Introduction to Fluorescence: 1st Edition (Hardback) book cover

Introduction to Fluorescence

1st Edition

By David M. Jameson

CRC Press

313 pages | 265 Color Illus.

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The phenomenon known as fluorescence is now widely used in the chemical and life sciences largely due to the development of highly sophisticated fluorescent probe chemistries and the commercial availability of these probes as well as the development of novel microscopy approaches. Introduction to Fluorescence helps readers acquire a sound understanding of basic fluorescence theory and practice. It describes general principles in a straightforward way and uses examples from a variety of disciplines to demonstrate them.

In color throughout, the book takes readers through the history of important discoveries to the most current advances. It introduces the fundamentals of the fluorescence phenomenon and gives detailed examples of fluorescence applications in the molecular life sciences, including biochemistry, biophysics, clinical chemistry and diagnostics, pharmaceutical science, and cell and molecular biology. The author presents the basic theories underlying the applications and offers in-depth information on practical aspects.

Along with a list of references in each chapter, the text incorporates more than 250 figures that clearly illustrate the concepts and gives the chemical structures of the most widely used fluorescent molecules. In addition, the appendix provides a "Rogue’s Gallery" of the most common errors and pitfalls to avoid.


"An essential contribution to educating scientists in the principles of fluorescence. It will also be an important addition to the libraries of practitioners applying the principles of molecular fluorescence."

—Ken Jacobson, Kenan Distinguished Professor of Cell Biology and Physiology, University of North Carolina at Chapel Hill

"An exquisite compendium of fluorescence and its applications in biochemistry enriched by a very exciting historical perspective. This book will become a standard text for graduate students and other scientists."

—Drs. Zygmunt (Karol) Gryczynski and Ignacy Gryczynski, University of North Texas Health Science Center

"… truly a masterwork, combining clarity, precision, and good humor. The reader, novice or expert, will be pleased with the text and will not stop reading. It is a formidable account of the fluorescence field, which has impacted the life sciences so considerably in the last 60 years."

—Jerson L. Silva, M.D., Ph.D., Professor and Director, National Institute of Science and Technology for Structural Biology and Bioimaging, Universidade Federal do Rio de Janeiro

"Dr. Jameson’s book fills a gap in the literature available for the student approaching the applications of fluorescence techniques as well as for the researcher practicing them daily in the laboratory setting. The subject is presented by the author in a clear didactic manner; the anecdotes and historical references make the book truly a delight to read."

—Dr. Beniamino Barbieri, President, ISS Inc.

"An excellent textbook! This comprehensive overview written at an introductory level covers fundamental aspects, principles of instrumentation, and practical applications of biological fluorescence, while providing many valuable tips. In essence, this book reflects the excellent didactic skills and vast experience of the author in research and teaching fluorescence. Highly recommendable for undergraduate teaching. Also excellent material for active researchers who are learning to apply fluorescence methods."

—Prof. Luis Bagatolli, Center for Biomembrane Physics, Department of Biochemistry and Molecular Biology, University of Southern Denmark

"I am very impressed by the author’s lucid treatment. This book is a ‘must read’ for students and researchers."

—Prof. Amitabha Chattopadhyay, Ph.D., FRSC, J.C. Bose Fellow, Outstanding Scientist, Centre for Cellular & Molecular Biology, Hyderabad

Table of Contents


What Is Fluorescence and How Is It Used?

A Nano-History of Fluorescence

General Concepts

And the Rest

Absorption of Light

Electromagnetic Radiation: Characterization


Relevant Wavelength Range

Absorption of Light by Molecules

Effect of Conjugation on Absorption

Effects of Molecular Environment on Absorption

Practical Considerations


Light Sources

Optical Filters




Miscellaneous Topics

Emission and Excitation Spectra

Emission Spectra

Excitation Spectra

Delayed Fluorescence

Advanced Scanning Methods

Polarization and Anisotropy

Basic Principles

Some Applications of Polarization/Anisotropy

Depolarization via FRET

Time-Resolved Fluorescence

Excited State Lifetimes

Instrumentation Details

Weber’s Algorithm

Time Domain versus Frequency Domain

Magic Angles

Anisotropy Decay/Dynamic Polarization

Dynamic Polarization


Quantum Yields and Quenching

Determination of QYs

Kasha–Vavilov Rule


Quenching and Membrane Systems

Förster Resonance Energy Transfer


Basic Principles

How Do We Determine the Efficiency of Energy Transfer (E)?

The Orientation Factor

How Do We Determine κ2?

Homo-Transfer of Electronic Excitation Energy

Examples of FRET Applications

Advanced Treatments

Brief Overview of Fluorescence Microscopy

Introduction of Fluorophores into Living Cells

Fluorescence Microscopy Approaches

Specialized Techniques


Where Do Fluorophores Come From?

How Does One Choose a Fluorophore?

Intrinsic Probes

Extrinsic Probes


Labeling Proteins In Vitro

Membrane Probes

Nucleic Acid Probes and Nucleotide/Nucleoside



Ion Probes

pH Probes

Molecular Beacons

Voltage-Sensitive Dyes

Miscellaneous Probes

Fluorescent Proteins

Intrinsic Protein Fluorescence




Electronic Energy Transfer in Proteins

Use of Site-Directed Mutagenesis

Tryptophan Analogs


Protein Unfolding/Refolding

Protein–Protein and Protein–DNA Interactions

Tyrosine Studies

Phasors and Protein Fluorescence

Appendix: Rogue’s Gallery of Fluorescence Artifacts and Errors


Additional Reading appears at the end of each chapter.

About the Author

David M. Jameson is a professor in the Department of Cell and Molecular Biology at the John A. Burns School of Medicine, University of Hawaii at Manoa, having previously served there as professor and chairman of the Department of Biochemistry and Biophysics. He earned his Ph.D. in biochemistry from the University of Illinois at Urbana-Champaign, where his graduate thesis advisor was Professor Gregorio Weber. Prior to his move to the University of Hawaii, he was a postdoctoral fellow at the University of Illinois at Urbana-Champaign and the CNRS, University of Paris-South and he was an assistant professor in the Department of Pharmacology at the University of Texas Southwestern Medical Center. Dr. Jameson is the co-organizer of the International Weber Symposia on Innovative Fluorescence Methodologies in Biochemistry and Medicine, which have been held every three years since 1986 (since 1995 in Hawaii). He serves on the editorial boards of The Scientific World Journal, Analytical Biochemistry, and Methods and Applications in Fluorescence. He is a member of the advisory board for the Laboratory for Fluorescence Dynamics (supported by the National Institutes of Health) at the University of California, Irvine. He was the recipient of the 2004 Gregorio Weber Award for Excellence in Fluorescence Theory/Application.

Subject Categories

BISAC Subject Codes/Headings:
SCIENCE / Chemistry / Industrial & Technical
SCIENCE / Microscopes & Microscopy
SCIENCE / Physics