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Bioactive Surfaces

The INP has many years of experience in the development of plasma-supported processes for refining product surfaces in the life sciences sector. The application spectrum of such surfaces covers the complete range of vascular replacement and load-bearing implants, but also disposables such as cell culture dishes, pipette tips or high-throughput screening systems, as well as complete infusion systems, including the refinement of valves, supply lines, and catheters. Another focus is the modification of surfaces for the immobilization of enzymes, bacteria or cells for biocatalytic systems and fermenters. To find the optimal solution for every application, INP offers a wide range of functional surfaces for use in the following areas:

  • Improvement of cell adherence 
  • Anti-adhesive surfaces
  • Microstructured cell growth
  • Antimicrobial surfaces
  • Optimized surfaces for immobilization of enzymes

Photocatalytic Surfaces Since process costs and the simple integration of plasma processes into existing production lines are of great importance, especially in industrial applications, INP offers low-pressure processes for maximum purity, as well as processes at atmospheric pressure for short process times.


APPLICATION FIELDS

The spreading of pathogenic microorganisms is omnipresent and difficult to prevent, even with the most stringent hygienic measures and frequent disinfection measures. In particular, multi-resistant germs are a major problem today Consequently hygiene plays a central role in health-sensitive areas, such as hospitals and medical practices. Accordingly, the requirements for excluding the possibility of contamination are high. Surfaces that have an antimicrobial effect can make an essential contribution toward solving this problem.

Through use of plasma-based or sol-gel based coating procedures properties with an antimicrobial effect can be generated. To avoid resistances, such as occur through use of antibiotics, or to kill off bacteria that have already developed a resistance, among other things metal ions (especially copper or silver) or transition metal oxides (for example, molybdenum trioxide) can be incorporated in the material surface or applied as a coating. Through these measures, germ-free services can be produced that are effective against a variety of bacteria, viruses and fungi. 

Through plasma-induced surface modification of biomaterials, according to the composition of the process gas used, protein adhesion and cell adhesion can be selectively controlled – positively (cell attracting) or negatively (cell repelling). One focus area is generation of bio-relevant surface properties of implant materials, with the aid of specially tuned plasma processes. Rapid acceptance by the surrounding tissue is crucial for integration of an implant or bone replacement material. In this regard the boundary surface between biomaterial and bone plays a decisive role. Selective optimization of adhesion, migration, and proliferation of the body's own cells on the boundary surfaces should cause improved ingrowth, a reduced risk of infection, and an increase in the stability of an implant. Inversely however, via plasma treatment the adhesion properties of surfaces can be controlled in such a manner that the linking of cells or microorganisms is inhibited. For example this is a great advantage for temporary implants.

Adhesion of bacteria and cells on biomaterials is also controlled through the morphologic surface shaping, among other things. Currently there are various methods for either reducing surface roughness or for selectively structuring the surface. For example, the plasma polishing method offers a number of advantages in one process step. In the manufacturing of medical products, process-related burrs and residues occur on edges and surfaces that can impair compatibility relative to tissue and cells. Through plasma polishing these surfaces can be deburred with controlled stock removal. Additional applications are decoating, improvement of corrosion protection r friction reduction, as well as hydrophilization of different materials and alloys. On the other hand, the plasma spraying method enables deposition of porous, bioactive coatings that favor bone integration through ingrowth of the bone tissue, and consequently the clamping and intermesh of the implant with the bone (e.g. endoprosthesis). 

A layer of metal oxide characterizes photocatalytic surfaces, usually TiO2, which is activated by irradiation in the UV or visible wavelength range. For example, in combination with a naturally occurring thin film of water, OH radicals are formed, which then interact with cells, microorganisms, fats and other fluids. In combination with an ideal wettability of the surface, self-cleaning or easy-to-clean surfaces are therefore often mentioned.

Such surfaces are particularly advantageous for implants. TiO2 is approved as a material for medical implants so that no major hurdles need to be overcome with regard to the approval of such a refined implant as a medical device. The following advantages characterize our coating systems:

  • Super-hydrophilic, very good wetting with blood and other human sera
  • Increased cell vitality by more than 40%
  • 50% reduction in microbial proliferation, even in mixed cultures
  • Excellent adhesion, grade 0 cross-cutting test
  • Targeted doping for combined effects, such as extreme hardness, significantly increased the antimicrobial effect
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Saos-2 bone epithelial cells on a reference sample
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on activated TiO2

For a variety of applications, it is important that proteins are immobilized on surfaces. For a protein that is anchored to a surface to show its natural activity depends, to a large extent, on the type of anchoring. Linkers and spacers are used for covalent coupling of proteins to surfaces. The decisive factor is that the protein retains its natural activity.

Enzymes may be particular and selective, but in many cases, they do not meet the process stability requirements of industrial applications. The enzyme or biocatalyst can be immobilized to improve stability, and the yield increased by plasma-assisted surface modification of the carrier material.

For the purification of bodily fluids, the exchange of substances in blood plasma filters takes place via membranes. Toxic biomolecules, such as, e.g., endotoxins, can be coupled by plasma treatment of membrane surfaces. The plasma-treated membranes significantly reduce the endotoxin concentration in the solution compared to the control (untreated membrane).

  • Plasma treatment increases the wettability of the porous material structure so that the aqueous enzyme solution can penetrate well
  • Enzyme activity increased by a factor of 40 compared to the untreated carrier material
  • Increased stability of adsorptively bound enzymes in an anhydrous environment
  • Reusability of the enzymes immobilized on plasma-modified carriers, in some cases without significant loss of activity in several cycles
  • Further possibilities through the use of linker and spacer
  • A significant increase in the binding capacity of proteins (increased by 30% for endotoxins)

Microfluidic systems are of great importance for bioanalytics and integrate analytical functions such as sample delivery, chemical and biochemical reaction and detection on a chip. With the help of plasmas and masks, chemically different microstructures can be produced on a variety of materials. Multi-step plasma process sequences can be used to create a combination of cell-attracting and cell-repellent areas on the surface.

Applications: tissue-like cell culture systems, neural networks, bio-artificial organs, DNA and protein bio-chips, implants, biosensors, drug screening.

  • Masking and thus generated growth structures are variable
  • Long-term stable plasma-chemical functionalizations and coatings with very good adhesion to a variety of surfaces
  • Cell growth locally increased or suppressed
  • Generation of local functionalities with structural dimensions from 30 µm
  • Successfully shown for different cell lines (e.g., HEK cells, keratinocytes, human umbilical cord endothelial cells/fibroblasts and mouse fibroblasts)
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3T3-cells (mouse fibroblasts) on masked amino-functionalized plastic
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3T3-cells (mouse fibroblasts) on masked amino-functionalized plastic
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3T3-cells after 2 days of cultivation
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Plasma-assisted generation of a triple chemical structure - detection with XPS-Imaging (N1s)

Ceramic surfaces are often used in technical and biomedical applications. The already established plasma spraying method allows the production and development of unique coatings with customized, multiple properties compared to conventional methods. Powders are used as the basic material, which is (partially) melted by plasma and shot at the product at high speed. The uniqueness of the process lies in the almost unlimited combination possibilities and mixtures of powders (metals, glasses, ceramics, polymers, etc.) and the high material deposition rate or layer thickness. Furthermore, the coatings are characterized by very high adhesion and, depending on the material combination, in a unique hardness. In the biomedical field, coatings withTiO2, CaP, CaCO3, copper, silver, ZnO and their mixtures are the core expertise of INP. However, the spectrum is also constantly being expanded using new ideas and coating processes. The system used at INP is an industrial system with a prevalent plasma source. This has the advantage that coatings and coating systems developed with this system can be taken over 1:1 by the customer or ordered from established contract coaters without process scaling.

We can create the following layers:

  • CaP/HAp (soft, bone-like substance, fast integration into the bones of implants, incl. admixture of active agents or biocides)
  • YSZ (Color: white - cosmetic effect on dental implants; hard and low-wear, e.g.: on dental implants, on turbine blades, roller bearings...)
  • Cu (offers additional anti-microbial properties, especially for implant surfaces)
  • TiO2( hard layer, good integration behavior with implants, slightly antimicrobial effect, easy approval)
  • Al2O3 (Color: white - cosmetic effect; hard coating, wear-resistant, very high chemical resistance)

We specialize in:

  • Development of new coatings as well as multi-layer and multi-material coating systems (e.g., ceramic/metal, ceramic/polymer, metal/polymer)
  • Resorbable surfaces (implants) 
  • Implants with/without anti-microbial effect
  • Optimization of existing processes with regard to process and cost efficiency
  • 3D-adaptation to different substrates and products

Advantages:

  • Fast process
  • High Coating Rates
  • No melting of the substrate
  • High adhesive strength
  • Areas of the application can be combined
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Plasma spraying of a hip stem with TiO2
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YSZ dental implant both untreated, and coated with TiO2
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antimicrobial HAp/Cu surface


Projects

In the modern food processing industry, pathogens that enter food via food residues in production or processing machines pose a considerable problem. The most dangerous pathogens include salmonella, noroviruses, and listeria. These microbes can cause various diseases, which in extreme cases can also be fatal for humans. Cleaning the equipment in the food industry, therefore, requires a great deal of time and financial resources. A further complication is that many of the cleaning agents used are very aggressive and attack the materials used in the production facilities. The aim of this R & D project is, therefore, to expand and in some cases, even to replace the chemical cleaning of conveyor belts in the food industry by environmentally friendly in-line disinfection and surfaces coating. The surfaces of the conveyor belts are to be dried, disinfected, conditioned and protected from recontamination, using antibacterial layers. Appropriate processes are developed to achieve this, like a conveyor belt specially modified to the processes, and an in-line module for integration into new and existing conveyor belt systems.

Over a thousand research laboratories around the world are investigating nerve cells from the brain and other tissues using electrophysiological methods. To demonstrate the function of individual cell building-blocks, it makes sense to cultivate the nerve cells isolated from each other in a nutrient solution in the incubator. In this way, it is possible that only one cell at a time responds to a single stimulus, without interference signals from thousands of other nerve cells. The process of the so-called autaptic-cell culture is established. The nerve cells are separated so that cells can only adhere to certain areas of the glass bottom of the culture dish, usually round spots ("islands") with a diameter of approx. 200 µm. In this project, cell carriers for autaptic-cell culture are developed and optimized with the aid of plasma polymer coatings.

For patients with permanent catheters, there is a high risk of urinary tract infection, which can currently only be treated with drugs. This leads not only to the microorganisms to be treated developing resistance but also in the known interactions and side effects for the patient and additional treatment costs for the clinics and practices. With plasma-assisted processes, it is possible to produce antimicrobial surfaces on medical devices as well; these, therefore, represent a promising possibility for the refinement of medical devices, especially for the gentle treatment of plastics for use in the urinal tract. The PlasKat project deals with options for realizing such plasma-based surface refinements, e.g., for reducing the infection rate/encrustation rate and thus a longer waiting time compared to standard products. Besides the development of a plasma process for the modification of plastic medical devices, the characterization of surface properties, e.g., with X-ray photoelectron spectroscopy (XPS), the determination of the release characteristics of the active substances and the antimicrobial testing of the test specimens are important tasks in the project.


Publications

Hempel, F.; Steffen, H.; Busse, B.; Finke, B.; Nebe, J.-B.; Quade, A.; Rebl, H.; Bergemann, C.; Weltmann, K.-D.; Schröder, K.:
On the Application of Gas Discharge Plasmas for the Immobilization of Bioactive Molecules for Biomedical and Bioengineering Applications "Biomedical Engineering - Frontiers and Challenges",
ISBN: 978-953-307-309-5, InTech, August 8, 2011 - edited by Reza, F.-R. 

Hempel, F.; Finke, B.; Zietz, C.; Bader, R.; Weltmann, K.-D.; Polak, M.:
"Antimicrobial surface modification of titanium substrates by means of plasma immersion ion implantation and deposition of copper", Surf. Coat. Technol.

Finke, B.; Polak, M.; Hempel, F.; Rebl, H.; Zietz, C.; Stranak, V.; Lukowski, G.; Hippler, R.; Bader, R.; Nebe, J.-B.; Weltmann, K.-D.; Schröder, K.: 
"Antimicrobial potential of copper-containing titanium surfaces generated by ion implantation and dual high power impulse magnetron sputtering", Adv. Eng. Mat. 14 (2012), B224-B230  

Finke, B.; Testrich, H.; Rebl, H.; Walschus, U.; Schlosser, M.; Zietz, C.; Staehlke, S.; Nebe, J.-B.; Weltmann, K.-D.; Meichsner, J.; Polak, M.:
Plasma-deposited fluorocarbon polymer films on titanium for preventing cell adhesion: a surface finishing for temporarily used orthopaedic implants,
DOI: 10.1088/0022-3727/49/23/234002, J. Phys. D: Appl. Phys. 49 (2016), p. 234002   

Kredl, J.; Kolb, J.-F.; Schnabel, U.; Polak, M.; Weltmann, K.-D.; Fricke, K.:
Deposition of Antimicrobial Copper-Rich Coatings on Polymers by Atmospheric Pressure Jet Plasmas
DOI: 10.3390/ma9040274, Materials 9 (2016), p. 274

Fricke, K.; Girard-Lauriault, P.-L.; Weltmann, K.-D.; Wertheimer, M.-R.:
Plasma polymers deposited in atmospheric pressure dielectric barrier discharges: Influence of process parameters on film properties, DOI: 10.1016/j.tsf.2016.01.057, Thin Solid Films 603 (2016), p. 119-125  

Polak, M.; Ihrke, R.; Quade, A.; Hempel, F.; Fröhlich, M.; Weltmann, K.-D.:
Plasmapolieren - Reinigung, Entfettung, Entgratung und Hochglanzpolitur in einem Prozessschritt,
ISBN: 978-3-87480-292-5, Jahrbuch Oberflächentechnik, Bad Saulgau: Leuze (2015)

Finke, B.; Rebl, H.; Hempel, F.; Schäfer, J.; Liefeith, K.; Weltmann, K.-D.; Nebe, J.-B.:
Aging of Plasma-Polymerized Allylamine Nanofilms and the Maintenance of Their Cell Adhesion Capacity,
DOI: 10.1021/la5019778, Langmuir 30 (2014), p. 13914-13924

Quade, A.; Ihrke, R.; Kellner, U.; Bergemann, C.; Schröder, K.:
Stimulierung des Zellwachstums in dreidimensionalen Stützgerüsten durch plasmagestützte Oberflächenveredelungen Knöcherne Geweberegeneration, ISBN: 978-3-8440-3049-5, Aachen: Shaker-Verl. (2014) 

Finke, B.; Testrich, H.; Rebl, H.; Nebe, B.; Bader, R.; Walschus, U.; Schlosser, M.; Weltmann, K.-D.; Meichsner, J.:
Anti-adhesive finishing of temporary implant surfaces by a plasma-fluorocarbon-polymer,
Mat. Sci. Forum 783-786 (2014), p. 1238-1243  

Fricke, K.; Duske, K.; Quade, A.; Nebe, B.; Schröder, K.; Weltmann, K.-D.; v. Woedtke, Th.:
Comparison of Nonthermal Plasma Processes on the Surface Properties of Polystyrene and Their Impact on Cell Growth,
IEEE Trans. Plasma Sci. 40 (2012), p. 2970-2979

Contact

Leibniz Institute for Plasma Science and Technology
Felix-Hausdorff-Str. 2
17489 Greifswald

Dr. Katja Fricke
Programme Manager "Bioactive Surfaces"

Phone: +49 3834 - 554 3841
Fax: +49 3834 - 554 301

k.frickeinp-greifswaldde
www.leibniz-inp.de

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