Bulletin of the American Physical Society
2007 APS March Meeting
Volume 52, Number 1
Monday–Friday, March 5–9, 2007; Denver, Colorado
Session J34: In vivo Imaging and Biomolecular Fibrils |
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Sponsoring Units: DBP Chair: Wolfgang Losert, University of Maryland Room: Colorado Convention Center 404 |
Tuesday, March 6, 2007 11:15AM - 11:27AM |
J34.00001: Crystallographic Properties of Physiological Hydroxyapatite as a Function of Age Th. Leventouri, R. Venturelli, A. Kyriacou Hydroxyapatite with 4-6 wt {\%} B-type carbonate substitution is the major mineral component in our teeth and bones. Crystal structure properties of human teeth as a function of age between 17 and 91 years are investigated. X-ray powder diffraction reveals a partial phase transition from the hexagonal Ca$_{5}$(PO$_{4})_{3}$OH (Hydroxyapatite) to the triclinic Ca$_{4}$H(PO$_{4})_{3}$.2H$_{2}$O (Calcium Hydrogen Phosphate Hydrate) at the 70 year old tooth. This phase becomes predominant in the diffraction pattern of a 91 year old tooth. Correlation of such transition with physical properties of synthetic hydroxyapatite could provide useful insights in dentistry and medicine. [Preview Abstract] |
Tuesday, March 6, 2007 11:27AM - 11:39AM |
J34.00002: Single fluorescent nanodiamond as a cellular biomarker Hsu-Yang Lee, Huan-Cheng Chang, Wunshain Fann Type Ib diamonds emit bright fluorescence at 550--800 nm from nitrogen-vacancy point defects, (N-V)0 and (N-V)-, by high-energy ion beam irradiation and subsequent thermal annealing. The absence of fluorescence intermittency and photobleaching in addition to its non-cytotoxicity and the easiness of surface functionalization make the fluorescent nano-sized diamonds (FND) a promising fluorescent probe for single-particle tracking in heterogeneous environments. We investigated the basic photophysical properties of surface-functionalized single FND particles with average diameter of 35-nm using single-photon and two-photon excitation. The application of tracking single FNDs in HeLa cells was also demonstrated. We found that the photostability of FNDs is not deteriorated by the surface treatment and the brightness of the fluorescence emitted by FNDs is much higher than typical organic dyes. The absorption and emission wavelength of FND, which are well separated from that of the intracellular components, further ensures the good signal to noise ratio for its application as a cellular biomarker. [Preview Abstract] |
Tuesday, March 6, 2007 11:39AM - 11:51AM |
J34.00003: Intracellular Osmolyte Distributions Assessed by $^{1}$H and $^{23}$Na Magnetic Resonance Microscopy Samuel Grant Recently, Magnetic Resonance Microscopy (MRM) has been applied to the high resolution imaging and localized spectroscopy of isolated cells$^{1,2}$. With resolutions $<$40 $\mu $m, these efforts have demonstrated the diverse intracellular environments that can be probed by proton MRM to provide insight into the compartmental diffusion and relaxation of intracellular water and metabolites. In this study, the intracellular distribution of the inorganic osmolyte sodium in isolated single neurons is assessed by MRM through the acquisition of three-dimensional (3D) microimages by direct observation of $^{23}$Na. These efforts are made possible through (a) the use of a specially constructed, double-tuned Radio Frequency (RF) microcoil and (b) the application of a unique, ultra-widebore 21.1-T magnet. Results show an increased sodium signal in the nucleus of the L7 neuron of \textit{aplysia Californica}. These $^{23}$Na findings are compared with MR data that display a heterogeneous distribution of the organic osmolyte betaine, which appears to be localized at high concentrations to the cytoplasm. The link between the intracellular distributions of sodium and other osmolytes in this single neuron model may shed light on intracellular osmoregulatory processes, particularly in response to toxic or pathological perturbations. $^{1}$S.C.Grant, \textit{et al.}, Magn. Reson. Med. 2000. $^{2}$S.C.Grant, \textit{et al.,} Magn. Reson. Med. 2001. [Preview Abstract] |
Tuesday, March 6, 2007 11:51AM - 12:03PM |
J34.00004: Harmonic Generation Spectroscopy of Enzymatic Activity in Live Organisms Jie Fang, Gustavo Cardenas, Shih-Ying Hsu, William Widge, John Miller We report on measurements of harmonics generated by whole cells, chloroplasts, and whole plants in response to applied sinusoidal electric fields. The frequency- and amplitude-dependence of the induced harmonics exhibit features that correlate with physiological processes. In particular, we find that harmonics generated by whole plants and suspensions of chloroplasts are dramatically increased by the presence of light. Systematic studies of the second and third harmonic generation spectra of chloroplast suspensions indicate the following: 1) a broad peak, centered around 10 kHz applied frequency (20 kHz for the second harmonic) appears when the photosynthetic electron transport chain is activated by light in the presence of a suitable electron acceptor, such as ferricyanide; changes observed in the time-dependent harmonic response for fixed frequency are correlated to the presence of light activation of photosynthetic election transport activity. 2) This feature correlates with oxygen evolution activity of photosynthesis. In whole plants, multiple peaks in the light-activated harmonic generation spectra suggest that the method may be able to selectively probe specific photosynthetic activity in plants. [Preview Abstract] |
Tuesday, March 6, 2007 12:03PM - 12:15PM |
J34.00005: Near-Infrared Fluorescence of the NBT/BCIP Chromogenic Stain M. D. McCutchen, L. A. Bumm, D. W. McCauley, L. A. Trinh, M. Bonner-Fraser, S. E. Fraser We demonstrate the previously unreported near infrared (NIR) fluorescence of the dark purple stain formed from 5-bromo-4-chloro-3-indolyl phosphate (BCIP) and nitro blue tetrazolium (NBT). Although the product is a solid with strong optical absorption, its fluorescence enables high cellular resolution imaging of gene expression. We use spectrofluorometry to identify NBT diformazan as the component of the stain that is the fluorophore exhibiting the strong fluorescence signal. The fluorescence shows an intense emission signal (780-910 nm) that is well separated from excitation (645-685 nm). The NBT diformazan fluorescence is also photostable. Because NBT/BCIP is a widely used chromogenic stain, existing staining protocols can also be applied to fluorescence imaging techniques to increase the resolution of gene expression patterns. [Preview Abstract] |
Tuesday, March 6, 2007 12:15PM - 12:27PM |
J34.00006: Development of a `Protein Microscope' to Map Peptide Distributions in Cells J.A. Hoffmann, M.E. Reeves We report on the development of a new instrument, dubbed a `Protein Microscope,' that uses near-field optical techniques to increase the spatial resolution of atmospheric pressure matrix-assisted laser desorption and ionization (AP-MALDI). This functions as a novel front-end for time-of-flight mass spectrometry. Standard protein identification techniques involve homogenization of a tissue sample, which destroys all spatial and temporal information about the expressed proteins. Our new NSOM-based instrument will allow the identification and mapping of proteins expressed in intact cells and tissues, which is of great interest as protein expression connects genomic information with the functioning of an organism. [Preview Abstract] |
Tuesday, March 6, 2007 12:27PM - 12:39PM |
J34.00007: The fate of cells in skin: from clonal analysis to cell kinetics Allon M. Klein, David P. Doupe, Douglas J. Winton, Phil H. Jones, Benjamin D. Simons Biologists are keen to understand the mechanisms of development and maintenance of tissues in mammals. As well as its intrinsic scientific interest, an understanding of the kinetics of cell division has important implications for mechanisms of aging and cancer development. Analysis of cell populations (clones) resulting from progenitor cells provides indirect access to the laws governing cell division and fate. Yet, until recently, the quality of clonal fate data acquired \emph{in vivo} has inhibited reliable quantitative analysis. By addressing a recent, detailed, and extensive experimental study of mammalian skin, we develop a general theoretical framework which shows that the wide range of clonal fate data are consistent with a remarkably simple cell kinetic model. As well as overturning the accepted paradigm for skin maintenance, the analysis introduces a general framework for analysing clone fate data in future experiments. We now have a robust platform to study the effect of drug treatments and the influence of cell mutations on the epidermis. [Preview Abstract] |
Tuesday, March 6, 2007 12:39PM - 12:51PM |
J34.00008: Photo-ionization Potential Threshold of Single Human Fibrinogen Molecule Absorbed onto Silicon Surfaces Xianhua Kong, Jacob Gauguilo, Robert Nemanich Human Plasma Fibrinogen (HPF), which is a protein involved in haemostasis and thrombosis, is known to readily adsorb onto artificial surfaces. Therefore, understanding the absorption process for specific surfaces is critical to establish biocompatibility. In this study, the photo-ionization potential of single HPF molecules adsorbed onto oxidized p-type silicon substrates was studied by photoelectron emission microscopy (PEEM). PEEM, using the spontaneous emission output of the Duke OK-4 free electron laser (FEL), were illuminated at tunable wavelengths between 248 and 310 nm. The photo-ionization potential threshold for single HPF molecules was found to be 4.6 $\pm $ 0.2 eV. The electronic states of the molecule were related to the electronic states of the oxidized Si surfaces. The deduced alignment of the electronic states is consistent with negative charge transfer from the adsorbed fibrinogen to the p-type silicon substrates which would proceed by tunneling through the thin oxidized layer. [Preview Abstract] |
Tuesday, March 6, 2007 12:51PM - 1:03PM |
J34.00009: In vitro, interaction of homotrimers with heterotrimers of type I collagen Sejin Han, Wolfgang Losert, Sergey Leikin The dominant mutations in type I collagen cause a group of diseases, often termed collagen, or connective tissue, diseases: for example, Osteogenesis Imperfecta (OI) characterized by bone fragility and skeletal deformity. The mechanism in which collagen mutations affect on the diseases is still unknown. To understand the fibril assembly and their interactions might provide a key to approaching the cause of the collagen diseases. This study demonstrates that the self-assembly, termed fibrillogenesis, of type I collagen homozygous mutations revealed substantial differences in the kinetics with the absence of lag time and in the morphology of 3D fibril network structure. The heterotrimers (normal) and homotrimers (mutant) in mixtures were segregated within the same fibrils during fibrillogenesis, in correspondence between confocal microscopy and thermodynamic measurements. The efficiency for self-assembly of the homotrimers into fibrils was markedly reduced, while that of the heterotrimers was not affected by the presence of homotrimers with no change in solubility. [Preview Abstract] |
Tuesday, March 6, 2007 1:03PM - 1:15PM |
J34.00010: Determining Beta Sheet Crystallinity in Fibrous Proteins by Thermal Analysis and Infrared Spectroscopy Xiao Hu, David Kaplan, Peggy Cebe We report a study of self-assembled beta pleated sheets in \textit{Bombyx mori} silk fibroin films using thermal analysis and infrared spectroscopy. Crystallization of beta pleated sheets was effected either by heating the films above the glass transition temperature (Tg) and holding isothermally, or by exposure to methanol. The fractions of secondary structural components including random coils, alpha helices, beta pleated sheets, turns, and side chains, were evaluated using Fourier self-deconvolution (FSD) of the infrared absorbance spectra. As crystalline beta sheets form, the heat capacity increment from the TMDSC trace at Tg is systematically decreased and is linearly well correlated with beta sheet content determined from FSD. This analysis of beta sheet content can serve as an alternative to X-ray methods and may have wide applicability to other crystalline beta sheet forming proteins. [Preview Abstract] |
Tuesday, March 6, 2007 1:15PM - 1:27PM |
J34.00011: Resonance Effects in the Ultraviolet Raman Spectroscopy of Collagen in Mineralized Tissues J. W. Ager III, M. Pugach, S. Habelitz, G. Balooch, J. H. Kinney, G. W. Marshall, R. O. Ritchie Ultraviolet resonance Raman spectroscopy (UVRRS) was used to investigate type I collagen in solid tissues including tendon, dentin, and bone. With 244 nm excitation, spectral features from both the amide backbone (amide I, II, and III) and resonance-enhanced side-chain vibrations (Y8a, tyrosine) were observed. This contrasts with reported Raman spectra of proteins in solution excited with similar UV wavelengths, where side chain vibrations, but not strong amide features, are observed. The height of the dominant amide I feature in teeth and bone can be reversibly increased/decreased in dentin by dehydration/rehydration cycles. Also, the amide I peak is relatively stronger in both human bone and dentin from older donors. The strong intensity of the amide I UVRRS feature in these mineralized tissues is attributed to an increase in the width of the $\pi\rightarrow$ $\pi$$^{\textrm{*}}$ amide resonance in collagen compared to the solution phase. These findings suggest that UVRRS can be used as a specific probe of the collagen environment in bone and dentin. [Preview Abstract] |
Tuesday, March 6, 2007 1:27PM - 1:39PM |
J34.00012: Surface modification of cotton and silk fabrics by SF$_{6}$ plasma Satreerat Hodak, Thidarat Supasai, Varong Pavarajarn, Boonchoat Paosawatyanyong Hydrophobic properties are of the interest in fabric and textile manufacturers. We have used SF$_{6}$ plasma to modify the surface of cotton and silk fabrics. We have found that SF$_{6}$ plasma enhances the hydrophobic property of both types of fabrics. The water contact angle of SF$_{6}$-treated fabrics increased from 20 degrees up to 140 degrees. The measured absorption time was found to depend upon the treatment time and RF power, only at the low SF$_{6}$ pressure of 0.005 and 0.05 torr. At higher pressure, all samples achieved high absorption time of about 200 min, regardless of the RF power and treatment time. The morphology changes of fabrics after plasma treatment were characterized by scanning electron microscopy and atomic force microscopy. After plasma treatment, the rms surface roughness of the fibre increased from about 20 nm to 40 nm. From X-ray photoelectron microscopy analysis, we found that the higher the F/C atomic ratio leads to the longer the absorption time or the improved hydrophocity of the fabric. [Preview Abstract] |
Tuesday, March 6, 2007 1:39PM - 1:51PM |
J34.00013: Fundamental Investigations of the Extracellular Proteins Fibrin and Collagen in Microchannel Devices Heather M. Evans, Sarah Koester, Thomas Pfohl Microfluidic structures are particularly amenable to controlled investigations of protein bundle and network formation. Hydrodynamic focusing is utilized to create a diffusion-controlled gradient of reactants, enabling non-equilibrium investigations. We present studies of the blood clotting protein fibrin, a three-dimensional network formed from the enzymatic cleavage of fibrinogen monomers by the protein thrombin. Fibrin is a vital component of blood clots, and has been implicated in a variety of diseases. Real-time fluorescence microscopy and x-ray micro-diffraction are used to quantify supramolecular assembly and provide snapshots of the evolution of fibrin network formation. We also show that collagen, a ubiquitous extracellular protein, can be bundled in situ through the use of a pH gradient. An outlook toward artificial blood vessels arises from the insight that both fibrin and collagen can easily be used to coat microchannel structures. The resulting mesh forms an ideal environment for red blood cells and other cell types. [Preview Abstract] |
Tuesday, March 6, 2007 1:51PM - 2:03PM |
J34.00014: Biomolecule surface patterning by aqueous polymer nanografting (APN) Robert Davis, Katherine Barnett, Jodi Knoebel, Matthew Linford We have demonstrated a method to chemically pattern aqueous polymer layers on the nanoscale. An atomic force microscope (AFM) was used to mechanically remove positively charged polymers from silica and mica surfaces with submicron resolution in liquid. Polyallyl amine (PAA) and polylysine were both been patterned creating 10 and 20 micron boxes with nanometer scale edge transition lengths. These patterns can serve as templates for patterning lipid and protein layers in buffer environments where pH and concentration can be controlled. [Preview Abstract] |
Tuesday, March 6, 2007 2:03PM - 2:15PM |
J34.00015: Mechanical Single Molecule Investigations of SNARE Protein Interactions Wei Liu, Vedrana Montana, Vladimir Parpura, Umar Mohideen We used an Atomic Force Microscope (AFM) to perform single molecule investigations of the SNARE (soluble N-ethyl maleimide-sensitive fusion protein attachment protein receptors) proteins, syntaxin, synaptobrevin and SNAP 25. These proteins are involved in the docking and release of neurotransmitters. The rupture force and extension of the interactions were measured. Chemical reaction rate theory was applied to obtain the energy barrier width and lifetime. Their temperature dependence was also explored. [Preview Abstract] |
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