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David Farrar

Publications and source records attributed to David Farrar.

13 recordsLinked to original sources
PubMed2006

Quantification of surface-bound proteins by fluorometric assay: Comparison with quartz crystal microbalance and amido black assay.

Protein adsorption is of major and widespread interest, being useful in the fundamental understanding of biological processes at interfaces through to the development of new materials. A number of techniques are commonly used to study protein adhesion, but few are directly quantitative. Here we describe the use of Nano Orange, a fluorometric assay, to quantitatively assess the adsorption of bovine fibrinogen and albumin onto model hydrophilic (OH terminated) and hydrophobic (CH3 terminated) surfaces. Results obtained using this method allowed the calibration of previously unquantifiable data obtained on the same surfaces using quartz crystal microbalance measurements and an amido black protein assay. Both proteins were found to adsorb with higher affinity but with lower saturation levels onto hydrophobic surfaces. All three analytical techniques showed similar trends in binding strength and relative amounts adsorbed over a range of protein concentrations, although the fluorometric analysis was the only method to give absolute quantities of surface-bound protein. The versatility of the fluorometric assay was also probed by analyzing protein adsorption onto porous superhydrophobic and superhydrophilic surfaces. Results obtained using the assay in conjunction with these surfaces were surface chemistry dependent. Imbibition of water into the superhydrophilic coatings provided greater surface area for protein adsorption, although the protein surface density was less than that found on a comparable flat hydrophilic surface. Superhydrophobic surfaces prevented protein solution penetration. This paper demonstrates the potential of a fluorometric assay to be used as an external calibration for other techniques following protein adsorption processes or as a supplemental method to study protein adsorption. Differences in protein adsorption onto hydrophilic vs superhydrophilic and hydrophobic vs superhydrophobic surfaces are highlighted.

Adsorption↗
PubMed2006

Surface tailoring for controlled protein adsorption: effect of topography at the nanometer scale and chemistry.

Protein adsorption behavior is at the heart of many of today's research fields including biotechnology and materials science. With understanding of protein-surface interactions, control over the conformation and orientation of immobilized species may ultimately allow tailor-made surfaces to be generated. In this contribution protein-surface interactions have been examined with particular focus on surface curvature with and without surface chemistry effects. Silica spheres with diameters in the range 15-165 nm with both hydrophilic and hydrophobic surface chemistries have been used as model substrates. Two proteins differing in size and shape, bovine serum albumin (BSA) and bovine fibrinogen (Fg), have been used in model studies of protein binding with detailed secondary structure analysis being performed using infrared spectroscopy (IR) on surface-bound proteins. Although trends in binding affinity and saturation values were similar for both proteins, albumin is increasingly less ordered on larger substrates, while fibrinogen, in contrast, loses secondary structure to a greater extent when adsorbing onto particles with high surface curvature. These effects are compounded by surface chemistry, with both proteins becoming more denatured on hydrophobic surfaces. Both surface chemistry and topography play key roles in determining the structure of the bound proteins. A model of the binding characteristics of these two proteins onto surfaces having differing curvature and chemistry is presented. We propose that properties of an adsorbed protein layer may be guided through careful consideration of surface structure, allowing the fabrication of materials/surface coatings with tailored bioactivity.

Adsorption↗
PubMed2006

Dynamic mechanical studies of hydrolytic degradation in isotropic and oriented Maxon B.

Hydrolytic degradation studies have been undertaken on Maxon B, a bioresorbable block copolymer of polyglycolic acid (PGA) and polytrimethylene carbonate (TMC). Isotropic and oriented samples were studied by dynamic mechanical measurements over a wide range of temperatures. In addition to mechanical tests, water content and mass loss were also determined on the degraded samples. At early stages of degradation water content was the dominant factor and plasticisation lead to reductions in the glass transition temperatures of the PGA and TMC components. Orientation was shown to give significant improvements in the mechanical properties, including overall increases in modulus and an increase in the glass transition temperature of the PGA component, which is important for the behaviour at body temperature (37 degrees C). Oriented samples also showed significantly less reduction in mechanical properties on degradation. Simple one-dimensional Takayanagi models were used to provide useful insight into the understanding of the mechanical behaviour.

Biocompatible Materials↗
PubMed2006

In vivo assessment of a rotary left ventricular assist device-induced artificial pulse in the proximal and distal aorta.

The increasing clinical use of rotary left ventricular assist devices (LVADs) suggests that chronic attenuation of arterial pulse pressure has no clinically significant detrimental effects. However, it remains possible that modulating LVAD rotor speed to produce an artificial pulse may be of temporary or occasional benefit. We sought to evaluate a pulse produced by a continuous-flow, centrifugal pump in an ovine thoracic and abdominal aorta. Both ventricles of an adult sheep were resected to eliminate all native cardiac contributions to pulsatility, each replaced by a continuous-flow Thoratec HeartMate III blood pump (Burlington, MA, USA). An LVAD-induced pulsatile flow was achieved by sharply alternating the speed of the magnetically levitated rotor of the left pump between 1,500 rpm (artificial diastole) and 5,500 rpm (artificial systole) at a rate of 60 bpm at a "systolic" interval of 30%. A catheter was advanced from the ascending aorta to the iliac bifurcation via the ventricular assist device outflow graft for pressure measurement and data acquisition. The mean LVAD-induced pulse pressures were 34, 29, 27, and 26 mm Hg in the ascending, thoracic, and abdominal aorta, and the iliac bifurcation, respectively. The maximum rate of pressure rise (deltap/deltat) was between 189 and 238 mm Hg/s, approaching that of the native pulse, although the energy equivalent pressure did not exceed the mean arterial pressure. The HeartMate III's relatively stiff speed control, low rotor mass, and robust magnetic rotor suspension result in a responsive system, enabling very rapid speed changes that can be used to simulate physiologic pulse pressure and deltap/deltat.

Animals↗
PubMed2005

Short and medium chain length chlorinated paraffins in UK human milk fat.

Chlorinated paraffins (also called polychlorinated n-alkanes -- PCAs) are a class of industrial chemicals comprising chlorinated straight chain hydrocarbons. They have a wide range of applications and are now found in a range of environmental compartments. We analysed a total of 25 human milk-fat samples, donated by 18 individuals from the urban London and more rural Lancaster areas in the UK, for short chain PCAs (C(10)-C(13) sPCAs) and medium chain PCAs (C(14)-C(17) mPCAs), using gas chromatography-ECNI high-resolution mass spectrometry. Our study confirms that trace quantities of PCAs can reach human milk-fat. sPCAs were detected in all but four samples, while mPCAs were detected in all samples. The median sPCA concentration was 180 ng/g fat (range of 49 to 820 ng/g fat -- detected values only) and the median mPCA concentration was 21 ng/g fat (range of 6.2 to 320 ng/g fat). No differences were noted in ranges of observed values for either sPCAs or mPCAs between samples from London and Lancaster. Most samples also exhibited similar patterns of sPCAs and mPCAs. One sample exhibited a different pattern for sPCAs and mPCAs, an observation that may be related to differences in exposure or biological factors for this individual.

Fats↗
PubMed2005

Interpretation of protein adsorption: surface-induced conformational changes.

Protein adhesion plays a major role in determining the biocompatibility of materials. The first stage of implant integration is the adhesion of protein followed by cell attachment. Surface modification of implants (surface chemistry and topography) to induce and control protein and cell adhesion is currently of great interest. This communication presents data on protein adsorption (bovine serum albumin and fibrinogen) onto model hydrophobic (CH(3)) and hydrophilic (OH) surfaces, investigated using a quartz crystal microbalance (QCM) and grazing angle infrared spectroscopy. Our data suggest that albumin undergoes adsorption via a single step whereas fibrinogen adsorption is a more complex, multistage process. Albumin has a stronger affinity toward the CH(3) compared to OH terminated surface. In contrast, fibrinogen adheres more rapidly to both surfaces, having a slightly higher affinity toward the hydrophobic surface. Conformational assessment of the adsorbed proteins by grazing angle infrared spectroscopy (GA-FTIR) shows that after an initial 1 h incubation few further time-dependent changes are observed. Both proteins exhibited a less organized secondary structure upon adsorption onto a hydrophobic surface than onto a hydrophilic surface, with the effect observed greatest for albumin. This study demonstrates the ability of simple tailor-made monochemical surfaces to influence binding rates and conformation of bound proteins through protein-surface interactions. Current interest in biocompatible materials has focused on surface modifications to induce rapid healing, both of implants and for wound care products. This effect may also be of significance at the next stage of implant integration, as cell adhesion occurs through the surface protein layer.

Adsorption↗
PubMed2005

A degradation study of PLLA containing lauric acid.

Addition of lauric acid to poly (L-lactide) (PLLA) has resulted in a new family of enhanced degradation biomaterials. Presented is PLLA4.5 (PLLA containing 4.5 wt% lauric acid), the fastest degrading of the family. Degradation was studied via mass changes, gel-permeation chromatography, wide- and small-angle X-ray scattering (WAXS and SAXS), simultaneous SAXS and tensile testing, and visual observation. The undegraded PLLA4.5 deformed by crazing, recognisable from the characteristic shape of the SAXS pattern. As water up-take and degradation proceeded, samples crystallised, decreasing the SAXS long period, until by 4 days the deformation mechanism had become that of crystal-mediated deformation. This resulted in a 'peanut-lemon'-shaped SAXS pattern, interpreted in terms of cavitation and fibrillated shear. Further degradation up to 12 days resulted in the same deformation mechanism at different sample displacements, with samples failing earlier during tensile testing until a ductile-brittle transition occurred. At 30-40 days water up-take and mass-loss increased significantly and global whitening of samples occurred, while the crystallinity and long period stabilised. Complete degradation had not occurred by the end of the study at 73 days. Through an understanding of how the changes in morphology during degradation affect the micromechanisms of deformation, it may be possible to design microstructures to give a tailored evolution of mechanical response in the body.

Absorbable Implants↗
PubMed2004

Computational fluid dynamics analysis of a maglev centrifugal left ventricular assist device.

The fluid dynamics of the Thoratec HeartMate III (Thoratec Corp., Pleasanton, CA, U.S.A.) left ventricular assist device are analyzed over a range of physiological operating conditions. The HeartMate III is a centrifugal flow pump with a magnetically suspended rotor. The complete pump was analyzed using computational fluid dynamics (CFD) analysis and experimental particle imaging flow visualization (PIFV). A comparison of CFD predictions to experimental imaging shows good agreement. Both CFD and experimental PIFV confirmed well-behaved flow fields in the main components of the HeartMate III pump: inlet, volute, and outlet. The HeartMate III is shown to exhibit clean flow features and good surface washing across its entire operating range.

Assisted Circulation↗
PubMed2003

Assessing the health risks following environmental exposure to hexachlorobutadiene.

Hexachloro-1,3-butadiene (HCBD) has been reported to be toxic to the rat kidney in a 2 year study at doses higher than 0.2 mg/kg/day. The toxicity is known to be a consequence of the metabolism of HCBD by glutathione conjugation and the renal beta-lyase pathway. Neither toxicity data, nor data on the metabolism of HCBD, are available in humans. In the current work, the potential of HCBD to cause kidney damage in humans environmentally exposed to this chemical has been assessed quantitatively by comparing the key metabolic steps in rats and humans. To that end, the hepatic conjugation of HCBD with glutathione, the metabolism of the cysteine conjugate by renal beta-lyases and N-acetyltransferases, and the metabolism of the N-acetylcysteine conjugate by renal acylases has been compared in vitro in rat and human tissues. Rates for each metabolic step were lower in humans than in rats; 5-fold for glutathione conjugation, 3-fold for beta-lyase and 3.5-fold for N-acetyltransferase. Acylase activity could not be detected in human kidney cytosol. Use of these data in a physiologically based toxicokinetic model to quantify metabolism by the beta-lyase pathway demonstrated that metabolism in humans was an order of magnitude lower than that in rats. At the no effect level for kidney toxicity in the rat the concentration of beta-lyase metabolites was calculated by the model to be 137.7 mg/l. In humans the same concentration would be achieved following exposure to 1.41 ppm HCBD. This is in contrast to the figure of 0.6 ppb which is obtained when it is assumed that the risk is associated with the internal dose of HCBD itself rather than beta-lyase metabolites.

Adult↗
PubMed2002

Alternative methods for the median lethal dose (LD(50)) test: the up-and-down procedure for acute oral toxicity.

The authors have developed an improved version of the up-and-down procedure (UDP) as one of the replacements for the traditional acute oral toxicity test formerly used by the Organisation for Economic Co-operation and Development member nations to characterize industrial chemicals, pesticides, and their mixtures. This method improves the performance of acute testing for applications that use the median lethal dose (classic LD50) test while achieving significant reductions in animal use. It uses sequential dosing, together with sophisticated computer-assisted computational methods during the execution and calculation phases of the test. Staircase design, a form of sequential test design, can be applied to acute toxicity testing with its binary experimental endpoints (yes/no outcomes). The improved UDP provides a point estimate of the LD50 and approximate confidence intervals in addition to observed toxic signs for the substance tested. It does not provide information about the dose-response curve. Computer simulation was used to test performance of the UDP without the need for additional laboratory validation.

Animal Testing Alternatives↗
PubMed2002

Toxicity of ammonium perfluorooctanoate in male cynomolgus monkeys after oral dosing for 6 months.

Ammonium perfluorooctanoate (APFO) is a processing aid in the production of fluoropolymers that has been shown to have a long half-life in human blood. To understand the potential toxicological response of primates, groups of male cynomolgus monkeys were given daily po (capsule) doses of either 0, 3, 10, or 30 (reduced to 20) mg/kg/day for 26 weeks. Two monkeys from each of the control and 10 mg/kg/day dose groups were observed for 90 days after the last dose. Clinical observations, clinical chemistry, determination of key hormones, gross and microscopic pathology, cell proliferation, peroxisomal proliferation, bile-acid determination, and serum and liver perfluorooctanoate (PFOA) concentrations were monitored. Toxicity, including weight loss and reduced food consumption, was noted early in the study at the 30 mg/kg/day dose; therefore, the dose was reduced to 20 mg/kg/day. The same signs of toxicity developed in 3 monkeys at 20 mg/kg/day, after which treatment of these monkeys was discontinued. One 30/20 mg/kg/day monkey developed the signs of toxicity noted above and a possible dosing injury, and this monkey was sacrificed in extremis on Day 29. A 3 mg/kg/day dose-group monkey was sacrificed in extremis on Day 137 for reasons not clearly related to APFO treatment. Dose-dependent increases in liver weight as a result of mitochondrial proliferation occurred in all APFO-treated groups. Histopathologic evidence of liver injury was not observed at either 3 or 10 mg/kg/day. Evidence of liver damage was seen in the monkey sacrificed in moribund condition at the highest dose. Body weights were decreased at 30/20 mg/kg. PFOA concentrations in serum and liver were highly variable, were not linearly proportional to dose, and cleared to background levels within 90 days after the last dose. A no observable effect level was not established in this study, and the low dose of 3 mg/kg/day was considered the lowest observable effect level based on increased liver weight and uncertainty as to the etiology leading to the moribund sacrifice of one low-dose monkey on Day 137. Other than those noted above, there were no APFO-related macroscopic or microscopic changes, changes in clinical chemistry, hormones, or urinalysis, or hematological effects. In particular, effects that have been associated with the development of pancreatic and testicular toxicity in rats were not observed in this study.

Acyl Coenzyme A↗
PubMedDate not supplied

Biostability of Thoralon left ventricular assist device blood pumping sacs after long-term clinical use.

The purpose of this paper is to investigate the biostability of Thoralon polyurethane blood pumping sacs that have been used for up to 14 months in the Thoratec ventricular assist device (VAD) in human subjects. Thoralon is a proprietary polyetherurethaneurea (PEUU) blended with siloxane based surface modifying additive (SMA). Samples of Thoralon blood sacs that were used in Thoratec LVAD pumps for 53, 171, 336, and 434 days were taken from three distinct regions identified as high flex, medium flex, and low flex for physicochemical analysis. Explanted samples exhibited a slight but not significant increase in physical properties. The molecular weight of the blood sacs had also increased significantly in all blood sacs regardless of duration of use. SEM images did not indicate any cracking, pitting, or surface erosion; however, in a small area of the high flex region of the 336 day sac some biofilm was seen. The Attenuated Total Reflectance Fourier Transform Infra Red Spectroscopy (ATR-FTIR) analysis showed no significant reduction of surface ether content. The surface chemistry analyzed using Electron Spectroscopy for Chemical Analysis (ESCA) also indicated no significant change compared with control. These results confirm the biostability of Thoralon for long-term clinical applications in the Thoratec VAD.

Biocompatible Materials↗