject of this book, technological as its title may sound, is one of the fortunate examples. It will report on lasers generating the coldest places in the Universe, and on table top laser microtools which can produce a heat "inferno" as it prevails in the interior of the Sun, or simulate, for specific plant cells, microgravity of the space around our plan et Earth. There will be some real surprises for the reader. The applications range from basic studies of the driving forces of cell division (and thus life) via genetic modification of cells (for example, for plant breeding) to medical applications such as blood cell analysis and finally in vitro fertilization. What are these instruments: laser microbeams and optical tweezers? Both are lasers coupled with a fluorescence microscope. The laser microbeam uses a pulsed ultraviolet laser. Light is focused, as well as possible, in space and time, in order to obtain extremely high light intensities - high enough to generate, for a very short instant, extremely hot spots which can be used to cut, fuse or perforate biological material.