Abstract-Pathologic analysis of the biologic effects and mechanisms of laser-tissue interactions requires correlation of the irradiation parameters with the biologic status and response of the target tissues over time. The photobiologic mechanisms of laser-induced tissue injury can be separated into three categories, photochemical, photothermal and photomechanical. Anatomic pathologic analysis of laser-induced lesions reveals alterations that represent either specific markers of the photobiologic mechanism or non-specific reactions to tissue injury. Repair, regeneration and wound healing of laser induced lesions appear to be non-specific responses to the type of tissue damage rather than the photobiologic mechanism producing the lesion.

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In this study, we investigate the potential of near-infrared Raman spectroscopy to differentiate cervical precancers from normal tissues, inflammation and metaplasia and to differentially diagnose low-grade and high-grade precancers. Near infrared Raman spectra were measured from 36 biopsies from 18 patients in vitro. Detection algorithms were developed and evaluated relative to histopathologic examination. Algorithms based on empirically selected peak intensities, ratios of peak intensities and a combination of principal component analysis for data reduction and Fisher discriminant analysis for classification were investigated. Spectral peaks were tentatively identified from measured spectra of potential chromophores. Empirically selected normalized intensities can differentiate precancers from other tissues with an average sensitivity and specificity of 88 +/- 4% and 92 +/- 4%. Ratios of unnormalized intensities can differentiate precancers from other tissues with a sensitivity and specificity of 82% and 88% and high-grade from low-grade lesions with a sensitivity and specificity of 100%. Using multivariate methods, intensities at eight frequencies can be used to differentiate precancers from all other tissues with a sensitivity and specificity of 82% and 92% in an unbiased test. Raman algorithms can potentially separate benign abnormalities such as inflammation and metaplasia from precancers. Comparison of tissue spectra to published and measured chromophore spectra indicate that the most likely primary contributors to the tissue spectra are collagen, nucleic acids, phospholipids and glucose 1-phosphate. These results suggest that near-infrared Raman spectroscopy can be used for cervical precancer diagnosis and may be able to accurately separate samples with inflammation and metaplasia from precancer.

A portable fluorimeter was developed and utilized to acquire fluorescence spectra from 381 cervical sites in 95 patients at 337, 380 and 460 nm excitation immediately prior to colposcopy. A multivariate statistical algorithm was used to extract clinically useful information from tissue spectra acquired in vivo. Two full-parameter algorithms were developed using tissue fluorescence emission spectra at all three excitation wavelengths (161 excitation-emission wavelength pairs) for cervical precancer (squamous intraepithelial lesion [SIL]) detection: a screening algorithm that discriminates between SIL and non-SIL with a sensitivity of 82 +/- 1.4% and specificity of 68 +/- 0.0%, and a diagnostic algorithm that differentiates high-grade SIL from non-high-grade SIL with a sensitivity and specificity of 79 +/- 2% and 78 +/- 6%, respectively. Multivariate statistical analysis was also employed to reduce the number of fluorescence excitation-emission wavelength pairs needed to redevelop algorithms that demonstrate a minimum decrease in classification accuracy. Two reduced-parameter algorithms that employ fluorescence intensities at only 15 excitation-emission wavelength pairs were developed: the screening algorithm differentiates SIL from non-SIL with a sensitivity of 84 +/- 1.5% and specificity of 65 +/- 2% and the diagnostic algorithm discriminates high-grade SIL from non-high-grade SIL with a sensitivity and specificity of 78 +/- 0.7% and 74 +/- 2%, respectively. Both the full-parameter and reduced-parameter screening algorithms discriminate between SIL and non-SIL with a similar specificity (+/-5%) and a substantially improved sensitivity relative to Pap smear screening. A comparison of the full-parameter and reduced-parameter diagnostic algorithms to colposcopy in expert hands indicates that all three have a very similar sensitivity and specificity for differentiating high-grade SIL from non-high-grade SIL.

Oblique incidence reflectometry is a simple and accurate method for measuring the absorption and the reduced-scattering coefficients of turbid media. We used this technique to deduce absorption and reduced-scattering spectra from wavelength-resolved measurements of the relative diffuse reflectance profile of white light as a function of source-detector distance. In this study, we measured the absorption and the reduced-scattering coefficients of chicken breast tissue in the visible range ͑400 -800 nm͒ with the oblique incidence probe oriented at 0°and 90°relative to the muscle fibers. We found that the deduced optical properties varied with the probe orientation. Measurements on homogenized chicken breast tissue yielded an absorption spectrum comparable with the average of the absorption spectra for 0°and 90°probe orientations measured on the unhomogenized tissue. The reduced-scattering spectrum for homogeneous tissue was greater than that acquired for unhomogenized tissue taken at either probe orientation. This experiment demonstrated the application of oblique-incidence, fiber-optic reflectometry to measurements on biological tissues and the effect of tissue structural anisotropy on optical properties.