Micromanipulation by Light in Biology and Medicine The Laser Microbeam and Optical Tweezers

Laser micromanipulation--implying perforation, cutting and welding of biological objects using. optical tweezers, as well as moving them just by using, forces generated by a laser microbeam--is becoming an increasingly popular technique for studying biological objects, especially automatedDNA sequencing. This book is an excellent introduction to the basics and primarily biological applications of optical trapping and laser microbeam techniques. The author, one of the pioneers of the microbeam technique, examines the field in depth, covering basic physical principles to applications in cell and molecular biology, and biomedicine. This book bridges the gap between physicists who understand the underlying principles of the method but are not aware of its numerous biological applications, and biologists who seek background information on the underlying principles. Series: Methods in Bioengineering Contents: Series Preface Preface Introduction: The history of using light as a working tool 1. From photons to the laser microtools 1.1 Light 1.1.1 Light is a special form of electromagnetic radiation 1.1.2 The energy carried by light 1.1.3 Photons and their social behavior 1.1.4 The (relativistic) mass of a photon 1.1.5 Light pressure: Where does it come from 1.2 Lasers: sources of peculiar light 1.2.1 Laser light: Spectral purity, coherence and divergence 1.2.2 Principle of lasing 1.2.3 Inversion, three and four state lasers 1.2.4 Pumping: The primary energy source for lasers 1.2.5 Laser resonators and laser modes 1.2.6 Changing the color: Frequency multiplication 1.2.7 Real lasers 1.3 Microscopes and cameras 1.3.1 Different types of light microscopy 1.3.2 Object illumination 1.3.3 The magnification of a microscope 1.3.4 A two lens microscope 1.3.5 Real microscopes 1.3.6 Microscope objectives 1.3.7 Microscope detectors 1.4 Laser microbeams and optical tweezers 1.4.1 Choice of the laser 1.4.2 Choice of microscope and objective 1.4.3 Building a microbeam and/or optical tweezers 1.4.4 The coupling unit 1.4.5 Mounting the parts and adjusting the lasers 1.4.6 Flying spot - and multiple beam optical tweezers 1.4.7 Combination of laser microbeam and optical tweezers 2. Why and how can light be used as a microtool 2.1 Pulsed laser microbeams 2.1.1 Pulsed lasers can be focused to extreme intensities 2.1.2 Bringing the Sun into the lab 2.1.3 Why is the laser microbeam so precise: The heat is diluted immediately 2.1.4 Not only heat: the electric fields are also impressive 2.1.5 Caution: X rays 2.2 Interaction of pulsed lasers with biological matter 2.2.1 High intensities in medical surgery 2.2.2 Photons cooperate at extreme intensities 2.2.3 Heat shocks from a laser microbeam: Too fast to damage proteins 2.2.4 Working in the interior of closed objects 2.3 Interaction of optical tweezers with biological material 2.3.1 The wavelength makes the difference 2.3.2 Thermal effects in continuous laser microbeams 2.3.3 Interaction of Nd YAG optical tweezers (1064nm) with cells 2.3.4 The surprise: Nd YAG lasers are not the best choice 2.3.5 Two photon excited fluorescence to detect damage in IR optical traps 2.4 Light pressure: Some quantitative relationships 2.4.1 Calculating some surprising facts 2.4.2 Mie particles: The color of the target affects the light force 2.4.3 Light forces on very small (Rayleigh) particles 2.5 Gradient forces: Full control in three dimensions 2.5.1. Axial and transversal effects 2.5.2 Why are particles pushed towards the optical axis? 2.5.3 Why are particles pulled against the light pressure force 2.5.4 Quantitative influence of objective illumination and of refractive indices 2.6 Unprecedented accuracy and precision: Nanometers and piconewtons 2.6.1 The escape force method for calibrating optical tweezers 2.6.2 The stiffness method 2.6.3 Calibration by thermal motion: The corner frequency method 2.6.4 Calibration by thermal motion: The minimal information method 2.6.5 Split photodiodes and interferometers for manometer accuracies 3. From the First simple experiments to sophisticated applications of laser microbeams 3.1 Microbeams in developmental biology 3.1.1 Drosophila: From the embryos to the organism 3.1.2 Laser microbeams in the study of the development of the roundworm Caenorhabditis elegans 3.1.3 The nerve system of Caenorhabditis elegans 3.1.4 Other nerve systems 3.1.5 Plant root redevelopment: Clonal or positional determination of cell fate? 3.2 Intermezzo I: Cells and cellular organelles 3.2.1 Prokaryotic: Bacteria 3.2.2 Cell membranes 3.2.3 Eukaryotic cells 3.2.4 The nucleus of eukaryotic cells 3.2.5 The cytoskeleton and structure-stabilizing intracellular strands 3.2.6. Subcellular structures: mitochondria and chloroplasts 3.2.7 Cell types for work with laser microbeams and optical tweezers 3.3 Laser microbeam studies on unicellular organisms, cells and subcellular structures 3.3.1 Negative and positive chemotaxis induced by laser ablation 3.3.2 Laser treatment of flagella, cilia and pseudopodia in unicellular organisms 3.3.3 Nerve cells: Inhibition of external stimulation and regeneration of neurites 3.3.4 Ablation of melanophores in fish 3.3.5 Ablation of mitochondria to study heart function 3.3.6 Nucleoli 3.3.7 Actomyosin fibrils and cytoplasmic strands 3.3.8 Laser ablation of interphase chromatin in the nucleus 3.4 Optical tweezers enter the stage 3.4.1 Optical trapping of viruses and bacteria 3.4.2 The step toward infrared optical tweezers 3.4.3 Single-cell sorters 3.4.4 Brakes for biological motors: The compliance of bacterial flagella. 3.4.5 The slipper animalcule survives moving subcellular organelles and the nucleus 3.4.6 Neural growth cone membranes 4. Mitosis, molecular motors and muscles: What laser microtools can teach us 4.1 Intermezzo II: Reproduction and motion 4.1.1 The cell cycle 4.1.2 Details of mitosis 4.1.3 Details of the interphase 4.1.4 Microtubules 4.1.5 Intracellular traffic: cytoplasmic streaming 4.1.6 Molecular motors 4.1.7 Forces generated by single molecules 4.2 What drives cell division? Laser microtools help to find the answer 4.2.1 From the early days to complete micromanipulation by laser light 4.2.2 Kinetochores and centrosomes: The secret of symmetric distribution of chromosomes 4.2.3 Microablation studies on intranuclear metaphase chromosomes 4.2.4 Microtubule polymerization dynamics and flexibility 4.3 Molecular motors: True nanotechnology 4.3.1 Optical tweezers and single motor protein mechanics 4.3.2 Molecular force measurements in vivo: Mitochondria transport in the amoeba Reticulomyxa 4.3.3 Kinesin 4.3.4 Dynein - not as exact as kinesin 4.3.5 Myosinactin: single heads can do the work 4.3.6 Differences between skeletal and smooth muscle 4.3.7 Molecular bungee: Titin 4.3.8 Laser microbeams for preparing the smaller functional unit of muscle 5. DNA as a molecular individuum 5.1 The ultimate DNA analytics: single molecules 5.1.1 DNA: Astronomical dimensions and unlimited variability 5.1.2 Handling of DNA 5.1.3 Laser microdissection of single DNA molecules 5.1.4 The elasticity of a single DNA molecule 5.1.5 Two types of elasticity in DNA 5.2 Single molecule DNA reactions 5.2.1 From bulk properties to single molecule reactions 5.2.2 Cutting of an individual DNA molecule by the restriction endonucleases Apa 1 5.2.3 Transcription against a force 5.3 Intermezzo III: Genes, chromosomes and genetic diseases 5.3.1 Packing DNA: Chromatin 5.3.2 From chromatin to chromosomes 5.3-3 Banded chromosomes and cytogenetic nomenclature 5.3.4 Genome sizes of different organisms 5.3.5 Genetic diseases 5.4 Laser microdissection of chromosomes 5.4.1 The motivation for microdissection of chromosomes 5.4.2 Microdissection of chromosomes 5.4.3 Submicroscopic effects of laser microdissection 5.4.4 Combined use of laser microbeam and optical tweezers 5.4.5 Microcloning 5.4.6 PCR 6 From biology to medicine 6.1 Laser microtools in plant cell biology 6. 1.1 Optical trapping in the interior of plant cells 6.1.2 Simulating microgr