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Biomedical subjects

M C Davies

Publications and source records attributed to M C Davies.

At least 19 recordsLinked to original sources
PubMed2000

TOF-SIMS characterization and imaging of controlled-release drug delivery systems.

Time-of-flight secondary ion mass spectrometry (TOF-SIMS) was used for the analysis of multilayer drug beads that serve as controlled-release drug delivery systems. TOF-SIMS analysis of a cross section of each bead system allowed molecular chemical information to be gained from all of the layers simultaneously, in situ. The integrity of each of the layers was evaluated through imaging of specific ion species for the drug, excipient, and coating materials. The three beads in this study each showed a unique distribution of ingredients. Images of the parent molecular ion for each drug (theophylline, paracetamol, prednisolone) showed their distribution ranged from micrometer-sized particles in one bead cross section to almost homogeneous in another bead cross section. The chemical composition of each of the layers in the beads was evaluated through mass spectrometry; the ingredients did not always match the manufacturer's specification. In addition, many common drug bead ingredients were analyzed as pure substances, providing TOF-SIMS reference spectra of these materials for the first time.

Drug Delivery Systems↗
PubMed2000

Observation of DNA-polymer condensate formation in real time at a molecular level.

Dynamic real time assembly of toroidal and rod-like DNA condensates has been visualised using atomic force microscopy. Imaging has been conducted in an aqueous environment allowing the visualisation of hydrated, pegylated-polymer DNA condensates undergoing dynamic structural movement and conformational change. A major hurdle in the field of gene delivery is cellular transfection and the subsequent transfer of condensed genetic material to the cell nucleus. An increased understanding of the process of DNA condensation will aid the development and optimisation of gene delivery vectors.

Cations↗
PubMed2000

Surface characterization of aspirin crystal planes by dynamic chemical force microscopy.

Tapping mode (TM) atomic force microscopy (AFM) has been applied in a novel fashion to characterize and distinguish the (001) and (100) surfaces of individual aspirin crystals. The surface characterization was achieved by amplitude-phase, distance (a-p,d) measurements employing gold-coated AFM probes functionalized with self-assembled monolayers (SAM). Experiments using model probes coated with -CH3 and -COOH terminated SAMs have been performed on the two aspirin crystal planes (001) and (100). Results indicate that the hydrophobic -CH3 terminated AFM probes had a greater degree of interaction with the crystal plane (001), whereas the -COOH terminated AFM probes had a larger interaction with the crystal plane (100). Interpretation of these data, based upon the chemistries of the probes, correlates with current understanding of the crystal surface chemistry derived from X-ray diffraction data and dissolution rate studies.

Anti-Inflammatory Agents, Non-Steroidal↗
PubMed2000

Atomic force microscopy of gastric mucin and chitosan mucoadhesive systems.

Atomic force microscopy has been utilized to probe, at a molecular level, the interaction between purified pig gastric mucin (PGM) and a mucoadhesive cationic polymer, chitosan (sea cure 210+), with a low degree (approx. 11%) of acetylation. Images were produced detailing the structures of both PGM and chitosan in 0.1 M acetate buffer (pH 4.5), followed by the complex of the two structures in the same buffer. PGM in 0.1 M acetate buffer revealed long linear filamentous structures, consistent with earlier electron microscopy and scanning tunnelling micoscopy studies. The chitosan molecules also adopted a linear conformation in the same buffer, although with a smaller average length and diameter. They appeared to adopt a stiff-coil conformation consistent with earlier hydrodynamic measurements. The complexes formed after mixing PGM and chitosan together revealed large aggregates. In 0.1 M ionic strength buffer they were of the order of 0.7 microm in diameter, consistent with previous electron microscopy studies. The effect of ionic strength of the buffer on the structure of the complex was also studied and, together with molecular hydrodynamic data, demonstrates that the interaction is principally electrostatic in nature.

Animals↗
PubMed2000

In-situ atomic force microscopy study of beta-amyloid fibrillization.

We report the use of atomic force microscopy to observe the initial stages of beta-amyloid fibrillization in situ. The growth of individual beta-amyloid protofibrils on a mica substrate was followed over several hours. The first in situ visualization of protofibril formation from single aggregate units of beta-amyloid is reported. The growth of these protofibrils through the subsequent addition of these aggregate units is also observed. Growth of the protofibrils is bi-directional and the outgrowth of protofibrils from a common amyloid/heterogeneous core is also observed. Elongation also occurred by the addition of protofibrils from solution. This data provides an exciting insight into the early stages of beta-amyloid fibrillization and can be used to enhance the understanding of the mechanism(s) by which beta-amyloid fibrillizes and may consequently enable inhibition of one or more stages of fibrillization as a potential therapeutic strategy.

Adsorption↗
PubMed2000

Characterization of the surfaces generated by liposome binding to the modified dextran matrix of a surface plasmon resonance sensor chip.

The dextran matrix of a surface plasmon resonance (SPR) sensor chip modified with hydrophobic residues (BIAcore sensor chip L1) provides an ideal substrate for liposome adsorption. Liposomes of different lipid compositions are captured on the sensor chips by inserting these residues into the liposome membrane, thereby generating stable lipid surfaces. To gain a more detailed understanding of these surfaces, and to prove whether the liposomes stay on the matrix as single particles or form a continuous lipid layer by liposome fusion, we have investigated these materials, using atomic force microscopy (AFM) and fluorescence microscopy. Force measurements with AFM probes functionalized with bovine serum albumin (BSA) were employed to recognize liposome adsorption. Analysis of the maximal adhesive force and adhesion energy reveals a stronger interaction between BSA and the dextran matrix compared to the lipid-covered surfaces. Images generated using BSA-coated AFM tips indicated a complete and homogeneous coverage of the surface by phospholipid. Single liposomes could not be detected even at lower lipid concentrations, indicating that the liposomes fuse and form a lipid bilayer on the dextran matrix. Experiments with fluorescently labeled liposomes concurred with the AFM studies. Surfaces incubated with liposomes loaded with TRITC-labeled dextran showed no fluorescence, indicating a complete release of the encapsulated dye. In contrast, surfaces incubated with liposomes containing a fluorescently labeled lipid showed fluorescence.

Animals↗
PubMed2000

Surface plasmon resonance analysis of dynamic biological interactions with biomaterials.

Surface plasmon resonance (SPR) is an optical technique that is widely gaining recognition as a valuable tool to investigate biological interactions. SPR offers real time in situ analysis of dynamic surface events and, thus, is capable of defining rates of adsorption and desorption for a range of surface interactions. In this review we highlight the diversity of SPR analysis. Examples of a wide range of applications of SPR are presented, concentrating on work relevant to the analysis of biomaterials. Particular emphasis is given to the use of SPR as a complimentary tool, showing the broad range of techniques that are routinely used alongside SPR analysis.

Animals↗
PubMed2000

Characterization of protein-resistant dextran monolayers.

A range of synthetic thiolated dextrans of varying molecular weights and degrees of thiol substitution have been investigated as well-defined monolayer coatings for the reduction of nonspecific protein adsorption. Atomic force microscopy and surface plasmon resonance (SPR) analysis revealed that the surface coverage of the dextran monolayers increased with an increasing degree of thiol substitution, but conversely decreased with increasing molecular weight. SPR was then employed to monitor bovine serum albumin protein adsorption to thiolated dextran monolayers from a flowing buffered solution. Whilst a significant reduction of protein adsorption to a thiolated dextran layer coated surface compared to an uncoated surface was observed, the degree of conversion of hydroxyls to thiol groups and molecular weight was shown to affect the protein-resistant performance of the dextran layer.

Animals↗
PubMed2000

Probing protein-peptide-protein molecular architecture by atomic force microscopy and surface plasmon resonance.

We demonstrate the creation of a protein multilayer which utilises the high affinity interaction between streptavidin and biotin and incorporates a peptidic spacer. Surface plasmon resonance measurements enabled us to monitor the construction of the multilayer in real time. Atomic force microscopy was utilised to determine surface functionality at each stage of the multilayer construction, allowing us to investigate the associated mechanical properties. In this context we observed an increase in biomolecular stretching on the formation of the multilayer. We demonstrate, utilising circular dichroism, that variations in the solvent can affect the secondary structure of the peptide linker and hence its mechanical properties. Trifluoroethanol titrations on the assembled system indicate that the multilayer properties are also stimuli responsive with regard to solvent conditions. These results indicate that the multilayer stretch before cleavage is increased in the presence of trifluoroethanol. This was not expected from the study of the individual linker alone, indicating the need to study the system as a whole as opposed to the isolated components.

Biosensing Techniques↗
PubMed2000

Atomic force microscopy studies of intercalation-induced changes in plasmid DNA tertiary structure.

Structural transitions in the tertiary structure of plasmid DNA have been investigated using atomic force microscopy. Changes in superhelical stress were induced by ethidium bromide intercalation, and conformational effects monitored by recording topographic images from DNA complexes of various ethidium bromide:base pair stoichiometry. Significant changes in the tertiary structure of individual DNA molecules were observed with increasing ethidium bromide concentration. The first distinct conformational transition was from a predominantly relaxed structure to one consisting solely of toroidal supercoils. A further increase in ethidium bromide concentration resulted in the formation of regions of plectonemic supercoiling. The ratio of plectonemic:toroidal supercoiling gradually increased until an extremely tightly interwound structure of solely plectonemic supercoiling was finally adopted. The toroidal form of supercoiling observed in this study is unusual as both atomic force microscopy and electron microscopy techniques have previously shown that plectonemic supercoiling is the predominant form adopted by plasmid DNA.

DNA, Superhelical↗
PubMed2000

Discrimination of polymorphic forms of a drug product by localized thermal analysis.

In chemical processing, it is important to distinguish between and identify polymorphic forms. We demonstrate the novel use of scanning thermal microscopy (SThM) and localized thermal analysis to distinguish and identify polymorphic forms of the drug cimetidine. These forms cannot be resolved by classical bulk thermal analysis. SThM reveals a sample consisting of a 50 : 50 mixture of the polymorphs contains regions of different thermal conductivity, corresponding to the different polymorphs. Localized thermal analysis of small volumes of pure polymorphic samples (approximately 50 microm3) shows that the origin of the thermal conductivity contrast lies, at least in part, with the presence of a surface water layer on the more hydrophilic polymorph.

Chemistry, Pharmaceutical↗
PubMed2000

Printing patterns of biospecifically-adsorbed protein.

The advancement of elastomeric patterning techniques in recent years has significantly enhanced our ability to spatially control biomaterial surface chemistry at the micrometre level. The application of this technology to the patterning of biomolecules onto solid surfaces has created many potential applications including the development of advanced biosensors, combinatorial library screening and the formation of tissue engineering templates. In this paper, we describe the direct patterning of protein by microcontact printing. An important consideration for the fabrication of protein micropatterns intended for these applications is the nature of the protein immobilization to a substrate. To date, the patterning of proteins by direct microcontact printing (microCP) has relied on the non-covalent adsorption to a substrate. Ideally, the proteins need to be firmly anchored onto a surface without adversely effecting their activity. Here, the high affinity avidin-biotin receptor-ligand interaction has been exploited to form arrays of avidin molecules onto a polymeric substrate expressing biotin moieties. This has created a generic technique by which any biotinylated species can be subsequently immobilized into defined patterns. Utilizing atomic force microscopy (AFM), the patterned surfaces have been characterized to molecular resolution. The micropatterned sample supported cell adhesion when biotin-(G)11-GRGDS was bound to the avidin bearing arrays.

3T3 Cells↗
PubMed1999

The influence of epitope availability on atomic-force microscope studies of antigen-antibody interactions.

The ability of the atomic-force microscope (AFM) to detect interaction forces between individual biological molecules has recently been demonstrated. In this study, force measurements have been obtained between AFM probes functionalized with the beta-subunit of human chorionic gonadotrophin (betahCG) and surfaces functionalized with anti-betahCG antibody. A comparison of the obtained results with previous anti-ferritin antibody-binding data identifies differences when the antigen molecule expresses only a single epitope (betahCG), rather than multiple epitopes (ferritin), for the monoclonal antibodies employed. Specifically, the probability of observing probe-sample adhesion is found to be higher when the antigen expresses multiple epitopes. However, the periodic force observed in the adhesive-force distribution, due to the rupture of single antigen-antibody interactions, is found to be larger and more clearly observed for the mono-epitopic system. Hence, these findings indicate the potential of the AFM to distinguish between multivalent and monovalent antibody-antigen interactions, and demonstrate the influence of the number of expressed epitopes upon such binding studies.

Adhesiveness↗
PubMed1999

Detection and determination of surface levels of poloxamer and PVA surfactant on biodegradable nanospheres using SSIMS and XPS.

The surface chemical characterisation of sub-200 nm poly(DL-lactide co-glycolide) nanospheres has been carried out using the complementary analytical techniques of static secondary ion mass spectrometry (SSIMS) and X-ray photoelectron spectroscopy (XPS). The nanospheres, which are of interest for site-specific drug delivery, were prepared using an emulsification-solvent evaporation technique with poly(vinyl alcohol), Poloxamer 407 and Poloxamine 908 respectively as stabilisers. The presence of surfactant molecules on the surface of cleaned biodegradable colloids was confirmed and identified on a qualitative molecular level (SSIMS) and from a quantitative elemental and functional group analysis (XPS) perspective. SSIMS and XPS data were also used in combination with electron microscopy to monitor the effectiveness of cleaning procedures in removing poorly bound surfactant molecules from the surface of nanospheres. The findings are discussed with respect to the development of nanoparticle delivery systems, particularly the composition of the surface for extending blood circulation times and achieving site-specific deposition.

Adsorption↗
PubMed1999

Morphological development of beta(1-40) amyloid fibrils.

The Alzheimer's disease-related peptide beta(1-40) amyloid self-associates to form fibrils exhibiting a morphology characteristic of amyloidogenic proteins. The mechanism of this fibrillization process has yet to be fully elucidated. In this study we have immobilized the beta(1-40) amyloid to flat gold surfaces using thiol-based self-assembled monolayers. Atomic force microscopy reveals the presence of spherical units of beta(1-40) amyloid immediately following the initiation of fibrillization. Short fibrillar structures, termed nascent fibrils, which appear to be formed by the association of these units are also present at this time point. At later time points extended, branching networks of fibrils are observed. Some fibrils exhibit a more beaded appearance and greater axial periodicity than others. No nascent fibrils are seen to be present. We believe that these data identify an early fibril structure which could act as an intermediate in beta-amyloid fibrillization. The oligomeric units of which these nascent fibrils are comprised are also determined.

Amino Acid Sequence↗
PubMed1999

Competitive protein adsorption as observed by surface plasmon resonance.

The competitive nature of protein adsorption has been investigated in situ by surface plasmon resonance (SPR) analysis. The adsorption from blood plasma solutions of albumin, fibrinogen and immunoglobulin-G (IgG), to a polystyrene surface was investigated as part of concentration- and time-dependent studies, to observe the sequential adsorption of the three proteins at the surface. Adsorption of plasma solutions at a range of concentrations or incubation times was performed and the resulting surfaces were probed by the addition of an appropriate antibody to the protein surface. The process was repeated for each antigen leading to a surface concentration profile of each protein with respect to plasma concentration and plasma incubation time. The SPR was able to detect changes in the relative surface concentration of each component demonstrating that the protein's residence time at the interface was dependent upon its molecular weight, bulk concentration and surface affinity. All ri,hts reserved

Adsorption↗
PubMed1999

Bone density in a population of long term oral contraceptive pill users does not differ from that in menstruating women.

Prevention of osteoporosis is a major public health issue. Amenorrhoeic women have lower bone density than normally menstruating women, which is related to the duration of amenorrhoea and the severity of oestrogen deficiency. Bone mineral density (BMD) in amenorrhoeic women can be improved by oestrogen replacement in the form of the combined oral contraceptive pill (COCP), so increased BMD might be an important non-contraceptive benefit of the COCP in menstruating women. Previous studies have been variably reported, but have used different methodologies for measurement of BMD. We measured BMD using the DEXA technique in long term COCP users and compared this with menstruating women who had never used the COCP. No differences in bone density were found, suggesting that the COCP does not improve bone mass in menstruating women who are adequately oestrogenised by their own ovaries.

Absorptiometry, Photon↗