|
|
Atomic force microscopy in the region of biomacromolecules
Vančura, Martin ; Kopecký, Vladimír (advisor) ; Bednárová, Lucie (referee)
Atomic force microscopy (AFM) enables sample imaging at the micro and nanoscale. Recently, the method is applied to investigate biomacromolecules. Here, we describe the basic principles of AFM with a special emphasis for bioapplications. We tested experimental abilities of Alpha 300 - the Raman microscope with AFM/SNOM accessory from WITec company. The ability of AFM to study objects of cellular dimensions was demonstrated on erythrocytes and green algae Desmodesmus quadricauda. We were able to observe growing of lysozyme protein fibrils on day scale - from dimensions of seeds (~3 nm height) up to fibrils itself (3-10 nm height and 100 nm up to micrometers length). Subsequently, we observed separate protein molecules of thyroglobulin (~6 nm) and also γ-globulin (~3 nm). It seems plausible to image objects up to 2 nm dimensions by the given device with respect to the signal/noise ratio.
|
|
|
Study of nucleic acids by means of drop coating deposition Raman microspectroscopy (DCDR)
Souček, Pavel ; Kopecký, Vladimír (advisor) ; Mojzeš, Peter (referee)
The work was focused on possibilities of the new technique of nonresonance Raman spectroscopy - drop coating deposition Raman (DCDR) spectroscopy upon study of nucleic acids. DCDR spectroscopy is based on deposition of a small droplet of the studied sample on a hydrophobic surface, where after evaporation of the solvent, ring of the studied material in glass phase is formed. That way the material is concentrated and upon measurements by means of Raman microspectrometry the increase of signal of several orders of magnitude is observed with respect to the standard technique of sample measurements in solvent. In this work we studied behavior of DNA 12-mer and DNA with 3000 base pairs. Our research showed that DCDR spectra of DNA dissolved in deionized water can be measured up to concentration of 30 M per base. The DCDR spectra are similar to those measured from solvents. Nevertheless, the deposited sample never forms a ring. It was shown that the size of nucleic acids didn't play dominant role upon ring formation but their charge which lead to repulsion of the molecules. After adding of sodium ions and subsequently also magnesium ions the ring formation was observed. Factor analysis of spectral maps demonstrated that formed rings are not fully homogenous and that upon their origin condensation of DNA could...
|
|
|
Raman spectroscopy as the tool for Alzheimer's disease diagnostics
Tesař, Adam ; Kopecký, Vladimír (advisor) ; Jelínek, Otakar (referee)
Alzheimer disease (AD) is the most frequent dementia. The prevalence is approximately 10% in 65 years old people. The current treatment is only progression protective, therefore it is crucial to find a new diagnostic approach for diagnosing AD in early stage. We analysed a set of 55 patients by the drop coating deposition Raman spectroscopy with the goal to verify previously published high sensitivity of the AD spectroscopic diagnosis in cerebral spinal fluid (CSF) and to find a new diagnostic method for blood serum (BS). We optimized measurement conditions for BS. The results were evaluated by the cluster analysis and the principal component analysis. The small set of samples exhibited high sensitivity in both CSF and BS but that distinctly decreased in the whole set. The results for CSF were affected by the choice of the analysed spectral interval. The best for AD diagnose was the interval containing peaks at 980, 1080 and 1249 cm-1.The results for BS have been the most sensitive in the whole spectral range. They have low sensitivity but high specificity for AD (92%). The usage of neural networks has conversely high sensitivity and low specificity in both sets of samples of BS and CSF. Powered by TCPDF (www.tcpdf.org)
|
|
|
Combination of AFM and micro-spectroscopy applied on nanoparticles
Doležal, Pavel ; Fučíková, Anna (advisor) ; Kopecký, Vladimír (referee)
This thesis is about two very modern scientific methods used to study the world in smaller than micro scale. The first one of them is atomic force microscopy (AFM) which is used to measure the topology of a sample at the atomic scale. The second one is micro-spectroscopy which is used to study the nature of emission spectra of nanoparticles. The goal of this thesis is to combine these two methods so that it can be used to measure in detail the size and the shape of nanoparticles and in the same time study their emission spectra. In the first part of the thesis I present both methods, their principles, benefits and applications. In the second part I describe performed measurements, such as scale and directional calibration of measuring apparatuses cameras and selection of suitable sample carrier. At the end of the second part, I present my proposed procedure how to pair the same sample location studied by AFM and micro-scpectroscopy. The procedure consist of 20 steps and with its use it will be possible to study luminescence emission in more detail, especially for example examine its dependence on the shape and size of the nanoparticle. The ability to measure the effect of the distance between two nanoparticles on their emission spectra is also very promising.
|
|
|
Drop coating deposition Raman microspectroscopy (DCDR) of carbohydrates
Novák, David ; Kopecký, Vladimír (advisor) ; Kočišová, Eva (referee)
The work is focused on the usage of drop-coating Raman (DCDR) spectroscopy in the study of carbohydrates. DCDR spectroscopy offers a lowering of the detection limits by the coffee-ring effect and minimization of the water signal while keeping similarity to the spectrum obtained from the solution. The similarity is achieved due to the glassy phase, in which molecules have a random orientation and probably retain the first solvation envelope. However, for small molecules, the situation is complicated by both the low efficiency of the coffee-ring effect and the tendency to crystallize. In this work, we studied glucose, galactose, lactose, maltose, trehalose and cyclodextrin to cover the basic structural diversity of sugars - from monosaccharides, disaccharides to oligosaccharides. It was successfully found the concentration range for DCDR spectroscopy of mono and disaccharides (ca. 1-30 mM), while maintaining the glassy phase of the deposit, and to point out the complications associated with the crystallization of polysaccharides. We compared the obtained DCDR spectra with the spectra of polycrystalline samples, from which they clearly differed. Subsequently, we measured the concentration dependences, up to saturated solutions, whose Raman spectra were analysed by factor analysis to capture the...
|
|
|
Phosphine and nitrous oxide as false-positive biosignature gases in planetary spectra
Kaiserová, Tereza ; Ferus, Martin (advisor) ; Kopecký, Vladimír (referee)
5 Abstract This study is dedicated to evaluating the potential for a novel pathway of abiotic synthesis of phosphine (PH3) from phosphorus trioxide (P4O6) by a photocatalytic mechanism on aerosols. The mechanism might explain the recent possible detection of phosphine on Venus. This scenario has recently been suggested theoretically by our team based on an analogy with methane production form carbon dioxide on acidic photochemically-active surfaces of materials, which may account for a possible source of methane on Mars. Methane, just like phosphine, was suggested as an indicator of life on terrestrial planets, including Mars. The theoretical testing of photochemical phosphine generation suggests that even if this gas is present in the atmosphere of Venus it cannot be considered as an indicator of life until the suggested mechanism is excluded theoretically, or by experimental results, or direct evidence of life on Venus. This thesis will be followed by preparation of sophisticated experiments and intensive laboratory research addressing this. Furthermore, the role of nitrous oxide as another false positive biosignature was evaluated in this study. The presence of nitrous oxide can also be explained by processes other than biological, particularly on early planets. This study specifically demonstrates the...
|
|
|
Consequences of heavy bombardment for chemical and prebiotic evolution on early Mars and Earth
Petera, Lukáš ; Ferus, Martin (advisor) ; Kopecký, Vladimír (referee)
The diploma thesis refers to the results of laboratory research dedicated to the influence of extraterestrial matter impacts onto a possible prebiotic synthesis on early Mars. The influence of understanding of early Mars chemistry for proceeding studies of terrestrian chemical habitus evolution and origin of life is discussed, as well as certain advantages to be made of this research within exoplanetary science. The prebiotic synthesis scenario purposed connects the hypothesis comprising a key role of extraplanetary bodies impacts played in the chemical evolution of an early Earth-like planet to the important processes studied within chemical evolution of life. Asteroid impacts are simulated by laser sparks, while post-impact thermochemical processes are mimetized in the laboratory by a controlled termolysis of certain key species in presence of meteorites, iron-rich clays and several other minerals and rocks presumably present in real impact craters. This particular field of research carried is likely to have an interesting application within current and future investigations of the Gale crater. Impactor mineraly is described on a particular example of a rare Porangaba meteorite, which, thanks to this research, has recently been addressed as a member of unique (comprising 32 bodies) family tree...
|
|
|
Mass spectrometric and optical spectroscopy studies of clusters and nanoparticles in molecular beams
Moriová, Kamila ; Fárník, Michal (advisor) ; Kopecký, Vladimír (referee)
Title: Mass spectrometric and optical spectroscopy studies of clusters and nano- particles in molecular beams Author: Kamila Moriová Institue: Institute of Physics of Charles University Supervisor: doc. Mgr. Michal Fárník, Ph.D., DSc., J. Heyrovský Institute of Phy- sical Chemistry of the CAS Consultant: RNDr. Václav Profant, Ph.D., Institute of Physics of Charles Uni- versity Abstract: Condensation of ices on the surface of dust grains with activation of these ices by cosmic rays in interstellar medium can lead to formation of complex chemical species. In the present work we primarily examine ammonia clusters, which represent a model system of ice particles. Large ammonia clusters were prepared in molecular beams and pass through a pickup chamber in which me- thanol molecules were captured. After electron ionization of (NH3)N (CH3OH)M clusters, a mass spectrum of ionic fragments was obtained in which fragments with methanol (NH3)n(CH3OH)mH+ predominated over the pure ammonia frag- ments (NH3)nH+ : at least one methanol molecule was present in 75 % fragments. This is in disproportion with the composition of the original neutral precursors, which contain less than 4 % of methanol. Based on the measurement results, we suggest that ammonia is ionized in the first place resulting in the NH+ 4 ion core solvated...
|
|
|
Study of protein structure and dynamics by means of optical spectroscopy
Pazderka, Tomáš ; Kopecký, Vladimír (advisor) ; Kaminský, Jakub (referee) ; Setnička, Vladimír (referee)
Title: Study of protein structure and dynamics by means of optical spectroscopy Author: Tomáš Pazderka Institute: Institute of Physics of Charles University Supervisor: RNDr. Vladimír Kopecký, Ph.D., Institute of Physics of Charles University Abstract: The aim of this thesis is to improve understanding of protein structure and dynamics and extend experimental setup and data processing for such stud- ies. We focus on the extension of experimental feasability of vibrational optical activity (VOA). We have demonstrated a usability of intensity calibration in the field of Raman optical activity. Advantages for measurements on multiple instru- ments and/or using different configurations have been shown. A new instrumental setup has been developed for microsampling measurements of vibrational circular dichroism spectra with a spatial resolution of 1 mm. Using this technique, spatial inhomogeneities in a sample of protein fibrils have been observed. Model com- pounds for amide nonplanarity have been investigated utilizing several methods of optical spectroscopy and key spectral features for determination of amide non- planarity and the absolute configuration have been identified. A comprehensive set of Raman spectra of proteinogenic amino acids has been measured. Sample concentration dependencies and consequent...
|
|
|
Raman spectroscopy as the tool for Alzheimer's disease diagnostics
Tesař, Adam ; Kopecký, Vladimír (advisor) ; Jelínek, Otakar (referee)
Alzheimer disease (AD) is the most frequent dementia. The prevalence is approximately 10% in 65 years old people. The current treatment is only progression protective, therefore it is crucial to find a new diagnostic approach for diagnosing AD in early stage. We analysed a set of 55 patients by the drop coating deposition Raman spectroscopy with the goal to verify previously published high sensitivity of the AD spectroscopic diagnosis in cerebral spinal fluid (CSF) and to find a new diagnostic method for blood serum (BS). We optimized measurement conditions for BS. The results were evaluated by the cluster analysis and the principal component analysis. The small set of samples exhibited high sensitivity in both CSF and BS but that distinctly decreased in the whole set. The results for CSF were affected by the choice of the analysed spectral interval. The best for AD diagnose was the interval containing peaks at 980, 1080 and 1249 cm-1.The results for BS have been the most sensitive in the whole spectral range. They have low sensitivity but high specificity for AD (92%). The usage of neural networks has conversely high sensitivity and low specificity in both sets of samples of BS and CSF. Powered by TCPDF (www.tcpdf.org)
|
guest ::
