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No description available.

No description available.

The studies presented in this thesis focus on the synthesis of air-stable solid supported lipid bilayers by anhydrobiotic mechanisms. Supported lipid bilayers (SLBs) serve as platforms that mimic cellular membrane surfaces in appearance and behavior. One of the most attractive aspects of the SLB is that it exhibits two-dimensional fluidity that allows for individual components to rearrange as they would in actual cellular membranes. The one thing that would allow the SLB to become an ideal biosensor is the ability to remain stable in the absence of bulk water. As it stands now, unprotected SLBs are unstable in the presence of air causing the membrane to rearrange and delaminate from the surface. Several biological organisms utilize the process of anhydrobiosis to persevere in severe dehydrated states. Anhydrobiosis occurs when organisms employ large amounts of sugars, particularly disaccharides, to protect their cell membranes. The sugars, often released as a stress response, protect the membrane by replacing the water around the lipid headgroups while also interacting with other sugars to form a glass atop the bilayer. One of the most successful anhydrobiotic sugars has been trehalose, although other sugars have been evaluated and are capable of protecting lipid bilayers minimally. The experimental section of this thesis involves the creation of SLBs that are examined with and without the presence of sugar molecules. Essentially, the SLB was created, exposed to sugar solutions, dried, and subsequently rehydrated. Successful experiments occurred when rehydrated bilayers exhibited little damage and were mobile and functional. In addition to trehalose, several other mono- and disaccharides were used as were glycolipids, lipids with sugar headgroups. Upon the completion of all experiments it was clear that trehalose afforded the most protection of all species tested and that glycolipids do not sufficiently protect the membrane during rehydration. Therefore, the addition of a sugar such as trehalose to an SLB could allow for the creation of an air-stable biosensor that would be both practical and require little maintenance.

This dissertation focuses on the applications of solid supported phospholipid membranes as mimics of the cellular membrane using lab-on-a-chip devices in order to study biochemical events such as ligand-receptor binding and the chemical mechanism for the preservation of the biomembrane. Supported lipid bilayers (SLBs) mimic the native membrane by presenting the important property of two-dimensional lateral fluidity of the individual lipid molecules within the membrane. This is the same property that allows for the reorganization of native membrane components and facilitates multivalent ligand-receptor interactions akin to immune response, cell signaling, pathogen attack and other biochemical processes. The study is divided into two main facets. The first deals with developing a novel lipopolymer supported membrane biochip consisting of Poly(ethylene glycol) (PEG)-lipopolymer incorporated membranes. The formation and characterization of the lipopolymer membranes was investigated in terms of the polymer size, concentration and molecular conformation. The lateral diffusion of the PEG-bilayers was similar to the control bilayers. The air-stability conferred to SLBs was determined to be more effective when the PEG polymer was at, or above, the onset of the mushroom-to-brush transition. The system is able to function even after dehydration for 24 hours. Ligandreceptor binding was analyzed as a function of PEG density. The PEG-lipopolymer acts as a size exclusion barrier for protein analytes in which the binding of streptavidin was unaffected whereas the binding of the much larger IgG and IgM were either partially or completely inhibited in the presence of PEG. The second area of this study presents a molecular mechanism for in vivo biopreservation by employing solid supported membranes as a model system. The molecular mechanism of how a variety of organisms are preserved during stresses such as anhydrobiosis or cryogenic conditions was investigated. We investigated the interaction of two disaccharides, trehalose and maltose with the SLBs. Trehalose was found to be the most effective in preserving the membrane, whereas maltose exhibited limited protection. Trehalose lowers the lipid phase transition temperature and spectroscopic evidence shows the intercalation of trehalose within the membrane provides the chemical and morphological stability under a stress environment.

Isotermas de adsorção de anfifílicos sintéticos formadores de bicamadas ou fosfolipídios a partir de vesículas sobre partículas de sílica hidrofílica (AEROSIL OX-50) foram obtidas em diversas condições experimentais. Vesículas pequenas de fosfatidilcolina (PC), dipahnitoilfosfatidilcolina (DPPC), dihexadecilfosfato (DHP) e brometo de dioctadecildimetilamônio (DODAB) dispersas em 10 mM de dois diferentes tampões (Tris ou HEPES) em 3 diferentes valores de pH apresentam afinidades pela sílica que seguem a seqüência: DODAB>DPPC>PC>DHP. Deposição de bicamada de fosfolipídio é favorecida pela presença de Tris como tampão em pH ≤ 7,4, e temperaturas acima da temperatura de transição de fase para bicamada de fosfolipídio. Interação a 65 ºC por 1 hora entre vesículas de DPPC e sílica eficientemente gera a deposição de bicamada com uma adsorção máxima, se Tris é o tampão utilizado. Consistentemente, a molhabilidade de superfícies planares de SiO2 em condições de deposição de bicamada (como descrita pelas isotermas de adsorção) na presença de vesículas lipídicas mostra um grande aumento da hidrofobicidade da superfície para vesículas de DPPC a 65 ºC e DODAB em temperatura ambiente. Redução na absorção de luz pela MC540 em 565 nm foi usada como indicativa de deposição de bicamada sobre partículas de sílica. Durante a interação com partículas sólidas, a absorbância a 565 nm mostra uma diminuição que corresponde a porcentagem de marcador que se intercala entre a bicamada e a partícula sólida e, conseqüentemente, se esconde da luz incidente. Para DPPC, à 65 ºC, e DODAB à temperatura ambiente, mas não para PC, a ocorrência de deposição de bicamada, descrita pelas isotermas de adsorção, é confirmada pelas três técnicas empregadas para detecção. / Adsorption isotherms of bilayer-forming synthetic amphiphiles or phospholipids from vesicles onto hydrophilic silica particles (AEROSIL 0X-50) are obtained over a range of experimental conditions. Phosphatidylcholine (PC), dipalmitoylphosphatidylcholine (DPPC), and dioctadecyldimethylammonium bromide (DODAB) dispersed in 10 mM of two different buffers (Tris or HEPES) at 3 different pH values as small unilamellar vesicles present affinities for silica following the sequence: DODAB > DPPC > PC. Deposition of phospholipid bilayers was favoured by the presence of Tris as buffer, pH≤ 7.4, and temperatures above the phase transition temperature (Tc) for the phospholipid bilayer. Interaction at 65 ºC for 1 h between DPPC vesicles and silica efficiently leads to bilayer deposition at maximal adsorption, if Tris is the buffer used. Consistently, wettability of SiO2 planar surfaces precisely under conditions of bilayer deposition (as depicted from the isotherms) in the presence of the lipidic vesicles yielded a large increase on surface hydrophobicity for DPPC at 65 ºC and DODAB at room temperature. Reduction of merocyanine 540 absorbance at 565 nm was used as a marker for bilayer deposition onto the silica particles. Upon interaction with the solid particle, absorbance at 565 nm displays a decrease with rime that corresponds to the percentage of dye that became sandwiched between the bilayer and the solid particle surface and thereby hidden from the incident light. For DPPC at 65 ºC and DODAB at room temperature, but not for PC, occurrence of bilayer deposition depicted from the adsorption isotherms is confirmed from the three techniques employed for detection.

Diss. (sammanfattning) Uppsala : Uppsala universitet, 2009. / Härtill 3 uppsatser.

The cell membrane is a highly complex structure consisting of a large diversity of phospholipids and macromolecules. There exist a variety of diseases that compromise the integrity of this key component of the cell. This thesis considers the investigation of interactions between β-amyloid peptide (Aβ₄₂) and lipid bilayers. To facilitate understanding of this complex system, it is advantageous to employ a model sample; supported lipid bilayers (SLB) and giant multilamellar vesicles (MLVs) are used as proxy cell membranes. These nanostructures are widely used as models of cellular membranes in many areas of scientific research. Phospholipid molecules self-organise into bilayer structures containing phase-separated microdomains, which are believed to be important in many biological processes. This study aims to develop model systems and experimental tools to explore hypothetical mechanisms through which the β-amyloid interacts with the lipid membranes. A lack of mechanistic understanding is the major challenge to our efforts to elucidate not only the interactions of the Aβ42 with the lipid membranes, but also the behaviour of these systems towards the changes of the environmental conditions (pH, concentration, temperature). Our results suggest that there are various different methods, such as AFM, CARS microscopy and Raman spectroscopy as well as neutron scattering that are capable of fast imaging. Overall, all these techniques contributed in a complementary study of Aβ₄₂ aggregation states under extreme and physiological conditions as well as to image Aβ₄₂ interactions with lipid bilayers consisted of specific lipids.

Diss. (sammanfattning) Uppsala : Uppsala universitet, 2009. / Härtill 5 uppsatser.

Thesis (Ph. D.)--University of Virginia, 2000. / Spine title: Phospholipase A₂ binding. Includes bibliographical references (p. 245-258). Also available online through Digital Dissertations.