Plasma processing of PDMS based spinal implants for covalent protein immobilization, cell attachment and spreading.

PDMS is widely used for prosthetic device manufacture. Conventional ion implantation is not a suitable treatment to enhance the biocompatibility of poly dimethyl siloxane (PDMS) due to its propensity to generate a brittle silicon oxide surface layer which cracks and delaminates. To overcome this limitation, we have developed new plasma based processes to balance the etching of carbon with implantation of carbon from the plasma source. When this carbon was implanted from the plasma phase it resulted in a surface that was structurally similar and intermixed with the underlying PDMS material and not susceptible to delamination. The enrichment in surface carbon allowed the formation of carbon based radicals that are not present in conventional plasma ion immersion implantation (PIII) treated PDMS. This imparts the PDMS surfaces with covalent protein binding capacity that is not observed on PIII treated PDMS. The change in surface energy preserved the function of bound biomolecules and enhanced the attachment of MG63 osteosarcoma cells compared to the native surface. The attached cells, an osteoblast interaction model, showed increased spreading on the treated over untreated surfaces. The carbon-dependency for these beneficial covalent protein and cell linkage properties was tested by incorporating carbon from a different source. To this end, a second surface was produced where carbon etching was balanced against implantation from a thin carbon-based polymer coating. This had similar protein and cell-binding properties to the surfaces generated with carbon inclusion in the plasma phase, thus highlighting the importance of balancing carbon etching and deposition. Additionally, the two effects of protein linkage and bioactivity could be combined where the cell response was further enhanced by covalently tethering a biomolecule coating, as exemplified here with the cell adhesive protein tropoelastin. Providing a balanced carbon source in the plasma phase is applicable to prosthetic device fabrication as illustrated using a 3-dimensional PDMS balloon prosthesis for spinal implant applications. Consequently, this study lays the groundwork for effective treatments of PDMS to selectively recruit cells to implantable PDMS fabricated biodevices.

Science of Water Leaks: Validated Theory for Moisture Flow in Microchannels and Nanochannels.

Water is ubiquitous; the science of its transport in micro- and nanochannels has applications in electronics, medicine, filtration, packaging, and earth and planetary science. Validated theory for water vapor and two-phase water flows is a "missing link"; completing it enables us to define and quantify flow in a set of four standard leak configurations with dimensions from the nanoscale to the microscale. Here we report the first measurements of water vapor flow rates through four silica microchannels as a function of humidity, including under conditions when air is present as a background gas. An important finding is that the tangential momentum accommodation coefficient (TMAC) is strongly modified by surface layers of adsorbed water molecules, in agreement with previous work on the TMAC for nitrogen molecules impacting a silica surface in the presence of moisture. We measure enhanced flow rates for two-phase flows in silica microchannels driven by capillary filling. For the measurement of flows in nanochannels we use heavy water mass spectrometry. We construct the theory for the flow rates of the dominant modes of water transport through each of the four standard configurations and benchmark it against our new measurements in silica and against previously reported measurements for nanochannels in carbon nanotubes, carbon nanopipes, and porous alumina. The findings show that all behavior can be described by the four standard leak configurations and that measurements of leak behavior made using other molecules, such as helium, are not reliable. Single-phase water vapor flow is overestimated by a helium measurement, while two-phase flows are greatly underestimated for channels larger than 100 nm or for all channels when boundary slip applies, to an extent that depends on the slip length for the liquid-phase flows.

Ball milling: a green mechanochemical approach for synthesis of nitrogen doped carbon nanoparticles.

Technological and scientific challenges coupled with environmental considerations have attracted a search for robust, green and energy-efficient synthesis and processing routes for advanced functional nanomaterials. In this article, we demonstrate a high-energy ball milling technique for large-scale synthesis of nitrogen doped carbon nanoparticles, which can be used as an electro-catalyst for oxygen reduction reactions after a structural refinement with controlled thermal annealing. The resulting carbon nanoparticles exhibited competitive catalytic activity (5.2 mA cm(-2) kinetic-limiting current density compared with 7.6 mA cm(-2) on Pt/C reference) and excellent methanol tolerance compared to a commercial Pt/C catalyst. The proposed synthesis route by ball milling and annealing is an effective process for carbon nanoparticle production and efficient nitrogen doping, providing a large-scale production method for the development of highly efficient and practical electrocatalysts.

An integrated solution for rapid biosensing with robust linker free covalent binding surfaces.

A novel integrated biosensor methodology is proposed and demonstrated. The methodology utilizes a nitrogen-containing plasma polymer to achieve linker-free binding of a layer of biorecognition molecules. The sensor surface has been shown to maintain its performance after freeze-drying providing a long shelf life under ambient conditions. The sensor is configured for single wavelength ellipsometric detection providing a low cost, versatile, and rapid sensing and diagnosis platform suitable for a wide range of applications and end-users. The merits of the methodology are demonstrated using three antigen-antibody pairs.

In vivo biocompatibility of a plasma-activated, coronary stent coating.

Bare metal and drug-eluting coronary stents suffer an inherent lack of vascular cell and blood compatibility resulting in adverse patient responses. We have developed a plasma-activated coating (PAC) for metallic coronary stents that is durable, withstands crimping and expansion, has low thrombogenicity and can covalently bind proteins, linker-free. This has been shown to enhance endothelial cell interactions in vitro and has the potential to promote biointegration of stents. Using the rabbit denuded iliac artery model, we show for the first time that PAC is a feasible coating for coronary stents in vivo. The coating integrity of PAC was maintained following implantation and expansion. The rate of endothelialization, strut coverage, neointimal response and the initial immune response were equivalent to bare metal stents. Furthermore, the initial thrombogenicity caused by the PAC stents showed a reduced trend compared to bare metal stents. This work demonstrates a robust, durable, non-cytotoxic plasma-based coating technology that has the ability to covalently immobilize bioactive molecules for surface modification of coronary stents. Improvements in the clinical performance of implantable cardiovascular devices could be achieved by the immobilization of proteins or peptides that trigger desirable cellular responses.

Facile preparation of free-standing carbon nanotube arrays produced using two-step floating-ferrocene chemical vapor deposition.

A two-step floating-ferrocene chemical vapor deposition method has been devised for the preparation of single-layered aligned carbon nanotube (CNT) arrays. In the first step, uniform Fe catalysts are in situ produced and coated on a Si substrate from ferrocene; single-layered CNT arrays are prepared on these catalysts from ethylene in the second step. The effect of ferrocene loading on the distribution of Fe catalysts, as well as the morphology, diameter, and height of the CNT arrays, was investigated. A novel vacuum extraction process was employed to release the as-prepared CNT array from the Si wafer after water etching at 750 °C. The structural integrity of the free-standing arrays was preserved after the detachment process. The interface between the substrate and the as-grown CNT array was examined. The Fe catalyst distribution on the Si substrate remained homogeneous when the CNT array was removed, and the tops and bottoms of the arrays had different structures, suggesting that the arrays were formed predominantly by a base-growth mode. These free-standing arrays could potentially be applied in membrane or electronic applications.

Free radical functionalization of surfaces to prevent adverse responses to biomedical devices.

Immobilizing a protein, that is fully compatible with the patient, on the surface of a biomedical device should make it possible to avoid adverse responses such as inflammation, rejection, or excessive fibrosis. A surface that strongly binds and does not denature the compatible protein is required. Hydrophilic surfaces do not induce denaturation of immobilized protein but exhibit a low binding affinity for protein. Here, we describe an energetic ion-assisted plasma process that can make any surface hydrophilic and at the same time enable it to covalently immobilize functional biological molecules. We show that the modification creates free radicals that migrate to the surface from a reservoir beneath. When they reach the surface, the radicals form covalent bonds with biomolecules. The kinetics and number densities of protein molecules in solution and free radicals in the reservoir control the time required to form a full protein monolayer that is covalently bound. The shelf life of the covalent binding capability is governed by the initial density of free radicals and the depth of the reservoir. We show that the high reactivity of the radicals renders the binding universal across all biological macromolecules. Because the free radical reservoir can be created on any solid material, this approach can be used in medical applications ranging from cardiovascular stents to heart-lung machines.

Stability of a therapeutic layer of immobilized recombinant human tropoelastin on a plasma-activated coated surface.

PURPOSE: To modify blood-contacting stainless surfaces by covalently coating them with a serum-protease resistant form of tropoelastin (TE). To demonstrate that the modified TE retains an exposed, cell-adhesive C-terminus that persists in the presence of blood plasma proteases. METHODS: Recombinant human TE and a point mutant variant (R515A) of TE were labeled with (125)Iodine and immobilized on plasma-activated stainless steel (PAC) surfaces. Covalent attachment was confirmed using rigorous detergent washing. As kallikrein and thrombin dominate the serum degradation of tropoelastin, supraphysiological levels of these proteases were incubated with covalently bound TE and R515A, then assayed for protein levels by radioactivity detection. Persistence of the C-terminus was assessed by ELISA. RESULTS: TE was significantly retained covalently on PAC surfaces at 88 ± 5% and 71 ± 5% after treatment with kallikrein and thrombin, respectively. Retention of R515A was 100 ± 1.3% and 87 ± 2.3% after treatment with kallikrein and thrombin, respectively, representing significant improvements over TE. The functionally important C-terminus was cleaved in wild-type TE but retained by R515A. CONCLUSIONS: Protein persists in the presence of human kallikrein and thrombin when covalently immobilized on metal substrata. R515A displays enhanced protease resistance and retains the C-terminus presenting a protein interface that is viable for blood-contacting applications.

The immobilization of recombinant human tropoelastin on metals using a plasma-activated coating to improve the biocompatibility of coronary stents.

Current endovascular stents have sub-optimal biocompatibility reducing their clinical efficacy. We previously demonstrated a plasma-activated coating (PAC) that covalently bound recombinant human tropoelastin (TE), a major regulator of vascular cells in vivo, to enhance endothelial cell interactions. We sought to develop this coating to enhance its mechanical properties and hemocompatibility for application onto coronary stents. The plasma vapor composition was altered by incorporating argon, nitrogen, hydrogen or oxygen to modulate coating properties. Coatings were characterized for 1) surface properties, 2) mechanical durability, 3) covalent protein binding, 4) endothelial cell interactions and 5) thrombogenicity. The N(2)/Ar PAC had optimal mechanical properties and did not delaminate after stent expansion. The N(2)/Ar PAC was mildly hydrophilic and covalently bound the highest proportion of TE, which enhanced endothelial cell proliferation. Acute thrombogenicity was assessed in a modified Chandler loop using human blood. Strikingly, the N(2)/Ar PAC alone reduced thrombus weight by ten-fold compared to 316L SS, a finding unaltered with immobilized TE. Serum soluble P-selectin was reduced on N(2)/Ar PAC and N(2)/Ar PAC + TE (p < 0.05), consistent with reduced platelet activation. We have demonstrated a coating for metal alloys with multifaceted biocompatibility that resists delamination and is non-thrombogenic, with implications for improving coronary stent efficacy.

A prototype scintillation dosimeter customized for small and dynamic megavoltage radiation fields.

A prototype plastic scintillation dosimeter has been developed with a small sensitive volume, rapid response and good dosimetric performance. The novelty of this design is the use of an air core light guide to transport the scintillation signal out of the primary radiation field. The significance of this innovation is that it eliminates the Cerenkov background signal that is generated in conventional optical fibres. The dosimeter performance was compared to existing commercial dosimeters in 6 MV and 18 MV photon beams and 6 MeV and 20 MeV electron beams, in both static and dynamic fields. The dosimeter was tested in small static fields and in dynamically delivered fields where the detector volume is shielded, while the stem is irradiated. The depth dose measurements for the photon beams agreed with ionization chamber measurements to within 1.6%, except in the build-up region due to positional uncertainty. For the 6 MeV and 20 MeV electron beams, the percentage depth dose measurements agreed with the ionization chamber measurements to within 3.6% and 4.5%, respectively. For field sizes of 1 cm x 1 cm and greater, the air core dosimeter readings agreed with diamond detector readings to within 1.2%. The air core dosimeter was accurate in dynamically delivered fields and had no measurable stem effect. The air core dosimeter was accurate over a range of field sizes, energies and dose rates, confirming that it is a sensitive and accurate dosimeter with high spatial resolution suitable for use in megavoltage photon and electron beams.

Cerenkov light spectrum in an optical fiber exposed to a photon or electron radiation therapy beam.

A Cerenkov signal is generated when energetic charged particles enter the core of an optical fiber. The Cerenkov intensity can be large enough to interfere with signals transmitted through the fiber. We determine the spectrum of the Cerenkov background signal generated in a poly(methyl methacrylate) optical fiber exposed to photon and electron therapeutic beams from a linear accelerator. This spectral measurement is relevant to discrimination of the signal from the background, as in scintillation dosimetry using optical fiber readouts. We find that the spectrum is approximated by the theoretical curve after correction for the wavelength dependent attenuation of the fiber. The spectrum does not depend significantly on the angle between the radiation beam and the axis of the fiber optic but is dependent on the depth in water at which the fiber is exposed to the beam.

Covalent immobilisation of tropoelastin on a plasma deposited interface for enhancement of endothelialisation on metal surfaces.

Currently available endovascular metallic implants such as stents exhibit suboptimal biocompatibility in that they re-endothelialise poorly leaving them susceptible to thrombosis. To improve the interaction of these implants with endothelial cells we developed a surface coating technology, enabling the covalent attachment of biomolecules to previously inert metal surfaces. Using horseradish peroxidase as a probe, we demonstrate that the polymerised surface can retain the presentation and activity of an immobilised protein. We further demonstrated the attachment of tropoelastin, an extracellular matrix protein critical to the correct arrangement and function of vasculature. Not only it is structurally important, but it plays a major role in supporting endothelial cell growth, while modulating smooth muscle cell infiltration. Tropoelastin was shown to bind to the surface in a covalent monolayer, supplemented with additional physisorbed multilayers on extended incubation. The physisorbed tropoelastin layers can be washed away in buffer or SDS while the first layer of tropoelastin remains tightly bound. The plasma coated stainless steel surface with immobilised tropoelastin was subsequently found to have improved biocompatibility by promoting endothelial cell attachment and proliferation relative to uncoated stainless steel controls. Tropoelastin coatings applied to otherwise inert substrates using this technology could thus have broad applications to a range of non-polymeric vascular devices.

Novel sphene coatings on Ti-6Al-4V for orthopedic implants using sol-gel method.

Hydroxyapatite (HAp) is commonly used to coat titanium alloys (Ti-6Al-4V) for orthopedic implants. However, their poor adhesion strength and insufficient long-term stability limit their application. Novel sphene (CaTiSiO5) ceramics possess excellent chemical stability and cytocompatibility. The aim of this study is to use the novel sphene ceramics as coatings for Ti-6Al-4V. The sol-gel method was used to produce the coatings and the thermal properties, phase composition, microstructure, thickness, surface roughness and adhesion strength of sphene coatings were analyzed by differential thermal analysis-thermal gravity (DTA-TG), X-ray diffraction (XRD), scanning electron microscopy (SEM), atom force microscopy (AFM) and scratch test, respectively. DTA analysis confirmed that the temperature of the sphene phase formation is 875 degrees C and XRD analysis indicated pure sphene coatings were obtained. A uniform structure of the sphene coating was found across the Ti-6Al-4V surface, with a thickness and surface roughness of the coating of about 0.5-1 microm and 0.38 microm, respectively. Sphene-coated Ti-6Al-4V possessed a significantly improved adhesion strength compared to that for HAp coating and their chemical stability was evaluated by testing the profile element distribution and the dissolution kinetics of calcium (Ca) ions after soaking the sphene-coated Ti-6Al-4V in Tris-HCl solution. Sphene coatings had a significantly improved chemical stability compared to the HAp coatings. A layer of apatite formed on the sphene-coated Ti-6Al-4V after they were soaked in simulated body fluids (SBF). Our results indicate that sol-gel coating of novel sphene onto Ti-6Al-4V possessed improved adhesion strength and chemical stability, compared to HAp-coated Ti-6Al-4V prepared under the same conditions, suggesting their potential application as coatings for orthopedic implants.