Fluorescence Spectroscopy : New Methods and Applications (176 Figures)

Fluorescence spectroscopy has found widespread application in many fields of science and medicine - including physics, chemistry, biology, biochemistry, clinical medicine, cytology, immunology, hydrogeology, and the environmental sciences. This text is divided into chapters on new methods in the field, applications, fluorescence immunoassays, fluorometric analysis and fluorescence spectroscopy in the biomedical sciences. It presents recent applications of and new developments in fluorescence spectroscopy written by experts in the field. The contents cover fluorescence spectroscopy using synchrotron radiation; picosecond fluorescence spectroscopy, fluorescence microscopy; fluorescence scattering by synthetic polymers; fluorescence immunoassays; fluorescence for environmental monitoring; fluorescence in flow injection analysis; hydrogeological studies; fluorescence of proteins, lipids and membranes; cell fluorescence; calcium transients.

1. New Methods in Fluorescence Spectroscopy.- 1. Fluorescence Spectroscopy: Where We Are and Where We’re Going.- 1 Introduction.- 2 The Marriage of Luminescence Spectroscopy With Separation Methods.- 3 Improvement of Selectivity Based Upon Narrow-Band Excitation.- 3.1 Fluorescence Line Narrowing (FLN) Spectroscopy….- 3.2 Shpol’skii Spectroscopy.- 3.3 Matrix Isolation Fluorescence Spectroscopy.- 3.4 Supersonic Jet Spectroscopy.- 4 Fiber Optic Sensors - Non-Destructive Spectroscopy….- 5 Semiconductor Lasers.- 6 References.- 2. Interactions and Kinetics of Single Molecules as Observed by Fluorescence Correlation Spectroscopy.- 1 Introduction.- 2 The Experimental Set up.- 3 Theoretical Background.- 4 Results.- 4.1 Translational Diffusion.- 4.2 Rotational Diffusion.- 4.3 Molecular Interactions and Kinetics.- 4.4 Single Molecule Events.- 5 Conclusions.- 6 References.- 3. Fast Optical Imaging Techniques.- 1 Introduction.- 2 High Speed Imaging Technologies.- 2.1 Resolution.- 2.2 Sensitivity.- 2.3 Dynamic Range.- 3 Imaging System Components.- 4 Examples of Detector Systems.- 4.1 Standard Silicon Photodiodes.- 4.2 Vidicon TV Cameras.- 4.3 Charge-Coupled Devices (CCD).- 5 Conclusions.- 4. Kinetic Studies on Fluorescence Probes Using Synchrotron Radiation.- 1 Introduction: Adiabatic Photochemical Product Formation and Fluorescence Probes.- 2 Fluorescence Kinetics and Its Application to TICT States.- 2.1 The Principle of TICT States.- 2.2 The Time Structure of Synchrotron Radiation.- 3 How to Construct TICT Fluorescence Probes.- 3.1 Principal Approaches.- 4 TICT Probes in Biology.- 5 Application of TICT and Other Relaxing Probes in Liquid and Rigid Media.- 6 Proton Transfer Fluorescence Probes.- 7 References.- 5. Dynamics and Geometry in Dimeric Flavoproteins from Fluorescence Relaxation Spectroscopy.- 1 Introduction.- 2 Lipoamide Dehydrogenase and Glutathione Reductase as Biophysical Systems.- 3 Fluorescence Methodology.- 3.1 Molecular Relaxation Spectroscopy.- 3.2 Fluorescence Lifetime Distributions.- 4 Examples.- 5 Conclusions.- 6 References.- 6. Fluorescence Spectroscopy of Light Scattering Materials.- 1 Introduction.- 2 Spatial Intensity Distribution of Fluorescence and Determination of Fluorescence Yields.- 3 Sensitivity and Concentration Dependence of Fluorimetric Analysis.- 4 Chemical Interactions Between Fluorophore and Substrate.- 5 Problems of Quantitative Fluorescence Spectroscopy.- 5.1 Reabsorption and Reemission.- 5.2 Inhomogeneous Distribution of the Fluorophores….- 5.3 Reflectance Spectroscopy of Fluorescent Samples….- 6 References.- 7. Optical Sensors Based on Fluorescence Quenching.- 1 Introduction.- 2 Some Principles of Fluorescence Quenching.- 3 Oxygen Sensors.- 4 Halothane Sensor.- 5 Sulphur Dioxide Sensor.- 6 Halide Sensors.- 7 Humidity Sensor.- 8 Oxygen Sensors as Transducers.- 8.1 Hydrogen Peroxide Sensors.- 8.2 Glucose Sensors.- 8.3 Other Biosensors.- 9 Applications.- 10 Future Aspects.- 11 References.- 2. New Applications of Fluorometry.- 8. Fluorescence in Forest Decline Studies.- 1 Introduction.- 2 Materials and Methods.- 2.1 Plant Materials.- 2.2 Measuring Devices.- 3 Results and Discussion.- 3.1 Chlorophyll Content.- 3.2 Fluorescence Spectra.- 3.3 Picosecond Kinetics.- 3.4 Long Term Delayed Luminescence (LDL).- 3.5 Induction Kinetics of Delayed Luminescence.- 4 Resume.- 5 References.- 9. Fluorescence Microscopy Studies of Structure Formation in Surfactant Monolayers.- 1 Introduction.- 2 Principles of the Microfiuorescence Technique.- 3 Investigations on the Plane Water Surface.- 3.1 Domain Superlattice.- 3.2 Domain Shape and Size.- 3.3 Number of Domains.- 4 Investigations During the Hydrophilic Langmuir-Blodgett Transfer.- 4.1 Substrate-Mediated Condensation with a Stationary Substrate.- 4.2 Morphological Instabilities of the LC Growth Front.- 4.3 Substrate-Mediated Condensation and Local Dye Distribution During Continuous LB-Transfer.- 5 Conclusions.- 6 References.- 10. Fluorescence Lifetime Imaging and Application to Ca2+ Imaging.- Abbreviations.- 1 Introduction.- 2 Creation of Lifetime Images.- 3 Instrumentation for FLIM.- 4 Experimental Methods.- 5 Results.- 5.1 FLIM of Lifetime Standards.- 5.2 FLIM of Quin-2.- 5.3 Phase Suppression Imaging of Ca2+.- 6 Discussion.- 7 References.- 11. Ether Phospholipids in Membranes: Applications of Phase and Steady-State Fluorometry.- 12. Pyrene-Labelled Lipids as Fluorescent Probes in Studies on Biomembranes and Membrane Models.- 1 Introduction.- 2 Basic Photophysics of Pyrene and Its Lipid Derivatives….- 3 Studies on Lipid Dynamics.- 3.1 Phospholipid Phase Separation.- 3.2 Formation of Lipid Superlattices.- 3.3 Estimation of Liposome Equilibrium Lateral Surface Pressure.- 4 Peripheral Ligand-Membrane Interactions.- 4.1 Formation of Superlattices of Membrane-Bound Cytochrome c.- 4.2 Binding of Polysomes to Rough Endoplasmic Reticulum Membrane.- 5 Future Perspectives.- 6 References.- 13. Optical Detection of Intracellular Ion Concentrations.- 1 Introduction.- 2 Theory of Color Change.- 3 Calibration Curve.- 4 How to Bring the Indicator Dye into the Cell.- 5 How Fluorescence is Measured.- 6 Limitations.- 6.1 Temporal Resolution.- 6.2 Spatial Resolution.- 6.3 Temporal and Spatial Resolution.- 7 Conclusion.- 8 References.- 3. Fluorimetric Analysis.- 14. Analytical Applications of Very Near-IR Fluorimetry.- 1 Introduction.- 2 VNIR Fluorophores.- 2.1 XantheneDyes.- 2.2 Phenoxazine Dyes.- 2.3 Carbocyanine and Merocyanine Dyes.- 3 Instrumentation.- 4 Results and Applications.- 5 Conclusions.- 6 References.- 15. Fluorescence Detection in Flow Injection Analysis.- 1 Introduction.- 2 Reactions Involved.- 3 Types of Manifold.- 4 Continuous Separation Techniques Coupled to FI Systems.- 5 (Bio)Chemical Fluorimetric Sensors in FI Systems.- 6 Final Remarks.- 7 References.- 16. Fluorescence Spectroscopy in Environmental and Hydrological Sciences.- 1 Introduction.- 2 EEM Studies, Multi-Component Analysis, Spectral Signatures, and “Fingerprinting”.- 3 Non-EEM Fluorescence Methods.- 4 Determination of Humic Dissociation Constants.- 5 Physical Characteristics and Fluorophore Composition of Fulvic and Humic Acids.- 6 Hydrologie Transport of Phenol and p-Cresol in the South Platte River Colorado.- 7 Gas Chromatographic Measurements.- 8 Fluorescence Measurements and Results.- 9 Pattern Recognition.- 10 Semi-Quantitative Method for Calculaton of Known Materials in EEM Spectra.- 11 EEM Spectra of Humic Fractions.- 12 Groundwater Pollution Mechanisms.- 13 Conclusions.- 14 References.- 4. Fluorescence Immunoassay.- 17. Fluorescence Polarisation Immunoassay.- 1 Introduction.- 2 Fluorescence Polarisation.- 3 Fluorophores.- 4 Design and Screening of Tracers.- 5 Summary.- 6 References.- 18. Progress in Delayed Fluorescence Immunoassay.- Abbreviations.- 1 Time-Resolved Fluorometry.- 2 Long-Decay-Time Luminescent Probes.- 2.1 Chelate Labels.- 3 Time-Resolved Fluoroimmunoassays.- 3.1 Fluorescence Enhancement Based Assays.- 3.2 Chelate Saturation Techniques.- 3.3 Assays with Stable Fluorescent Chelate Labels.- 4 Double-, Triple- and Quadruple-Label Assays.- 5 Homogeneous Assays.- 6 DNA Hybridization Assays.- 7 Future Prospects.- 8 References.- 19. Chemiluminescence Detection in Immunochemical Techniques. Applications to Environmental Monitoring.- 1 Introduction.- 2 Chemiluminescent Labels in Immunoassays.- 2.1 Transient Labels.- 2.2 Enzyme Labels.- 2.3 Advantages of Enzyme Labels.- 2.4 Disadvantages of Enzyme Labels.- 2.5 Comparison of Enzyme Labels.- 3 Detection of Chemiluminescence.- 3.1 Luminometers.- 3.2 Other Detection Devices.- 4 Applications to Environmental Monitoring.- 4.1 Immunoassays.- 4.2 Biotransformation Enzymes.- 4.3 Luminous Bacteria.- 4.4 Hygiene Monitoring.- 5 Conclusions.- 6 References.- 5. Fluorescence in Biomedical Sciences.- 20. Fluorescence Transients in Neurobiology: Applications of Voltage Sensitive and Ion Indicator Dyes.- 1 Introduction.- 2 Voltage Sensitive Dyes.- 3 Calcium Indicator Dyes.- 4 Sodium Indicator Dyes.- 5 Ion Diffusion.- 6 Imaging Technology.- 7 Future Directions.- 8 References.- 21. Optical Monitoring of Postsynaptic Potential in the Early Embryonic Avian Brain Stem Using a Voltage-Sensitive Dye.- 1 Introduction.- 2 Optical Methods for Monitoring Cellular Electrical Activity..- 2.1 Voltage-Sensitive Dyes.- 2.2 Optical Recording System.- 3 Embryonic Chick Brain Stem Preparations.- 4 Optical Detection of Evoked Action Potentials.- 5 Postsynaptic Potential Signals.- 5.1 Synaptic Fatigue.- 5.2 Effects of External Ca2+.- 6 Evidence for Glutaminergic EPSPs.- 7 Discussion.- 8 References.