A solid tissue phantom made of agar, Intralipid and black ink is described and characterized. The preparation procedure is fast and easily implemented with standard laboratory equipment. An instrumentation for time-resolved transmittance measurements was used to determine the optical properties of the phantom. The absorption and the reduced scattering coefficients are linear with the ink and Intralipid concentrations, respectively. A systematic decrease of the reduced scattering coefficient dependent on the agar content is observed, but can easily be managed. The phantom is highly homogeneous and shows good repeatability among different preparations. Moreover, agar inclusions can be easily embedded in either solid or liquid matrixes, and no artefacts are caused by the solid-solid or solid-liquid interfaces. This allows one to produce reliable and realistic inhomogeneous phantoms with known optical properties, particularly interesting for studies on optical imaging through turbid media.

Fluorescence lifetime imaging is a rather new and effective tool that can be used to study complex biological samples, either at microscopic or macroscopic levels. The map of the fluorescence lifetime allows one to discriminate amongst different fluorophores and to achieve valuable insights into the behaviour of emitting molecules, leading to information like local pH, oxygen concentration in cells, etc. Moreover, the distribution in space of any fluorescent marker achievable with this technique can be exploited for diagnostic purposes in medicine. After a brief introduction on the motivations for applying fluorescence lifetime imaging in biology and medicine, the basic principles of this technique will be addressed. Then, the two possible implementations of fluorescence lifetime imaging (i.e. the frequency domain and the time domain methods) will be presented. For this purpose, special attention will be devoted to practical aspects of image acquisition and processing, especially for what concerns the time domain method. Then, the analysis of the state-of-the-art systems will include a brief discussion on new concepts that have recently been introduced in this research field. Finally, two interesting applications of fluorescence lifetime imaging will be presented. The former refers to skin tumour detection and has been successfully applied in a preliminary clinical trial, the latter regards DNA chips reading and has been tested only at laboratory level, yet it has produced promising results for its future implementation in commercial systems.

Diffuse optical imaging and spectroscopy of the female breast is an area of active research. We review the present status of this field and discuss the broad range of methodologies and applications. Starting with a brief overview on breast physiology, the remodeling of vasculature and extracellular matrix caused by solid tumors is highlighted that is relevant for contrast in optical imaging. Then, the various instrumental techniques and the related methods of data analysis and image generation are described and compared including multimodality instrumentation, fluorescence mammography, broadband spectroscopy, and diffuse correlation spectroscopy. We review the clinical results on functional properties of malignant and benign breast lesions compared to host tissue and discuss the various methods to improve contrast between healthy and diseased tissue, such as enhanced spectroscopic information, dynamic variations of functional properties, pharmacokinetics of extrinsic contrast agents, including the enhanced permeability and retention effect. We discuss research on monitoring neoadjuvant chemotherapy and on breast cancer risk assessment as potential clinical applications of optical breast imaging and spectroscopy. Moreover, we consider new experimental approaches, such as photoacoustic imaging and long-wavelength tissue spectroscopy.

In vivo absorption and reduced scattering spectra of the human calcaneous from 650 to 1000 nm were assessed using a laboratory system for time-resolved transmittance spectroscopy. Measurements were performed on the calcaneous of seven female volunteers ranging from 26 to 82 years of age. The analysis of the absorption spectra, using a linear combination of the key tissue absorbers (bone mineral, water, lipids, oxy- and deoxyhemoglobin), revealed a general decrease in bone mineral content and an increase in lipids with age, which is in agreement with the aging transformations that occur in bone tissues. The scattering spectra were less effective in detecting such changes in older subjects, showing only a minor decrease in the coefficient for these subjects. The capability to noninvasively quantify bone tissue composition suggests a possible use of optical biopsy for the diagnosis of bone pathologies such as osteoporosis, which are characterized by a progressive reduction and transformation of the mineral in the bone matrix.

Different approaches for absorption and scattering spectroscopy of living tissues are discussed. In particular, a unique system for time-resolved reflectance and transmittance spectroscopy is presented, capable of acquiring in vivo absorption and scattering spectra of diffusive media between 600 and 1000 nm. A review of typical spectra obtained from a variety of tissue structures is shown, including female breast, forearm, abdomen, and forehead. A second-level analysis of the measured spectra permits an estimation of the concentrations of the key tissue absorbers, as well as of the Mie-equivalent scattering radii. Further, absorption and scattering spectra can be used to estimate the penetration depth of light in tissues as a function of wavelength, which is a crucial parameter in view of the possible application of optical in vivo molecular imaging in clinical diagnosis. Finally, an example of the applicability of the methodology to other biological media such as fruits and vegetables is shown.