Susie Boydston-White

Infrared Spectroscopy of Cells and Tissues: Shining Light onto a Novel Subject

T his focal point article aims to introduce the readers to a new spectroscopic endeavor, infrared spectroscopy of entire cells and tissues, and to present an overview of the results from, and the potential of, this research. The ® eld has seen quite a bit of activity, as judged by the number of papers published and patents ® led. But unfortunately, the level of activity in a ® eld is no measure of the quality of the work published: the earlier reports of ``detecting cancer’ ’ by comparing infrared spectra of healthy and diseased samples were sometimes based on rather naive interpretations of the observations and bypassed all ground

Raman and Infrared Microspectral Imaging of Mitotic Cells

We report the first ever Raman and infrared microspectroscopic images of human cells at different stages of mitosis. These spectroscopic methods monitor the distribution of condensed nuclear chromatin, and other biochemical components, utilizing inherent protein and DNA spectral markers, and, therefore, do not require the use of any stains. In conjunction with previously reported data from the G1, S, and G2 phases of the cell cycle, the complete cell division cycle has now been mapped by spectroscopic methods. Although the results reported here do not offer new insights into the distribution of biochemical components during mitosis, the recognition of cell division without the use of stains, and the possibility of doing so on living cells, may be useful for an automatic, spectroscopic determination of the proliferation rates of cells and tissues. Spectral images were constructed by plotting spectral intensities of DNA or protein versus the coordinates from which spectra were recorded. We found that both Raman and infrared intensities depend on the overall chromatin density variation among the individual subphases of mitosis.

Human brain cancer studied by resonance Raman spectroscopy

Abstract. The resonance Raman (RR) spectra of six types of human brain tissues are examined using a confocal micro-Raman system with 532-nm excitation in vitro. Forty-three RR spectra from seven subjects are investigated. The spectral peaks from malignant meningioma, stage III (cancer), benign meningioma (benign), normal meningeal tissues (normal), glioblastoma multiforme grade IV (cancer), acoustic neuroma (benign), and pituitary adenoma (benign) are analyzed. Using a 532-nm excitation, the resonance-enhanced peak at 1548  cm−1 (amide II) is observed in all of the tissue specimens, but is not observed in the spectra collected using the nonresonance Raman system. An increase in the intensity ratio of 1587 to 1605  cm−1 is observed in the RR spectra collected from meningeal cancer tissue as compared with the spectra collected from the benign and normal meningeal tissue. The peak around 1732  cm−1 attributed to fatty acids (lipids) are diminished in the spectra collected from the meningeal cancer tumors as compared with the spectra from normal and benign tissues. The characteristic band of spectral peaks observed between 2800 and 3100  cm−1 are attributed to the vibrations of methyl (─CH3) and methylene (─CH2─) groups. The ratio of the intensities of the spectral peaks of 2935 to 2880  cm−1 from the meningeal cancer tissues is found to be lower in comparison with that of the spectral peaks from normal, and benign tissues, which may be used as a distinct marker for distinguishing cancerous tissues from normal meningeal tissues. The statistical methods of principal component analysis and the support vector machine are used to analyze the RR spectral data collected from meningeal tissues, yielding a diagnostic sensitivity of 90.9% and specificity of 100% when two principal components are used.

Resonance Raman and Raman Spectroscopy for Breast Cancer Detection

Raman spectroscopy is a sensitive method to detect early changes of molecular composition and structure that occur in lesions during carcinogenesis. The Raman spectra of normal, benign and cancerous breast tissues were investigated in vitro using a near-infrared (NIR) Raman system of 785 nm excitation and confocal micro resonance Raman system of 532 nm excitation. A total number of 491 Raman spectra were acquired from normal, benign and cancerous breast tissues taken from 15 patients. When the 785 nm excitation was used, the dominant peaks in the spectra were characteristic of the vibrations of proteins and lipids. The differences between the normal and cancerous breast tissues were observed in both the peak positions and the intensity ratios of the characteristic Raman peaks in the spectral region of 700–1800 cm−1. With 532 nm excitation, the resonance Raman (RR) spectra exhibited a robust pattern of peaks within the region of 500–4000 cm−1. The intensities of four distinct peaks at 1156, 1521, 2854 and 3013 cm−1 detected in the spectra collected from normal breast tissue were found to be stronger in comparison with those collected from cancerous breast tissue. The twelve dramatically enhanced characteristic peaks, including the enhanced amide II peak at 1548 cm−1 in the spectra collected from cancerous breast tissue, distinguished the cancerous tissue from the normal tissue. Principal component analysis (PCA) combined with support vector machine (SVM) analysis of the Raman and RR spectral data yielded a high performance in the classification of cancerous and benign lesions from normal breast tissue.

Zirconia Phase Transformation, Metal Transfer, and Surface Roughness in Retrieved Ceramic Composite Femoral Heads in Total Hip Arthroplasty

Ceramic femoral heads have had promising results as a bearing surface in total hip arthroplasty. Our objective was to evaluate a series of retrieved alumina-zirconia composite ceramic femoral heads for evidence of the tetragonal to monoclinic zirconia phase transformation, metal transfer and articular surface roughness. Raman spectra showed evidence of the zirconia phase transformation in all retrieved specimens, with distinct monoclinic peaks at 183, 335, 383, and 479 cm(-1). All components displayed metal transfer. An increase in the zirconia phase transformation was seen with increasing time in vivo. No correlation between extent of zirconia phase transformation and the surface roughness was found. These short-term results suggest that the use of an alumina-zirconia composite ceramic is a viable option for femoral heads in THA.

Resonance Raman of BCC and normal skin

The Resonance Raman (RR) spectra of basal cell carcinoma (BCC) and normal human skin tissues were analyzed using 532nm laser excitation. RR spectral differences in vibrational fingerprints revealed skin normal and cancerous states tissues. The standard diagnosis criterion for BCC tissues are created by native RR biomarkers and its changes at peak intensity. The diagnostic algorithms for the classification of BCC and normal were generated based on SVM classifier and PCA statistical method. These statistical methods were used to analyze the RR spectral data collected from skin tissues, yielding a diagnostic sensitivity of 98.7% and specificity of 79% compared with pathological reports.

Optical pathology study of human abdominal aorta tissues using confocal micro resonance Raman spectroscopy

Resonance Raman (RR) spectroscopic technique has a high potential for label-free and in-situ detection of biomedical lesions in vivo. This study evaluates the ability of RR spectroscopy method as an optical histopathology tool to detect the atherosclerotic plaque states of abdominal aorta in vitro. This part demonstrates the RR spectral molecular fingerprint features from different sites of the atherosclerotic abdominal aortic wall tissues. Total 57 sites of five pieces aortic samples in intimal and adventitial wall from an autopsy specimen were examined using confocal micro Raman system of WITec 300R with excitation wavelength of 532nm. The preliminary RR spectral biomarkers of molecular fingerprints indicated that typical calcified atherosclerotic plaque (RR peak at 964cm-1) tissue; fibrolipid plaque (RR peaks at 1007, 1161, 1517 and 2888cm-1) tissue, lipid pool with the fatty precipitation cholesterol) with collagen type I (RR peaks at 864, 1452, 1658, 2888 and 2948cm-1) in the soft tissue were observed and investigated.

Analysis of microscopic infrared spectra of individual dried and live human cells

The ability of infrared (IR) spectroscopy to distinguish and map cancerous and non-cancerous tissue has opened the question of the origin of spectral differences between normal and cancerous cells. In this contribution, we report IR spectral maps of individual dried cancer cells, some of them in the process of cell division (mitosis), IR spectra of cells suspended in growth medium, and preliminary results of a statistical analysis of thousands of individual dried cancer cells.

Vulnerable atherosclerotic plaque detection by resonance Raman spectroscopy

Abstract. A clear correlation has been observed between the resonance Raman (RR) spectra of plaques in the aortic tunica intimal wall of a human corpse and three states of plaque evolution: fibrolipid plaques, calcified and ossified plaques, and vulnerable atherosclerotic plaques (VPs). These three states of atherosclerotic plaque lesions demonstrated unique RR molecular fingerprints from key molecules, rendering their spectra unique with respect to one another. The vibrational modes of lipids, cholesterol, carotenoids, tryptophan and heme proteins, the amide I, II, III bands, and methyl/methylene groups from the intrinsic atherosclerotic VPs in tissues were studied. The salient outcome of the investigation was demonstrating the correlation between RR measurements of VPs and the thickness measurements of fibrous caps on VPs using standard histopathology methods, an important metric in evaluating the stability of a VP. The RR results show that VPs undergo a structural change when their caps thin to 66  μm, very close to the 65-μm empirical medical definition of a thin cap fibroatheroma plaque, the most unstable type of VP.

The cell-cycle dependence of the spectra of proliferating normal and neoplastic single cells using confocal resonance Raman microspectroscopy

Confocal resonance Raman (RR) spectra were collected from single proliferating cells and analyzed to detect spectral patterns that are cell-cycle dependent, as a consequence of cellular proliferation — normal or abnormal. The cells’ biochemical age at each time point was confirmed by immunohistochemical staining to identify the presence or absence of cellular components that appear and/or disappear as the cells proceed through the cell-cycle. The RR spectra were collected and compared for each time point as the cells proceeded through the cell cycle to determine what spectral vibrational patterns are cell-cycle dependent. In this study, the question is whether the cell-cycle dependent RR spectral patterns of the vibrational modes observed in proliferating normal and neoplastic single cells are due to a state of cancer or are simply the consequences of the cells’ changing internal biochemistry due to the process of cellular proliferation --- normal or abnormal.