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Contents :
1. Two-Photon Excitation and Anisotropy Decays in Membranes and Oriented
Systems
B. Wieb Van der Meer and S.-Y. Simon Chen
1.1 Introduction
1.2 Two-Photon Anisotropy Theory in Isotropic Suspensions
1.3 Two-Photon Anisotropy Theory in Oriented Membranes
1.4 Conclusions
1.5 Appendix I
1.5.1 Appendix 1.1
1.5.2 Appendix 1.2
1.6 Appendix 2
1.6.1 Appendix 2.1
1.6.2 Appendix 2.2
1.6.3 Appendix 2.3
1.7 Appendix 3
1.8 Appendix 4
1.9 Appendix 5
1.10 Appendix 6
1.11 Appendix 7
1.12 Appendix 8
1.13 References
2. Ultrafast Stimulated Emission Spectroscopy
Gary J. Blanchard
2.1 Introduction
2.1.1 Stimulated Emission Pumping Spectroscopies
2.2 Experimental Implementation of Ultrafast Stimulated Emission
Pump-Probe Spectroscopy
2.3 Applications of Stimulated Emission Spectroscopy to Rotational
Diffusion Dynamics Measurements
2.4 Measuring Vibrational Population Relaxation Using Stimulated
Emission
2.4.1 Chemical Information Content of Ultrafast Stimulated
Emission T1Measurements: Perylene and l-Methylperylene
in n-alkanes
2.4.2 Solvent Dependent T1 Relaxation Dynamics of S0 Perylene
2.4.3 Understanding the Length Scales Over Which Intermolecular
Vibrational Energy Transfer Operates.
2.4.4 1-Methylperylene Vibrational Population Relaxation
2.4.5 Reorientation Dynamics of 1-Methylperylene in the Alkanes
2.4.6 Comparison of T1 and tauOR Data for 1-Methylperylene
2.5 Summary and Overview
2.6 References
3. Fluorescence and Multiwave Mixing Induced by Photon Absorption of
Excited Molecules
Valery Bogdanov
3.1 Introduction
3.2 Emission of Highly Excited Molecules
3.2.1 Methods of Excitation
3.2.2 Radiationless Decay of Highly Excited States
3.2.3 Emission Spectra and Relaxations of Short-Lived Highly
Excited States
3.2.4 Quantum Yield of Fluorescence From Highly Excited States
3.2.5 Anisotropy of Fluorescence from Highly Excited States
3.2.6 Vibrational Relaxation and Fluorescence Spectra of Highly
Excited Molecules
3.2.7 Energy Transfer from Highly Excited States
3.2.8 Stepped Excitation of Biflurophore
3.3 Transient Third-Order Optical Nonlinearity and Relaxation of
Excited Molecules
3.3.1 Preliminary Results
3.3.2 Dispersion of Transient Susceptibility and Ultrafast
Relaxation of Highly Excited Molecules
3.3.3 Transient Nonlinear Susceptibility and Orientational
Anisotropy of Excited Molecules
3.3.4 Time-resolved Dispersion of Transient Susceptibility
and Spectral Dynamics of Excited Molecules
3.4 Summary
3.5 References
4. The Theory of Two-Photon Induced Fluorescence Anisotropy
Patrik R. Callis
4.1 Introduction
4.2 Two-Photon Absorption
4.2.1 Qualitative Aspects
4.2.2 A Brief History
4.2.2.1 Use of Symmetry
4.2.2.2 Beyond the Use of Symmetry: Electronic Effects
4.2.2.3 Vibrationally-Induced Two-Photon Absorption
4.2.3 Quantitative Description
4.2.3.1 General
4.2.3.2 Involvement of Direct Transition Dipole
4.2.3.3 Pseudo Involvement of Direct Transition Dipole
4.2.3.4 Aromatic 1Lb Transitions
4.3 Two-Photon Anisotropy Theory
4.3.1 Introduction and Scope
4.3.2 Excitation with Linearly Polarized Light
4.3.3 Excitation with Circularly Polarized Light
4.3.3.1 Other ratios
4.3.4 The Two-Dimensional (2D) Limit with Linear Polarization
4.3.4.1 Determination of QX from r and Omega
4.3.4.2 Allowed Spaced of QX
4.3.4.3 Tensor Shapes
4.3.4.4 Tensor-Transition Moment Angle
4.3.4.5 Some Special 2D Cases
4.3.5 Three Dimensional Tensors
4.3.5.1 The [+,+] Sector
4.3.5.2 The [-,-] Sector
4.3.5.3 The [+,-] Sector
4.3.5.4 The [-,+] Sector
4.3.5.5 Exclusive Domains
4.4 Comparison with Experiment
4.4.1 Introduction
4.4.2 Polyenes
4.4.3 Benzene Derivatives
4.4.3.1 Benzene: The Pure Vibrational Baseline
4.4.3.2 Phenol
4.4.3.3 Tyrosine
4.4.3.4 RNAse A Behavior
4.4.4 Indoles
4.4.4.1 1La Excitation
4.4.4.2 1Lb Excitation
4.4.4.3 Out-of-Plane Contributions
4.4.4.4 INDO/S Predictions
4.5 Some Predictions
4.5.1 Phenylalanine
4.5.2 Tyrosine
4.5.2.1 S1 Maximum
4.5.2.2 S2
4.5.3 Indole in Non-Polar Solvent
4.5.4 5-Methoxyindole
4.5.5 Nucleotides
4.5.5.1 TMP
4.5.5.2 CMP
4.5.5.3 GMP
4.5.5.4 AMP
4.6 Concluding Remarks
4.7 References
5. Time-Resolved Stimulated-Emission and Transient-Absorption
Microscopy and Spectroscopy
P.T.C. So, C.Y. Dong, K.M. Berland, T. French and E. Gratton
5.1 Introduction
5.1.1 Pump-Probe Spectroscopy
5.1.2 Time-Resolved Fluorescence Microscopy
5.1.3 Overview
5.2 Theory
5.2.1 Pump-Probe Spectroscopy
5.2.2 Pump-Probe Microscopy
5.2.2.1 Radial and Axial Resolution
5.2.2.2 Confocal Pump-Probe Fluorescence Microscopy
5.2.2.3 Optical Transfer Function
5.2.2.4 Axial Depth Discrimination
5.3 Instrumentation and Methods
5.3.1 Pump-Probe Light Sources
5.3.2 Optics for Absorption Mode Measurement
5.3.3 Optics for Stimulated Emission Mode
5.3.4 Scanner Optics for Microscopy
5.3.5 Signal Detection and Processing
5.4 Spectroscopy Applications and Performance
5.4.1 Pump-Probe Absorption Spectroscopy
5.4.2 Stimulated Emission Pump-Probe Spectroscopy
5.4.2.1 One-Photon Stimulated Emission Spectroscopy
5.4.2.2 Two-Photon Stimulated Emission Spectroscopy
5.5 Applications and Performances of Pump-Probe Fluorescence
Microscope
5.5.1 Linearity of Pump-Probe Signal Response on Laser
Power Level
5.5.2 Experimental Verification of Pump-Probe Microscopy
Resolution
5.5.3 Time-Resolution of Pump-Probe Microscopy
5.5.4 Comparison Between Conventional Microscopy and
Pump-Probe Fluorescence Microscopy in Human
Erythrocytes and Mouse Fibroblasts
5.5.5 Multiple Dye Labeling Application
5.6 Conclusions
5.7 References
6. Anisotropy Decays Induced by Two-Photon Excitation
Carey K. Johnson and Chaozhi Wan
6.1 Introduction
6.1.1 Introduction and Overview
6.1.2 Theoretical Background
6.1.3 Time-Resolved Two-Photon Excitation
6.2 Orientational Distributions Induced by Two-Photon Excitation
6.2.1 Orientational Averages and Spherical Tensors
6.2.2 Spherical Tensors - A Primer
6.2.3 Two-Photon Photoselection
6.2.4 Two-Photon Anisotropies
6.2.5 Two-Photon Anisotropic Distributions
6.3 Two-Photon Anisotropy Decay
6.3.1 Rotational Diffusion
6.3.2 Two-Photon Induced Reorientational Decay
6.3.3 Time Dependence of the Two-Photon Anisotropies
6.3.4 Example: Perylene
6.4 Conclusion
6.5 References
7. Multi-Photon Excitation of Biochemical Fluorophores
Joseph R. Lakowicz and Ignacy Gryczynski
7.1 Introduction
7.2 Two-Photon Excitation of DPH
7.2.1 Emission Spectra and DPH Intensity Decays
for 1PE and 2PE
7.2.2 Anisotropy Spectra with Two-Photon Excitation
7.3 Theory of Photoselection with Multi-Photon Excitation
7.3.1 Anisotropy Decays of DPH with Two-Photon Excitation
7.4 Two-Photon Excitation of DPH in Membranes
7.5 Two-Photon Excitation of Fluorophore-Stained DNA
7.5.1 Emission Spectra of Hoechst 33342
7.5.2 Fluorescence Intensity Decays of Hoechst 33342
7.5.3 Anisotropy Excitation Spectra of Hoechst 33342
in Glycerol at - 20 C
7.5.4 Anisotropy Excitation Spectra of Hoechst 33342-DNA
Complex
7.6 Two-Photon Excitation of Indole, Tryptophan and Proteins
7.6.1 Emission Spectra
7.6.2 Anisotropy Spectra
7.6.3 Two-Photon Excitation of HSA
7.7 Two-Photon Induced Fluorescence of Alkanes
7.8 Standards for Two-Photon Excitation
7.8.1 A ps Two-Photon Standard
7.9 Three-Photon Excitation of Fluorescence
7.9.1 Three-Photon Excitation of 2,5-Diphenyloxazole (PPO)
7.9.2 Three-Photon Excitation of DPH in Membranes
7.9.3 Three-Photon Excitation of the Calcium Probe Indo-1
7.9.4 Three-Photon Excitation of Tryptophan and Proteins
7.9.5 Perspectives on Three-Photon Excitation
7.10 Two-Color Two-Photon Excitation
7.10.1 Perspectives on Two-Color Two-Photon Excitation
7.11 Future Applications of Multi-Photon excitation
7.12 References
8. Fluorescence Quenching by Stimulated Emission
Joseph R. Lakowicz and Ignacy Gryczynski
8.1 Introduction
8.2 One-Pulse and Two-Pulse Quenching
8.2.1 One-Pulse Time-Coincident Light Quenching
8.2.2 Two-Pulse Delayed Light Quenching
8.3 Effects of Light Quenching on Time-Resolved Intensity
and Anisotropy Decays
8.3.1 Lifetime-Resolved Experiments by Two-Beam Light
Quenching
8.4 Experimental Results One-Pulse Light Quenching
8.4.1 One-Pulse Light Quenching of DCM
8.4.2 One-Pulse Light Quenching with Two-Photon Excitation
8.4.3 Two-Pulse Light Quenching
8.4.4 Steady-State Measurements with Two-Beam Light Quenching
8.4.5 Time-Domain and Frequency-Domain Studies of Light
Quenching
8.4.6 Wavelength-Selective Light-Quenching with Time-Dependent
Spectral Relaxation
8.5 Applications of Light Quenching
8.5.1 Wavelength or Lifetime-Selective Light Quenching
8.5.2 Light Quenching with Total Internal Reflectance
8.6 Discussion
8.6.1 Comparison of Light Quenching and Amplified Stimulated
Emission
8.6.2 Biophysical Application of Light Quenching
8.6.3 Pulse-Width Dependent Light Quenching
8.6.4 Perspective on Light Quenching
8.7 References
9. Two-Photon Induced Fluorescence of Proteins
Borys Kierdaszuk, Ignacy Gryczynski and Joseph R. Lakowicz
9.1 Introduction
9.2 Excitation and Emission Spectra of Aromatic Aminoacids for One-
and Two-Photon Excitation
9.2.1 Studies of Tryptophan, Tyrosine and Phenylalanine in Solution
9.2.2 Tryptophan and Tyrosine in Proteins
9.3 Fluorescence Anisotropy of One- and Two-Photon Excitation of
Tryptophan and Tyrosine
9.3.1 Limiting Anisotropy Spectra
9.3.2 Anisotropy of Tryptophan and/or Tyrosine Containing Proteins
9.4 Two-Photon Induced Fluorescence of Protein Bound Chromophores
9.5 Experimental Methods
9.6 References
10. Increasing the Resolution of Far-Field Fluorescence Light ~Tfflcroscopy
by Point Spread Functional Engineering
Stefan W. Hell
10.1 Introduction
10.1.1 Why Far-Field Light Microscopy?
10.1.2 Imaging with a Lens: The Point Spread Function (PSF)
10.1.3 Scanning Fluorescence Microscopy
10.2 PSF-Engineering by Fluorescence Inhibition: STED and GSD
Microscopy
10.2.1 STED-Fluorescence Microscopy
10.2.1.1 Depletion of the Excited Singlet State by
Stimulated Emission
10.2.1.2 Resolution in the Concept of STED Fluorescence
Microscopy
10.2.1.3 STED Fluorescence Microscopy with Incomplete
Depletion
10.2.1.4 Toward the Practical Realization of STED
Fluorescence Microscopy: Studies on Depletion by
Stimulated Emission
10.2.2 Ground State Depletion (GDS) Fluorescence Microscopy
10.3 PSF Engineering Through Aperture Increase: 4Pi-Microscopy
10.3.1 4Pi-Illumination and Detection PSF
10.3.2 4Pi-Confocal Microscopy and Its Imaging Modes
10.3.3 Two-Photon Excitation 4Pi Microscopy
10.3.4 Confocal Two-Photon Excitation of 4Pi-Microscopy
10.3.5 Limitations and Potentials of 4Pi-Confocal Microscopy
10.4 Other Examples of PSF Engineering
10.4.1 Offset-beam Overlap Microscopy
10.4.2 Theta-(4Pi) Confocal Microscopy
10.5 Recent Developments
10.6 Conclusion
10.7 References
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