Raman spectroscopy is a potentially important clinical tool for real-time diagnosis of disease and in situ evaluation of living tissue. The purpose of this article is to review the biological and physical basis of Raman spectroscopy of tissue, to assess the current status of the field and to explore future directions. The principles of Raman spectroscopy and the molecular level information it provides are explained. An overview of the evolution of Raman spectroscopic techniques in biology and medicine, from early investigations using visible laser excitation to present-day technology based on near-infrared laser excitation and charge-coupled device array detection, is presented. State-of-the-art Raman spectrometer systems for research laboratory and clinical settings are described. Modern methods of multivariate spectral analysis for extracting diagnostic, chemical and morphological information are reviewed. Several in-depth applications are presented to illustrate the methods of collecting, processing and analysing data, as well as the range of medical applications under study. Finally, the issues to be addressed in implementing Raman spectroscopy in various clinical applications, as well as some long-term directions for future study, are discussed.

/pdf/prospects-for-in-vivo-raman-spectroscopy-1fkovf1b8p.pdf

Prospects for in vivo Raman spectroscopy.

Prospects forin vivoRaman spectroscopy

We have applied Raman spectroscopy to analyze the chemical composition of microcalcifications occurring in benign and malignant lesions in the human breast. Microcalcifications were initially separated into two categories based on their Raman spectrum: type I, calcium oxalate dihydrate, and type II, calcium hydroxyapatite. Type I microcalcifications were diagnosed as benign, whereas type II were subdivided into benign and malignant categories using principal component analysis, a statistical technique. Although type II microcalcifications are primarily composed of calcium hydroxyapatite, they also contain trace amounts of several biological impurities. Using principal component analysis, we were able to highlight subtle chemical differences in type II microcalcifications that correlate with breast disease. On the basis of these results, we believe that type II microcalcifications formed in benign ducts typically contain a larger amount of calcium carbonate and a smaller amount of protein than those formed in malignant ducts. Using this diagnostic strategy, we were able to distinguish microcalcifications occurring in benign and malignant ducts with a sensitivity of 88% and a specificity of 93%. This is a significant improvement over current X-ray mammography techniques, which are unable to reliably differentiate microcalcifications in benign and malignant breast lesions.

https://cancerres.aacrjournals.org/content/canres/62/18/5375.full.pdf

Identifying Microcalcifications in Benign and Malignant Breast Lesions by Probing Differences in Their Chemical Composition Using Raman Spectroscopy

Raman spectroscopy has the potential to provide real-time, in situ diagnosis of breast cancer during needle biopsy or surgery via an optical fiber probe. Understanding the chemical/morphological basis of the Raman spectrum of breast tissue is a necessary step in developing Raman spectroscopy as a tool for in situ breast cancer diagnosis. To understand the relationship between the Raman spectrum of a sample of breast tissue and its disease state, near-infrared Raman spectroscopic images of human breast tissue were acquired using a confocal microscope. These images were then compared with phase contrast and hematoxylin- and eosin-stained images to develop a chemical/morphological model of breast tissue Raman spectra. This model fits macroscopic tissue spectra with a linear combination of basis spectra derived from spectra of the cell cytoplasm, cell nucleus, fat, β-carotene, collagen, calcium hydroxyapatite, calcium oxalate dihydrate, cholesterol-like lipid deposits and water. Each basis spectrum represents data acquired from multiple patients and, when appropriate, from a variety of normal and diseased states. The model explains the spectral features of a range of normal and diseased breast tissue samples, including breast cancer. It can be used to relate the Raman spectrum of a breast tissue sample to diagnostic parameters used by pathologists. Copyright © 2002 John Wiley & Sons, Ltd.

/pdf/raman-microspectroscopic-model-of-human-breast-tissue-2wh04ra199.pdf

Raman microspectroscopic model of human breast tissue: implications for breast cancer diagnosis in vivo

Infrared (IR) and Raman spectroscopy are emerging biophotonic tools to recognize various diseases. The current review gives an overview of the experimental techniques, data-classification algorithms and applications to assess soft tissues, hard tissues and body fluids. The methodology section presents the principles to combine vibrational spectroscopy with microscopy, lateral information and fiber-optic probes. A crucial step is the classification of spectral data by a variety of algorithms. We discuss unsupervised algorithms such as cluster analysis or principal component analysis and supervised algorithms such as linear discriminant analysis, soft independent modeling of class analogies, artificial neural networks support vector machines, Bayesian classification, partial least-squares regression and ensemble methods. The selected topics include tumors of epithelial tissue, brain tumors, prion diseases, bone diseases, atherosclerosis, kidney stones and gallstones, skin tumors, diabetes and osteoarthritis.

Disease recognition by infrared and Raman spectroscopy.

Phosphate-type kidney stones have been examined by the Raman spectroscopic technique by merely focusing laser light on the cut surface of the stones. Before examination of the kidney stones, Raman spectra of several standard phosphate compounds—such as calcium monobasic phosphate [Ca(H2PO4)2], calcium dibasic phosphate (CaHPO4), calcium tribasic phosphate [Ca10(PO4)6(OH)2], calcium orthophosphate [Ca3(PO4)2], brushite (CaHPO4·2H2O), struvite (MgNH4PO4·6H2O), and hydroxyapatite [Ca10(PO4)6(OH)9],—were obtained. Hydroxyapatite has a distinctive line at 961 cm−1, and one kidney stone examined showed a comparable band. It was concluded that one stone is primarily hydroxyapatite and another one is brushite. The analysis of the kidney stones by Raman spectroscopy is direct, fast, and nondestructive, and does not require tedious sample preparation.

Raman Spectroscopic Identification of Phosphate-Type Kidney Stones:

The Raman scattering technique is used to obtain spectra from four kidney stones. An argon-ion laser (514.5 nm) is focused directly on the surface of the stones. Three stones were known to be composed mainly of hydrates of calcium oxalate, CaC2O4·H2O, and CaC2O4·2H2O. The scattered Raman shift is examined and compared with that of the standard hydrates. Two stones have spectra almost identical to that of calcium oxalate monohydrate. The characteristic bands at 1465 and 1492 cm−1 are easily observed. The spectrum from the third stone shows a band at 1477 cm−1 characteristic of calcium oxalate dihydrate, but it also shows weaker bands at 1465 and 1492 cm−1, which are observed in spectra of calcium oxalate monohydrate. It is concluded that the first two stones are primarily composed of CaC2O4·H2O, while the third stone includes both hydrates in its structure. The fourth stone, from a Russian woman, shows bands at 627, 998, 1038, 1405, 1465, and 1492 cm−1. The first few bands are characteristic of uric acid, while the 1465- and 1492-cm−1 bands indicate calcium oxalate monohydrate. It is possible not only to identify these two components but also to show their relative distribution within the stone.

Raman Spectroscopic Identification of Calcium-Oxalate-Type Kidney Stone:

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Raman Spectroscopic Identification of Phosphate-Type Kidney Stones:

Raman Spectroscopic Identification of Calcium-Oxalate-Type Kidney Stone: