This monograph focuses on modern femtosecond laser microscopes for two photon imaging and nanoprocessing, on laser tweezers for cell micromanipulation as well as on fluorescence lifetime imaging (FLIM) in Life Sciences. The book starts with an introduction by Dr. Wolfgang Kaiser, pioneer of nonlinear optics and ends with the chapter on clinical multiphoton tomography, the novel high resolution imaging technique. It includes a foreword by the nonlinear microscopy expert Dr. Colin Sheppard. Contents Part I: Basics Brief history of fluorescence lifetime imaging The long journey to the laser and its use for nonlinear optics Advanced TCSPC-FLIM techniques Ultrafast lasers in biophotonics Part II: Modern nonlinear microscopy of live cells STED microscopy: exploring fluorescence lifetime gradients for super-resolution at reduced illumination intensities Principles and applications of temporal-focusing wide-field two-photon microscopy FLIM-FRET microscopy TCSPC FLIM and PLIM for metabolic imaging and oxygen sensing Laser tweezers are sources of two-photon effects Metabolic shifts in cell proliferation and differentiation Femtosecond laser nanoprocessing Cryomultiphoton imaging Part III: Nonlinear tissue imaging Multiphoton Tomography (MPT) Clinical multimodal CARS imaging In vivo multiphoton microscopy of human skin Two-photon microscopy and fluorescence lifetime imaging of the cornea Multiscale correlative imaging of the brain Revealing interaction of dyes and nanomaterials by multiphoton imaging Multiphoton FLIM in cosmetic clinical research Multiphoton microscopy and fluorescence lifetime imaging for resection guidance in malignant glioma surgery Non-invasive single-photon and multi-photon imaging of stem cells and cancer cells in mouse models Bedside assessment of multiphoton tomography

This monograph demonstrates the latest developments in two-photon fluorescence microscopy and second-harmonic generation (SHG) microscopy, including coverage of high-resolution microscopy methods, such as STED microscopy. A special focus lies on clinical applications of these methods, e.g. in dermatology, ophtalmology, neuro sciences and cell biology.

The use of nanoparticles has increased in consumer products in recent decades; however, concerns regarding their safety remain. Zinc oxide is used in sunblocking and may generate free radicals in response to UV illumination, leading to DNA damage and an immunological response. With high-resolution, high-contrast imaging in biological tissue, multiphoton microscopy is able to separate nanoparticles signals from endogenous fluorophores. It has been proven to be very useful in imaging penetration of zinc oxide nanoparticles in skin and in combination with fluorescence lifetime imaging microscopy study cellular function as well. This chapter aims to review the use of these imaging techniques in studying the uptake and distribution of nanoparticles in skin and liver. Due to the questionable clinical use and possible toxicity of nanoparticles, it is important to study their pharmacokinetics. Some nanomaterials have been identified as relatively toxic to humans and a few metal nanoparticles have been reported to penetrate and be detected in blood. Multiphoton microscopy has high resolution and is able to visualize nanoparticles, due to their optical properties, in vivo. The addition of fluorescence lifetime imaging makes it possible to measure the physiochemical environment, with outputs that can be statistically analyzed, posing an advantage over fluorescence intensity imaging only.

During the past two decades, there has been an increasing appreciation of the significant value that lifetime-based techniques can add to biomedical studies and applications of fluorescence. Bringing together perspectives of different research communities, Fluorescence Lifetime Spectroscopy and Imaging: Principles and Applications in Biomedical Diagnostics explores the remarkable advances in time-resolved fluorescence techniques and their role in a wide range of biological and clinical applications. Broadly accessible, the book captures the state-of-the-art of fluorescence lifetime metrology and imaging and provides current perspectives on their applications to biomedical studies of intact tissues and medical diagnosis. The text introduces these techniques within the wider context of fluorescence spectroscopy and describes basic principles underlying current instrumentation for fluorescence lifetime imaging and metrology (FLIM). It also covers the wide range of methods, including single channel (point) spectroscopy, fluorescence lifetime imaging microscopy, and single- and multi-photon excitation. Edited by pioneers in this field, with contributions from leading experts, the book includes an overview of complementary techniques that help researchers beginning FLIM research. It offers a comprehensive treatment of fundamental principles, instrumentation, analytical methods, and applications. It also provides an overview of the label-free contrast available from lifetime measurements of tissue autofluorescence and the prospects for exploiting this for clinical applications and biomedical research including drug discovery.

Nanophotonics has emerged rapidly into technological mainstream with the advent and maturity of nanotechnology available in photonics and enabled many new exciting applications in the area of biomedical science and engineering that were unimagined even a few years ago with conventional photonic engineering techniques. Handbook of Nanophotonics in Biomedical Engineering is intended to be a reliable resource to a wealth of information on nanophotonics that can inspire readers by detailing emerging and established possibilities of nanophotonics in biomedical science and engineering applications. This comprehensive reference presents not only the basics of nanophotonics but also explores recent experimental and clinical methods used in biomedical and bioengineering research. Each peer-reviewed chapter of this book discusses fundamental aspects and materials/fabrication issues of nanophotonics, as well as applications in interfaces, cell, tissue, animal studies, and clinical engineering. The organization provides quick access to current issues and trends of nanophotonic applications in biomedical engineering. All students and professionals in applied sciences, materials, biomedical engineering, and medical and healthcare industry will find this essential reference book highly useful.