We have studied the optical properties of mammalian cell suspensions to provide a mechanistic basis for interpreting the optical properties of tissues in vivo. Measurements of the wavelength dependence of the reduced scattering coefficient and measurements of the phase function demonstrated that there is a distribution of scatterer sizes. The volumes of the scatterers are equivalent to those of spheres with diameters in the range between ~0.4 and 2.0 mum. Measurements of isolated organelles indicate that mitochondria and other similarly sized organelles are responsible for scattering at large angles, whereas nuclei are responsible for small-angle scattering. Therefore optical diagnostics are expected to be sensitive to organelle morphology but not directly to the size and shape of the cells.

Predictions from Mie theory regarding the wavelength dependence of scattering in tissue from the near UV to the near IR are discussed and compared with experiments on tissue phantoms. For large fiber separations it is shown that rapid, simultaneous measurements of the elastic scatter signal for several fiber separations can yield the absorption coefficient and reduced scattering coefficient. With this information, the size of the scattering particles can be estimated, and this is done for Intralipid. Measurements made at smaller source detector separations support Mie theory calculations, demonstrating that the sensitivity of elastic scatter measurements to morphological features, such as scatterer size, is enhanced when the distance between the source and detector fibers is small.

A diagnostic algorithm for distinguishing malignant from nonmalignant tissue based on the values of the slopes over the wavelength range 330-370 nm has a sensitivity of 100% and specificity of 97% for the limited number of patients in this study.

As part of our ongoing efforts to understand the fundamental nature of light scattering from cells and tissues, we present data on elastic light scattering from isolated mammalian tumor cells and nuclei. The contribution of scattering from internal structures and in particular from the nuclei was compared to scattering from whole cells. Roughly 55% of the elastic light scattering at high-angles (> 40 degrees) comes from intracellular structures. An upper limit of 40% on the fractional contribution of nuclei to scattering from cells in tissue was determined. Using cell suspensions isolated from monolayer cultures at different stages of growth, we have also found that scattering at angles greater than about 110 degrees was correlated with the DNA content of the cells. Based on model calculations and the relative size difference of nuclei from cells in different stages of growth, we argue that this difference in scattering results from changes in the internal structures of the nucleus. This interpretation is consistent with our estimate of 0.2 micron as the mean size of the scattering centers in cells. Additionally, we find that while scattering from the nucleus accounts for a majority of internal scattering, a significant portion must result from scattering off of cytoplasmic structures such as mitochondria.

An understanding of the relationship between tissue structures and light scattering from tissue will help facilitate the development and acceptance of noninvasive optical diagnostics including elastic scattering spectroscopy, diffuse reflectance, and optical coherence tomography. For example, a quantitative model of the structures that scatter light in epithelial cells would allow determination of what structures control the characteristics of in vivo light transport measurements and subsequently could provide a detailed relationship between cellular structures and optical measurements. We have determined the size distribution of refractive index structure variations in epithelial cells as well as in nuclei isolated from epithelial cells from measurements of the angular dependence of polarized light scattering. The quantitative size distributions we obtained for both whole cells and isolated nuclei include particles with effective radii of 2 microm to 10 nm or less and contain orders of magnitude more small particles than large particles. These results demonstrate that not only are biological cells very heterogeneous, but so are the nuclei within them. Light scattering is likely sensitive to structures smaller than those commonly investigated by standard pathology methods.