Biomedical subjects

Bayden R Wood

Publications and source records attributed to Bayden R Wood.

14 recordsLinked to original sources

PubMed2007

Focal plane array infrared imaging: a new way to analyse leaf tissue.

* Here, a new approach to macromolecular imaging of leaf tissue using a multichannel focal plane array (FPA) infrared detector was compared with the proven method of infrared mapping with a synchrotron source, using transverse sections of leaves from a species of Eucalyptus. * A new histological method was developed, ideally suited to infrared spectroscopic analysis of leaf tissue. Spatial resolution and the signal-to-noise ratio of the FPA imaging and synchrotron mapping methods were compared. * An area of tissue 350 microm(2) required approx. 8 h to map using the synchrotron technique and approx. 2 min to image using the FPA. The two methods produced similar infrared images, which differentiated all tissue types in the leaves according to their macromolecular chemistry. * The synchrotron and FPA methods produced similar results, with the synchrotron method having superior signal-to-noise ratio and potentially better spatial resolution, whereas the FPA method had the advantage in terms of data acquisition time, expense and ease of use. FPA imaging offers a convenient, laboratory-based approach to microscopic chemical imaging of leaves.

Eucalyptus↗

PubMed2006

Resonance Raman spectroscopy of red blood cells using near-infrared laser excitation.

Resonance Raman spectra of oxygenated and deoxygenated functional erythrocytes recorded using 785 nm laser excitation are presented. The high-quality spectra show a mixture of enhanced A(1g), A(2g), B(1g), B(2g), E(u) and vinyl modes. The high sensitivity of the Raman system enabled spectra from four oxygenation and deoxygenation cycles to be recorded with only 18 mW of power at the sample over a 60-minute period. This low power prevented photo-/thermal degradation and negated protein denaturation leading to heme aggregation. The large database consisting of 210 spectra from the four cycles was analyzed with principal components analysis (PCA). The PC1 loadings plot provided exquisite detail on bands associated with the oxygenated and deoxygenated states. The enhancement of a band at 567 cm(-1), observed in the spectra of oxygenated cells and the corresponding PC1 loadings plot, was assigned to the Fe-O(2) stretching mode, while a band appearing at 419 cm(-1) was assigned to the Fe-O-O bending mode based on previous studies. For deoxygenated cells, the enhancement of B(1g) modes at 785 nm excitation is consistent with vibronic coupling between band III and the Soret transition. In the case of oxygenated cells, the enhancement of iron-axial out-of-plane modes and non-totally symmetric modes is consistent with enhancement into the y,z-polarized transition a(iu)(pi)-->d(xz)+O(2)(pi(g)) centered at 785 nm. The enhancement of non-totally symmetric B(1g) modes in oxygenated cells suggests vibronic coupling between band IV and the Soret band. This study provides new insights into the vibrational dynamics, electronic structure and resonant enhancement of heme moieties within functional erythrocytes at near-IR excitation wavelengths.

Cells, Cultured↗

PubMed2006

A three-dimensional multivariate image processing technique for the analysis of FTIR spectroscopic images of multiple tissue sections.

BACKGROUND: Three-dimensional (3D) multivariate Fourier Transform Infrared (FTIR) image maps of tissue sections are presented. A villoglandular adenocarcinoma from a cervical biopsy with a number of interesting anatomical features was used as a model system to demonstrate the efficacy of the technique. METHODS: Four FTIR images recorded using a focal plane array detector of adjacent tissue sections were stitched together using a MATLAB routine and placed in a single data matrix for multivariate analysis using Cytospec. Unsupervised Hierarchical Cluster Analysis (UHCA) was performed simultaneously on all 4 sections and 4 clusters plotted. The four UHCA maps were then stacked together and interpolated with a box function using SCIRun software. RESULTS: The resultant 3D-images can be rotated in three-dimensions, sliced and made semi-transparent to view the internal structure of the tissue block. A number of anatomical and histopathological features including connective tissue, red blood cells, inflammatory exudate and glandular cells could be identified in the cluster maps and correlated with Hematoxylin & Eosin stained sections. The mean extracted spectra from individual clusters provide macromolecular information on tissue components. CONCLUSION: 3D-multivariate imaging provides a new avenue to study the shape and penetration of important anatomical and histopathological features based on the underlying macromolecular chemistry and therefore has clear potential in biology and medicine.

Journal Article↗

PubMed2006

Haemozoin (beta-haematin) biomineralization occurs by self-assembly near the lipid/water interface.

Several blood-feeding organisms, including the malaria parasite detoxify haem released from host haemoglobin by conversion to the insoluble crystalline ferriprotoporphyrin IX dimer known as haemozoin. To date the mechanism of haemozoin formation has remained unknown, although lipids or proteins have been suggested to catalyse its formation. We have found that beta-haematin (synthetic haemozoin) forms rapidly under physiologically realistic conditions near octanol/water, pentanol/water and lipid/water interfaces. Molecular dynamics simulations show that a precursor of the haemozoin dimer forms spontaneously in the absence of the competing hydrogen bonds of water, demonstrating that this substance probably self-assembles near a lipid/water interface in vivo.

Animals↗

PubMed2006

Elucidation of interaction between metal-free tetraphenylporphine and surface Ag atoms through temporal fluctuation of surface-enhanced resonance Raman scattering and background-light emission.

We have observed simultaneously temporal fluctuation of surface-enhanced resonance Raman scattering (SERRS) and its background-light emission from single Ag nanoaggregates that were adsorbed with metal-free tetraphenylporphine (H(2)TPP) molecules. We found that temporally stable SERRS spectra showed clearly a SERRS band that is attributed to a stretching mode of a chemical bond between a carbon atom and a non-hydrogenated nitrogen atom (C(alpha)-N). This stretching mode was not observed in regular resonance Raman spectra which are free from surface enhancement. On the other hand, we also found that temporally unstable SERRS spectra did not clearly show a C(alpha)-N stretching mode in SERRS bands. Furthermore, temporally stable SERRS spectra were accompanied by temporally stable background-light emission. Kobayashi et al. [J. Phys. Chem. 1985, 89, 5174] reported that formation of an Ag-N bond between surface Ag atoms and non-hydrogenated N atoms in a pyrrole ring enhances the intensity of a C(alpha)-N stretching mode. Thus, the observed relationship between clear appearance of a C(alpha)-N stretching mode and temporal stability of SERRS plus background-light emission strongly suggests that formation of a stable Ag-N bond suppresses fluctuation of both SERRS and background-light emission. Furthermore, the observed relationship implies that chemical contribution to SERRS is stabilization of H(2)TPP molecules that are adsorbed on SERRS-active sites by formation of Ag-N bonds. Additionally, we attributed background-light emission to luminescence of complexes between H(2)TPP molecules and surface Ag atoms considering possible formation of Ag-N bonds, synchronized SERRS intensity with background-light emission intensity, blue-shifted background-light emission maxima from normal fluorescence maxima, and previous reports related to electronic structures of H(2)TPP molecules on Ag surfaces.

Journal Article↗

PubMed2006

Resonance Raman spectroscopy in malaria research.

In recent years, the field of Raman spectroscopy has witnessed a surge in technological development, with the incorporation of ultrasensitive, charge-coupled devices, improved laser sources and precision Rayleigh-filter systems. This has led to the development of sensitive confocal micro-Raman spectrometers and imaging spectrometers that are capable of obtaining high spatial-resolution spectra and images of subcellular components within single living cells. This review reports on the application of resonance micro-Raman spectroscopy to the study of malaria pigment (hemozoin), a by-product of hemoglobin catabolization by the malaria parasite, which is an important target site for antimalarial drugs. The review aims to briefly describe recent studies on the application of this technology, elucidate molecular and electronic properties of the malaria pigment and its synthetic analog beta-hematin, provide insight into the mechanism of hemozoin formation within the food vacuole of the parasite, and comment on developing strategies for using this technology in drug-screening protocols.

Animals↗

PubMed2005

Mapping of nutrient-induced biochemical changes in living algal cells using synchrotron infrared microspectroscopy.

High quality Fourier transform infrared (FTIR) spectra were acquired from living Micrasterias hardyi cells maintained in an IR transparent flow-through cell using a FTIR microscope coupled to a synchrotron light source. Spectral maps of living, nutrient-replete cells showed band intensities consistent with the known location of the nucleus and the chloroplasts. These were very similar to maps acquired from fixed, air-dried cells. Bands due to lipids were lowest in absorbance in the region of the nucleus and highest in the chloroplast region and this trend was reversed for the absorbance of bands attributed to protein. Spectra acquired in 10 microm steps across living phosphorus-starved (P-starved) cells, repeated approximately every 30 min, were consistent over time, and bands correlated well with the known position of the nucleus and the observed chloroplasts, corroborating the observations with replete cells. Experiments in which missing nutrients were re-supplied to starved cells showed that cells could be maintained in a functional state in the flow-through cell for up to one day. Nitrogen-starved cells re-supplied with N showed an increase in lipid in all positions measured across the cell over a 23 h period of re-supply, with the largest increases occurring in positions where the chloroplasts were observed. Re-supply of phosphorus to P-starved cells produced no changes in bands attributable to lipid or protein. Due to their thin cell body ( approximately 12 microm) and large diameter ( approximately 300 microm) Micrasterias sp. make an ideal spectroscopic model to study nutrient kinetics in algal cells.

Eukaryota↗

PubMed2005

A portable Raman acoustic levitation spectroscopic system for the identification and environmental monitoring of algal cells.

We report the coupling of a portable Raman spectrometer to an acoustic levitation device to enable environmental monitoring and the potential taxonomic identification of microalgae. Spectra of living cells were recorded at 785 nm using a fiber-optic probe coupled to a portable Raman spectrometer. The spectra exhibit an excellent signal-to-noise ratio and clearly show bands from chlorophyll a and beta-carotene. Spectra of levitated photobleached microalgae clearly show a reduction in chlorophyll a concentration relative to beta-carotene after 10 min of exposure to a quartz halogen lamp. Spectra recorded from levitated nitrogen-limited cells also show a significant reduction in bands associated with chlorophyll a, as compared to nitrogen-replete cells. To investigate the diagnostic capability of the technique, four species of microalgae were analyzed. Good quality spectra of all four species were obtained showing varying ratios of beta-carotene to chlorophyll. The combination of an acoustic levitation device and a portable Raman spectrometer shows potential as a taxonomic and environmental monitoring tool with direct application to field studies in remote environments.

Chlorophyta↗

PubMed2005

Mie-type scattering and non-Beer-Lambert absorption behavior of human cells in infrared microspectroscopy.

We report infrared microspectral features of nuclei in a completely inactive and contracted (pyknotic) state, and of nuclei of actively dividing cells. For pyknotic nuclei, the very high local concentration of DNA leads to opaqueness of the chromatin and, consequently, the absence of DNA signals in the IR spectra of very small nuclei. However, these nuclei can be detected by their scattering properties, which can be described by the Mie theory of scattering from dielectric spheres. This scattering depends on the size of the nucleus; consequently, quite different scattering cross-sections are calculated and observed for pyknotic and mitotic nuclei.

Absorption↗

PubMed2004

Resonance Raman spectroscopy reveals new insight into the electronic structure of beta-hematin and malaria pigment.

Resonance Raman spectra of beta-hematin and hemin are reported for a range of excitation wavelengths including 406, 488, 514, 568, 633, 780, 830, and 1064 nm. Dramatic enhancement of A(1g) modes (1570, 1371, 795, 677, and 344 cm(-1)), ring breathing modes (850-650 cm(-1)), and out-of-plane modes including iron-ligand modes (400-200 cm(-1)) were observed when irradiating with 780- and 830-nm laser excitation wavelengths for beta-hematin and to a lesser extent hemin. Absorbance spectra recorded during the transformation of hemin to beta-hematin showed a red-shift of the Soret and Q (0-1) bands, which has been interpreted as excitonic coupling resulting from porphyrin aggregation. A small broad electronic transition observed at 867 nm was assigned to a z-polarized charge-transfer transition d(xy) --> e(g)(pi). The extraordinary band enhancement observed when exciting with near-infrared excitation wavelengths in beta-hematin when compared to hemin is explained in terms of an aggregated enhanced Raman scattering hypothesis based on the intermolecular excitonic interactions between porphyrinic units. This study provides new insight into the electronic structure of beta-hematin and therefore hemozoin (malaria pigment). The results have important implications in the design and testing of new anti-malaria drugs that specifically interfere with hemozoin formation.

Animals↗

PubMed2003

Raman imaging of hemozoin within the food vacuole of Plasmodium falciparum trophozoites.

Micro-Raman spectra of hemozoin encapsulated within the food vacuole of a Plasmodium falciparum-infected erythrocyte are presented. The spectrum of hemozoin is identical to the spectrum of beta-hematin at all applied excitation wavelengths. The unexpected observation of dramatic band enhancement of A(1g) modes including nu(4) (1374 cm(-1)) observed when applying 780 nm excitation enabled Raman imaging of hemozoin in the food vacuole. This unusual enhancement, resulting from excitonic coupling between linked porphyrin moieties in the extended porphyrin array, enables the investigation of hemozoin within its natural environment for the first time.

Animals↗

PubMed2003

Removal of blood components from cervical smears: implications for cancer diagnosis using FTIR spectroscopy.

Red and white cell lysis buffers were investigated to determine their ability to remove blood components from cervical smears prior to IR microspectroscopy. A white cell lysis buffer was effective in increasing the spectral reproducibility and sample homogeneity and in reducing the presence of inflammatory exudate, particularly leukocytes, from cervical smears. The reduction of leukocytes appeared to cause abnormal samples to be grouped with normal samples, resulting in poor discrimination. Despite differences between the cytological and histological diagnoses of cervical abnormalities, the results indicate that the differences seen in the spectra of exfoliated malignant and nonmalignant cervical cells could be due to the presence of inflammation.

Blood Cells↗

PubMed2002

Micro-Raman characterization of high- and low-spin heme moieties within single living erythrocytes.

Raman microspectroscopy was applied to monitor porphyrin perturbation associated with the tense-relaxed (T --> R) state transition of hemoglobin (Hb) within a single red blood cell. The spectra exhibited dramatic changes in the spin state region (1650-1500 cm(-1)) and the methine C-H deformation region (1250-1200 cm(-1)) between the 27- and 54-min interval following reoxygenation. Principal components analysis revealed a sigmoidal curve when the PC1 scores versus time were plotted. The inflection point on the curve corresponded to the T --> R transition point spectrum, indicating the cell is composed of Hb in a mixture of spin states. The results of this work illustrate the dynamics of porphyrin perturbation during erythrocyte respiration and may have applications in the diagnosis and treatment of red blood cell disorders including thalassemia and sickle cell anemia.

Erythrocytes↗

PubMedDate not supplied

Raman microspectroscopy and imaging provides insights into heme aggregation and denaturation within human erythrocytes.

The oxygenation process of a human erythrocyte is monitored using a Raman microimaging technique. Raman images of the 1638 cm(-1) band are recorded in the oxygenated and deoxygenated state using only 120 s of laser exposure and approximately 1 mW of defocused laser power. The images show hemoglobin oxygenating and deoxygenating within the cell. Prolonged laser imaging exposure (<180 s) at low temperatures results in photoinduced and/or thermal degradation. The effect of thermal degradation is investigated by recording spectra of erythrocytes as a function of temperature between 4 and 52 degrees C. Five bands at 1396, 1365, 1248, 972, and 662 cm(-1) are identified as markers for heme aggregation. Raman images recorded of cells after prolonged laser exposure appear to show heme aggregation commencing in the middle and moving toward the periphery of the cell. UV-visible spectra of erythrocytes show the Soret band to be broader and red shifted (approximately 3 nm) at temperatures between 45 and 55 degrees indicative of excitonic interactions. It is postulated that the enhancement of the aggregation marker bands observed at 632.8-nm excitation results primarily from excitonic interactions between the aggregated hemes in response to protein denaturation. The results have important medical implications in detecting and monitoring heme aggregation associated with hemopathies such as sickle cell disease.

Erythrocytes↗