Method and device for creating a nuclease-free surface
Patent Information
- Authority / Receiving Office
- EP · EP
- Patent Type
- Applications
- Current Assignee / Owner
- 3CON PHARMA MEDICAL GMBH
- Filing Date
- 2024-09-07
- Publication Date
- 2026-07-01
Smart Images
Figure EP2024075049_13032025_PF_FP_ABST
Abstract
Description
[0001] Method and device for creating a nuclease-free surface
[0002] Field of the invention
[0003] The present invention relates to a method and a device for creating a nuclease-free surface, in particular a nuclease-free polymer surface.
[0004] background
[0005] RNA (ribonucleic acid) and DNA (deoxyribonucleic acid) are two important nucleic acids that perform different functions in living organisms. As is well known, DNA carries the genetic information that influences, among other things, the development, growth, function, and reproduction of an organism. DNA is inherited from one generation to the next. RNA plays a key role in protein biosynthesis. Messenger RNA (mRNA) transfers the genetic information from the DNA in the cell nucleus to the ribosomes in the cytoplasm, where proteins are produced.
[0006] Novel vaccines and medical treatments based on RNA or DNA are currently being developed. For example, so-called mRNA vaccines have been successfully used as a COVID-19 vaccine. Other mRNA vaccines against diseases such as Lyme disease, chlamydia, cytomegalovirus, dengue fever, herpes, influenza, HIV, and malaria (to name just a few) are currently in development.
[0007] In addition, RNA- or DNA-based drugs are also being researched in cancer therapy, particularly cancer immunotherapy, and for the treatment of genetic defects. These RNA and / or DNA-based technologies have great potential to successfully treat diseases that are currently untreatable or only inadequately treatable, or at least to limit their symptoms.
[0008] However, even minor contamination with nucleases (ribonuclease, or RNase for short, and deoxyribonuclease, or DNase for short) can damage or even render the novel vaccines or active substances unusable. RNases are enzymes that catalyze the hydrolytic cleavage of phosphodiester bonds in ribonucleic acid (RNA) chains. Deoxyribonuclease is an enzyme that catalyzes the hydrolysis of deoxyribonucleic acid (DNA) molecular chains into shorter molecular chains or individual building blocks. If this catalysis occurs, the vaccine or active substance can be damaged, inactivated, or even rendered unusable.
[0009] Therefore, all containers and other items that come into contact with the vaccines or active substances (e.g., during production, storage, or administration) must be thoroughly cleaned and freed of active nucleases. This cleaning includes, for example, autoclaving, treatment with hydrogen peroxide, diethyl pyrocarbonate, or ribonuclease inhibitors, treatment with heat, UV radiation, or other processes.
[0010] These cleaning processes are often expensive, time-consuming, and can cause lasting damage to the treated surfaces or objects. For example, it is known that RNases and DNases are destroyed by autoclaving for 15 minutes at a minimum of 121°C or by heating to 180°C for at least eight hours. The high temperatures and / or the chemicals used cause lasting damage to many materials, especially polymers, so that these materials cannot be used, or can only be used to a limited extent, for the production and storage of the new vaccines and active substances.
[0011] In conventional bioreactors, the vaccines or active substances to be produced are usually stored in stainless steel tanks lined with a polymer film or a polymer film bag and / or connected via polymer tubes.
[0012] Typical polymers for use in bioreactors are
[0013] • ETFE - ethylene-tetrafluoroethylene copolymer,
[0014] • FEP - tetrafluoroethylene-hexafluoropropylene copolymer,
[0015] • HDPE - High Density Polyethylene,
[0016] • LDPE - Low Density Polyethylene,
[0017] • PC - Polycarbonate Polyethylene terephthalate,
[0018] • PFA - perfluoroalkoxy polymers,
[0019] • PP / PCO - Polypropylene / Polycyclooctene,
[0020] • PMP - polymethylpentene,
[0021] • Barex - acrylonitrile plastic,
[0022] • HIPS - High Impact Polystyrene,
[0023] • PVC - polyvinyl chloride,
[0024] • TPE - thermoplastic elastomer,
[0025] • and the like.
[0026] However, these are not or only partially suitable for being freed from active DNase and RNase using conventional methods.
[0027] JW Lackmann et al. "A dielectric barrier discharge terminally inactivates RNase A by oxidizing sulfur-containing amino acids and breaking structural disulfide bonds"; 2015, J. Phys. D: Appl. Phys. 48 494003 describes the inactivation of RNase A by means of a dielectric barrier discharge.
[0028] Description of the invention
[0029] It is therefore the object of the present invention to provide a method and a device for creating a nuclease-free surface, which method is particularly applicable to polymer surfaces. This object is achieved by a method according to claim 1 and a device according to claim 14. Further aspects of the invention are set forth in the dependent claims and the following description.
[0030] In particular, the object is achieved by a method for creating a nuclease-free surface, in particular a nuclease-free polymer surface. The terms nuclease-free and / or DNase-free are used synonymously below. The term nuclease-free surface refers to a surface that is essentially free of active RNases and DNases. An essentially RNase- and DNase-free surface is a surface whose RNase and DNase contamination is so low that RNA- or DNA-containing products (such as drugs or vaccines) that come into contact with this surface are no longer damaged or inactivated.
[0031] The method is applicable to all types of surfaces, especially polymer surfaces. However, it is understood that other surfaces, such as metallic surfaces, ceramic surfaces, glass surfaces, or the like, can also be rendered free of active nucleases using the method according to the invention.
[0032] To test whether a surface is nuclease-free (i.e., RNase or DNase-free), a sample of this surface can be extracted for 1 hour in RNase-free water (UltraPure Distilled Water DNase / RNase Free) or DNase-free water. If the surface is the internal surface of a volume (e.g., a polymer film pouch), the volume can be filled with RNase (or DNase)-free water and extracted for 1 hour. 45 μl of this RNase-free water can be treated with an RNase test kit (e.g., RNaseAlert™ Lab Test Kit, catalog number: AM1964) or DNase test kit and analyzed in a fluorometer (Mithras LB 940, Berthold Technologies). If the amount of RNase (or DNase) in the sample water relative to the amount of RNase (or DNase) in the RNase / DNase-free water (blank) is below 1.5, the surface can be considered RNase (or DNase) free. In the range of 1.5 to 2.5, the sample is slightly contaminated.A value of 2.5 or higher indicates severe contamination. The method according to the invention comprises the following:
[0033] • Generating a first non-thermal plasma under atmospheric pressure, wherein the first non-thermal plasma is generated at a discharge surface with a first power;
[0034] • Generating a second non-thermal plasma under atmospheric pressure, wherein the second non-thermal plasma is generated at a discharge surface with a second power; wherein the second power is higher than the first power,
[0035] • Generating a plasma flow from the first non-thermal plasma and the second non-thermal plasma;
[0036] • Directing the plasma flow onto a surface to be cleaned, and
[0037] • Inactivation of nucleases on the surface to be cleaned.
[0038] Investigations by the inventors show that using two different non-thermal plasmas with different power levels allows surfaces to be freed of nucleases significantly more efficiently than using only one non-thermal plasma type. The first non-thermal plasma (hereinafter sometimes referred to as "ozone-dominated non-thermal plasma") has a higher ozone content, while the second non-thermal plasma (hereinafter occasionally referred to as "nitrogen-dominated non-thermal plasma") has a higher nitrogen oxide content. The inventors attribute this to the low solubility of O3, NO, and NO2 in water (atmospheric humidity). Using two plasma types, high-valent NOx such as N2O5 is formed, which is otherwise not observed to this extent. It has been found that N2O5 can be generated effectively and stably by mixing the escaping gases from the first air plasma and the second air plasma.By using surfaces treated with the mixed gas, a reduction of nucleases by a factor of around 10 was achieved. 6 achieved.
[0039] Optionally, the inactivated nuclease can be removed from the surface to be cleaned, for example by rinsing with RNase-free water. Often, however, inactivation of the RNase and / or DNase is sufficient, since the inactivated enzymes can no longer cleave the RNA / DNA. Furthermore, optionally, after removal of the inactivated RNase and / or DNase from the surface to be cleaned, plasma flow can be directed onto the cleaned surface again. This renewed treatment with plasma flow allows the cleaning medium (e.g., RNase-free water) to be removed from the surface and the surface to be dried. Non-thermal plasma (NTP) is a plasma that is not in thermal equilibrium. The temperatures of the particle types it contains (neutral particles, ions, electrons) therefore differ, sometimes even significantly.Ozone-dominated non-thermal plasma is a non-thermal plasma that predominantly leads to the formation of ozone (O3) in the plasma gas. Nitrogen-dominated non-thermal plasma is a non-thermal plasma that predominantly leads to the formation of nitrogen compounds such as NO, NO2, NOx, N2O, HNO3, HNO2, and / or N2O5 in the plasma gas.
[0040] The first, ozone-dominated non-thermal plasma can be generated, for example, using a dielectric barrier discharge or a corona discharge. A dielectric barrier discharge is an alternating voltage gas discharge in which at least one electrode is electrically isolated from a gas space by galvanic separation using a dielectric.
[0041] A gas space filled with (or flowing through) a process gas (e.g., air, especially (filtered) ambient air) can be ionized by electrically insulated electrodes to generate a plasma. For this purpose, an alternating voltage is applied to the electrodes, causing a discharge. Displacement currents maintain the discharge even through the insulation, allowing electrical power to be continuously transferred into the plasma.
[0042] The frequency of the alternating voltage is preferably selected so that it corresponds to a resonant frequency of the plasma generator. The resonant frequency of the plasma generator depends, among other things, on the size and material of the plasma generator, as well as on the process gas used. In one example, the frequency of the alternating voltage is approximately 40 kHz (+-10%). The higher the frequency selected, the higher the concentration of reactive nitrogen species (RNS, e.g., NO). The lower the frequency selected, the higher the proportion of reactive oxygen species (ROS, such as ozone). Furthermore, the plasma density increases with decreasing frequency.
[0043] The electrodes can be designed, for example, as parallel plates, as a wire mesh on a dielectric, a tip opposite a plate and / or the like.
[0044] A corona discharge is an electrical discharge in a non-conductive medium, for example, in a process gas (e.g., air, especially (filtered) ambient air). The discharge requires ions as charge carriers. These can either already be present (plasma) or they form in the medium as a result of field ionization if the electric field strength is high enough (e.g., > 100 kV / m). The first non-thermal plasma (ozone-dominated non-thermal plasma), for example, is generated at a power at a discharge area of below 0.5 W / cm 2 , especially below 0.4 W / cm 2 or below 0.3 W / cm 2 , or below 0.2 W / cm 2 A preferred range for generating ozone-dominated non-thermal plasma is between 0.1 and 0.2 W / cm 2In particular, the surface temperature of the electrodes for generating the ozone-dominated non-thermal plasma may be less than 90°C, in particular less than 75°C or less than 60°C.
[0045] In a further aspect, the second non-thermal plasma (nitrogen-dominated non-thermal plasma) is generated by means of a dielectric barrier discharge or a corona discharge. For example, the nitrogen-dominated non-thermal plasma is generated at a power at a discharge area of more than 0.1 W / cm 2 , especially more than 0.2 W / cm 2 , or more than 0.5 W / cm 2 , or more than 1.5 W / cm 2 , or more than 2 W / cm 2 A preferred range is between 0.5 and 2.2 W / cm 2 In particular, the surface temperature of the electrodes for generating the nitrogen-dominated non-thermal plasma can be greater than 90°C.
[0046] Preferably, however, the first non-thermal plasma is generated at a power at the discharge surface of 0.2 W / cm 2 up to < 0.45 W / cm 2 , preferably 0.3 W / cm 2 up to < 0.45 W / cm 2 is generated, wherein the second non-thermal plasma (20) at a power at the discharge surface of > 0.45 W / cm 2 up to 0.7 W / cm 2 , preferably > 0.45 W / cm 2 up to 0.6 W / cm 2 is generated. Preferably, a dielectric barrier discharge or a corona discharge is provided.
[0047] The process gas used can be, for example, air, in particular dried and / or sterilized air and / or filtered ambient air, argon, and / or the like. Mixtures of at least two different process gases can also be used.
[0048] The process gas can be purified before generating the plasma (ozone-dominated and / or nitrogen-dominated), for example, by at least one filter. The same process gas (e.g., air, in particular (filtered) ambient air) can be used to generate the first non-thermal plasma and the second non-thermal plasma, or different process gases or mixtures can be used.
[0049] In particular, the first, ozone-dominated non-thermal plasma and the second, nitrogen-dominated non-thermal plasma can be generated simultaneously, preferably in a common plasma generator. For this purpose, the electrode(s) can be designed so that different power levels are applied to their surfaces. For example, in a region with a lower power density, the first non-thermal plasma can be generated primarily, and in a region with a higher power density, the second non-thermal plasma can be generated primarily.
[0050] It is also possible for the first non-thermal plasma and the second non-thermal plasma to be generated serially, i.e., in successive periods. For this purpose, the power of the plasma generator can be adjusted (periodically).
[0051] In a further aspect of the invention, the first non-thermal plasma is generated in a first plasma generator and the second non-thermal plasma is generated in a second plasma generator different from the first.
[0052] The generated plasmas (ozone-dominated and nitrogen-dominated) are then used to generate a plasma flow. The ionized process gas is discharged from the plasma generator(s). This discharge can be achieved, for example, by at least one fan and / or at least one pump. The plasma generator(s) can also be designed so that the process gas is expelled directly.
[0053] This plasma stream is then directed onto the surface to be cleaned. The plasma stream thus comes into contact with the surface. To increase the effectiveness of the plasma stream, the plasma stream can be specifically directed onto the surface to be cleaned. This can be achieved using plasma gas nozzles, fans, and / or pumps. This allows for particularly rapid inactivation of the nuclease present on the surface.
[0054] In a further preferred embodiment, the plasma flow is mixed with an aerosol and directed onto the surfaces to be cleaned. The aerosol is generated and can comprise water, in particular distilled water, RNase / DNase-free water, and / or plasma-activated water. The aerosol can be generated, for example, using a nebulizer or atomizer.
[0055] In particular, the addition of fine water droplets, preferably with a size of less than 100 pm, particularly preferably with a size of less than 80 pm, leads to a high inactivation rate of nucleases. Preferably, distilled water (and / or RNase- and / or DNase-free water) is atomized with an ultrasonic atomizer, and this aerosol mixture is mixed with the plasma flow (generated from ozone-dominated and / or nitrogen-dominated non-thermal plasma) and directed onto the surface to be cleaned.
[0056] In another, particularly preferred embodiment, plasma-activated water (PAW) is generated and applied as an aerosol mixture to the surface to be cleaned. Particularly preferably, the aerosol mixture is generated using an ultrasonic atomizer, and ultrafine PAW aerosols are applied to the surface to be cleaned.
[0057] The surface to be cleaned can be a polymer surface, such as a polymer film, a polymer sheet, or any other shaped polymer (injection-molded, extruded, etc.). The use of non-thermal plasma or plasma flow does not damage the polymer. The use of ozone-dominated and nitrogen-dominated plasma allows for rapid and reliable inactivation of RNases and / or DNases.
[0058] It has been shown that a ratio of first to second non-thermal plasma of 70:30 (based on the discharge area) leads to good inactivation rates. The NOx preferably comprises NO, NO2 and / or higher N x O y (x > 1; y > 3), with the NO component dominating.
[0059] The ratio of the components of the plasma flow can be determined and / or varied by the plasma generator power, in particular by the alternating voltage applied to the generator (frequency, amplitude, signal shape, ...) and / or the generator geometry.
[0060] The surface to be cleaned can be located inside a hollow body. This hollow body can be formed, for example, by at least one polymer film bag or by a decontamination container. To inactivate any nucleases, the plasma flow is directed into the interior of the hollow body. In particular, the plasma flow can be directed into the hollow body through at least one inlet. It is also possible to direct the plasma flow into the hollow body through multiple inlets. This increases the cleaning performance.
[0061] If the surface to be cleaned comprises several interior spaces of several hollow bodies, these can be connected to one another via at least one plasma gas line so that the plasma flow can be introduced into the corresponding interior spaces. For example, several polymer film bags can be connected to one another, or several polymer film bags can be filled with / flowed through with a plasma flow in parallel via a corresponding distributor. It is also possible to clean the inner surface of so-called single-use assemblies. For this purpose, the plasma flow is introduced into the single-use assembly and the inner surface is cleaned. Single-use assemblies typically comprise one or more polymer film bags and at least one hose line. Valves and / or the like can also be part of the single-use assemblies. These single-use assemblies are used, among other things, for the production of active ingredients or vaccines, or their precursors or intermediate products.In addition to the inner surface of the interior, objects within the hollow body can also be freed of active nuclease. For example, filters can be incorporated into the hollow body through which the plasma flow is directed. Likewise, other objects to be cleaned can be provided, for example, in a decontamination container. These can include, for example, pipettes, pipette tips, cannulas, vials, disposable medical or pharmaceutical plastic items, disposable bioprocess containers, laboratory equipment, centrifuges, PCR units, or the like. The inflowing or overflowing plasma flow then inactivates nucleases.
[0062] The method can further comprise a leak test of at least the hollow body, wherein a plasma gas from the generated plasma flow is used as the test gas. If the plasma gas is introduced into the hollow body and the pressure in the hollow body is measured, the pressure drop can be used to determine whether the hollow body (for example a polymer film bag) is leak-tight or has a leak. This means that an additional leak test is not necessary. Immediately after inactivation of the RNases / DNases, it can be determined whether the hollow body, for example a polymer film bag, is leak-tight and RNase / DNase-free. A hollow body which fulfills both criteria can then be used, for example, in a bioreactor in a bioprocess line for the production of vaccines or active ingredients, for the transport and / or storage of the vaccines or active ingredients. Likewise, precursors and / or intermediates of the vaccines or active ingredients can be stored in the polymer film bags.Active ingredients are manufactured, transported and / or stored.
[0063] The method can further comprise unwinding a polymer film and / or extruding a polymer film, and wherein the plasma flow is directed onto at least one side of the extruded and / or unwound polymer film. Thus, the method can be directly integrated into the film manufacturing process or into the manufacturing process of film products, such as polymer film bags. A separate inactivation step can therefore be omitted, since the film or film products produced are immediately RNase-free or DNase-free, respectively. It is also possible to treat surfaces multiple times with the plasma flow. For example, the plasma flow can be directed onto at least one surface of the film (in particular a later inner side of a polymer film bag) immediately after unwinding the film, after extruding the film, and / or after stretching (unidirectionally or bidirectionally) a film.For this purpose, a plasma gas nozzle can be provided, preferably spanning the entire width of the film, or several plasma gas nozzles can be provided, arranged side by side or offset from one another (in a film removal direction), so that the surface of the film is preferably exposed to plasma flow across its entire width. Likewise, the at least one plasma gas nozzle can be movably arranged and moved across the surface of the film to direct the plasma flow onto the surface to be cleaned.
[0064] In one aspect, the film is guided through a decontamination zone, where the plasma flow is initiated. The decontamination zone can have a longitudinal extension. The treatment time of the film's surface with the plasma flow can then be determined by the film's withdrawal speed. Within the decontamination zone, the film can be deflected (multiple times) (e.g., by means of rollers).
[0065] In particular, the plasma flow can be directed onto the surface to be cleaned for a period of at least 10 minutes, or at least 15 minutes, or at least 20 minutes.
[0066] Furthermore, the method can comprise creating at least one volume, in particular at least one polymer film bag. The volume is preferably produced from at least one polymer film. For this purpose, a polymer film can be folded and the edge regions of the folded polymer film are joined to one another (preferably by a material bond), or several polymer films (at least two) are placed on top of one another and joined to form a volume having an inner surface (e.g., a polymer film bag). Creating the at least one volume can comprise polymer bonding and / or polymer welding. For example, the polymer welding can be heat welding, high-frequency welding, pulse welding, ultrasonic welding, friction welding, laser welding, and / or the like.
[0067] The nuclease-free inner surface of the volume can be created before and / or after polymer bonding and / or polymer welding. For example, nuclease-free films can be welded together, or the volume can be created first and then the plasma flow can be introduced into it.
[0068] The object is further achieved by a device for creating a nuclease-free surface. The device is configured to carry out the method described above. The device comprises at least one plasma reactor with at least one discharge surface on a discharge device, wherein a first non-thermal plasma with a first power can be generated at the discharge surface under atmospheric pressure using the plasma reactor. Furthermore, a second non-thermal plasma with a second power can be generated at the discharge surface under atmospheric pressure using the plasma reactor, wherein the second power is higher than the first power.Furthermore, the device comprises at least one plasma gas line which is connected to the at least one plasma reactor and is configured to direct a plasma flow generated from the first non-thermal plasma and the second non-thermal plasma onto a surface to be cleaned in order to inactivate nucleases on the surface to be cleaned.
[0069] It is preferably provided that the plasma reactor with the discharge surface on the discharge device comprises a dielectric discharge device or a corona discharge device.
[0070] With regard to the generation of the non-thermal plasma, it is preferably provided that the discharge device for generating the first non-thermal plasma at a power at the discharge surface of 0.2 W / cm 2 up to < 0.45 W / cm 2 , preferably 0.3 W / cm 2 up to < 0.45 W / cm 2and that the discharge device for generating the second non-thermal plasma at a power at the discharge surface of > 0.45 W / cm 2 up to 0.7 W / cm 2 , preferably > 0.45 W / cm 2 up to 0.6 W / cm 2 For this purpose, a control device can be provided, for example, which controls the discharge device to generate the first non-thermal plasma at a power at the discharge surface of 0.2 W / cm 2 up to < 0.45 W / cm 2 , preferably 0.3 W / cm 2 up to < 0.45 W / cm 2 and the discharge device for generating the second non-thermal plasma at a power at the discharge surface of > 0.45 W / cm 2 up to 0.7 W / cm 2 , preferably > 0.45 W / cm 2 up to 0.6 W / cm 2Two separate discharge devices can be provided, one of which is set by the control device to the first power and one of which is set by the control device to the second power.
[0071] Surprisingly, it was found that with a discharge area of the discharge device of 1.0 cm 2 up to 6.0 cm 2 , preferably at 2.5 cm 2 up to 3.5 cm 2When operated in both power modes, preferably RNA (reactive nitrogen species) is generated by the plasma reactor in the plasma generation unit at low ozone concentrations. RNA, in particular nitrogen oxides such as N2O5 and NO, are reactive with nucleases. This results in a reduction in the amount of nucleases across the entire treated surface. Therefore, it is preferably provided that the plasma reactor is designed such that the first non-thermal plasma and the second non-thermal plasma can be generated simultaneously with a plasma reactor. This occurs via the discharge surface of the discharge device. A control device can actuate the plasma reactor in this way.
[0072] The plasma reactor is suitable for atmospheric dielectric barrier discharge and comprises, for example, a glass tube and an inner electrode made of bundled stainless steel fibers, wherein the bundled stainless steel fibers are arranged spirally inside the glass tube, wherein an outer electrode designed as a sieve cylinder is provided. The inner electrode is preferably made of bundled stainless steel fibers with a fiber diameter of approximately 1.0 to 1.2 μm, preferably approximately 1.0 μm, wherein the length of the fibers is approximately 3 to 6 cm. When installed, the fibers predominantly overlap. The sieve cylinder is preferably made of a metal wire mesh, wherein the wire diameter is preferably between approximately 0.18 and 0.36 mm and the spacing of the longitudinal wires is preferably between approximately 0.4 and 1.25 mm.
[0073] The glass tube forms a dielectric in conjunction with the sieve cylinder as the outer electrode. With a glass tube material thickness of 0.18 to 0.36 mm and a spacing of 0.4 to 1.25 mm between the longitudinal wires and the inner electrode, which consists of bundled stainless steel filaments with a thickness of 1.0 to 1.2 μm, a particularly homogeneous dielectric discharge occurs.
[0074] During operation of the device, the discharge tubes convert oxygen in the air into hydroxyl radicals. This effect is best achieved by applying high-energy alternating voltage pulses with a frequency of preferably 25 to 44 kHz.
[0075] For example, the nitrogen-dominated non-thermal plasma with a power at a discharge area of more than 0.1 W / cm 2 , especially more than 0.2 W / cm 2 , or more than 0.5 W / cm 2 , or more than 1.5 W / cm 2 , or more than 2 W / cm2 A preferred range is between 0.5 and 2.2 W / cm 2 In particular, the surface temperature of the electrodes for generating the nitrogen-dominated non-thermal plasma can be greater than 90 °C
[0076] The plasma reactor for generating the first non-thermal plasma under atmospheric pressure and the plasma reactor for generating the second non-thermal plasma under atmospheric pressure can be designed as an integral unit. The generation of the first non-thermal plasma and the second non-thermal plasma can occur at electrodes with different power densities, at different electrodes, or in series. It is also possible to use two or more different plasma generators.
[0077] Furthermore, the plasma gas line can be configured to connect the plasma reactors to at least one decontamination container, and / or to at least one hollow body having an inner surface, in particular at least one polymer film bag, and / or to at least one plasma gas nozzle. The plasma gas nozzle can direct the plasma flow into a hollow body having an inner surface or directly onto a surface to be cleaned (e.g., onto an unwound polymer film). In particular, the plasma gas nozzle can be directed onto at least one polymer film, wherein the polymer film is an extruded film, an unwound film, and / or a film to be welded, and wherein the plasma gas nozzle is optionally configured to be movable.
[0078] In a further aspect, the device forms a closed circuit for the plasma flow. The generated plasma gas flow is thus circulated. After generation, the plasma gas flow is directed onto the surface to be cleaned and then, after optional cleaning (e.g., filtering), fed back to the plasma generator(s). This allows the plasma gas flow to be generated in an energy-efficient manner. Furthermore, additional process gas (e.g., air, in particular (filtered) ambient air) can be fed into the circulated plasma gas flow. The process gas is preferably fed upstream of the plasma generator(s).
[0079] The device may further comprise a polymer welding device (e.g., a heat welding device, an ultrasonic welding device, a friction welding device, a laser welding device, a high-frequency welding device, an impulse welding device, and / or the like) configured to weld polymer film to form polymer film bags. Inactivation may occur before or after welding.
[0080] The device may further comprise an unwinding device for unwinding polymer film. Inactivation may occur immediately after unwinding. Additionally and / or alternatively, inactivation may also occur immediately after film production, e.g., after extrusion and / or stretching, i.e., before the film is wound up or further processed.
[0081] Furthermore, the device can comprise a decontamination container connected to the at least one plasma gas line. Thus, the decontamination container and its contents can be freed of active RNase / DNase.
[0082] Furthermore, the object is achieved by a polymer film bag for use in a bioreactor, wherein the polymer film bag is made from an RNase-free polymer film and / or wherein the polymer film bag has been freed of active RNase / DNase after its production. The inner surface of the polymer film bag is thus free of RNase and / or DNase.
[0083] Short description of the characters
[0084] Specific embodiments of the present invention are illustrated in the accompanying figures. These are intended to assist in understanding the invention. In particular, Figure 1 shows a schematic representation of a method for creating an RNase-free and / or DNase-free surface;
[0085] Fig. 2 is a schematic representation of a first device for creating an RNase-free and / or DNase-free surface;
[0086] Fig. 3 is a schematic representation of a second device for creating an RNase-free and / or DNase-free surface;
[0087] Fig. 4 is a schematic representation of a third device for creating an RNase-free and / or DNase-free surface;
[0088] Fig. 5 is a schematic representation of a fourth device for creating an RNase-free and / or DNase-free surface, and
[0089] Fig. 6 is a schematic representation of a fifth device for creating an RNase-free and / or DNase-free surface.
[0090] Fig. 7 is a schematic representation of a sixth device for creating an RNase-free and / or DNase-free surface.
[0091] Fig. 8 shows schematically investigations on five plastic surfaces contaminated with RNase (Bl to B5) before and after the method according to the invention.
[0092] Detailed description of the characters
[0093] Figure 1 shows a schematic representation of a method 1000 for creating an RNase-free and / or DNase-free surface. The method 1000 comprises the following:
[0094] • Generating 1100 a first, ozone-dominated non-thermal plasma at atmospheric pressure;
[0095] • Generating 1200 a second, nitrogen-dominated non-thermal plasma 20 under atmospheric pressure;
[0096] • Generating 1300 a plasma flow from the ozone-dominated non-thermal plasma and the nitrogen-dominated non-thermal plasma;
[0097] • Directing 1400 of the plasma flow onto a surface to be cleaned, and
[0098] • Inactivation of RNase and / or DNase on the surface to be cleaned.
[0099] Optionally, the inactivated RNase and / or DNase can also be removed from the surface to be cleaned. 1600. The 1000 procedure was applied to polymer film bags with a volume of 1 liter. First, the polymer film bags were filled with 50 mL of RNase-free water (UltraPure Distilled Water DNase / RNase Free, Thermo Fisher Scientific) and extracted for at least 1 h. Subsequently, 45 μl of the sample was treated with an RNase test kit and subsequently analyzed in a fluorometer (Mithras LB 940, Berthold Technologies). The RNase test kit used (RNaseAlert™ Lab Test Kit Catalog number: AM1964 from Thermo Fisher Scientific) has a detection limit of 3.5 x 100 -7 Units (~0.5 pg) RNase A. Limits that standardize the terms RNase / DNase-free have not yet been established.
[0100] It was shown that a longer extraction time leads to an increase in the measured RNase content. With an extraction time of 12 hours, the RNase content was almost twice as high, and with an extraction time of 24 hours, the RNase content was almost 7.5 times higher compared to the one-hour extraction. By treating the inner surface of the polymer film bags for 20 minutes according to method 1000, the relative amount of RNase compared to a sample of pure, RNase-free water was reduced from an initial 11.5 to below 1.5, specifically to 1.17. The treated surface was thus once again RNase-free.
[0101] The effect of the method according to the invention is also illustrated in Fig. 8, which shows a test using five plastic surfaces. The measurements for the presence of RNase were carried out using the RNaseAlert™ Lab Test Kit Catalog number: AM1964 from Thermo Fisher Scientific. The ordinate shows relative fluorescence units. The measurement results in a value of 51 for a control (= RNase-free) both before and after treatment. The five samples were treated with different amounts of RNase. After treatment, all samples were below the threshold of 100, which is considered RNase-free. Most samples (B1, B2, B4, and B5) were at the control level after treatment.
[0102] Figure 2 shows a schematic representation of a first device 100 for creating an RNase-free and / or DNase-free surface 102, 104, 106. The device 100 comprises at least one plasma reactor 112 for generating an ozone-dominated non-thermal plasma 12 under atmospheric pressure and at least one plasma reactor 114 for generating a nitrogen-dominated non-thermal plasma 14 under atmospheric pressure.
[0103] Two different plasma reactors 112, 114 can be used to generate the first non-thermal plasma 12 and the second non-thermal plasma 14, or a common plasma reactor 110 can be used. The plasma reactors for generating the ozone-dominated non-thermal plasma 12 and the nitrogen-dominated non-thermal plasma 14 can thus be integrally formed in a housing.
[0104] A plasma flow 26 generated from the ozone-dominated non-thermal plasma 12 and the nitrogen-dominated non-thermal plasma 14 can be directed onto a surface 102, 104, 106 to be cleaned via a plasma gas line 126 connected to the plasma reactors 110; 112, 114 in order to inactivate RNase and / or DNase on the surface 102, 104, 106 to be cleaned.
[0105] In the embodiment shown in Figure 2, the device 100 comprises a decontamination container 50, which is connected to the plasma gas line 126. The generated plasma flow 26 is thus introduced into the decontamination container 50 and directed onto the surfaces 102, 104, 106 to be cleaned. Thus, not only the inner surface 102 of the decontamination container 50 can be freed of active RNase / DNase, but also the surfaces 104, 106 of the
[0106] Items arranged in the decontamination container 50. These items may include, for example, pipettes, pipette tips, cannulas, vials, and / or the like.
[0107] The plasma flow 26 can be circulated via a plasma gas line 127. After flowing through the decontamination container 50, the plasma flow 26 is cleaned, for example, in a filter 210 and passed back to the plasma reactor(s) 110; 112, 114. Furthermore, additional process gas (e.g., air, in particular (filtered) ambient air) can be fed into the plasma reactor(s) 110; 112, 114 via a gas inlet 212. The process gas can be cleaned, in particular filtered, before entering the plasma reactor(s) 110; 112, 114.
[0108] Fig. 3 shows a schematic representation of a second device 100' for creating an RNase-free and / or DNase-free surface. The plasma flow is generated essentially as described with reference to Figure 1. However, as shown, the plasma gas line 126 is not connected to a decontamination container; instead, the plasma flow 26 is introduced directly into a polymer film bag 42 in order to free its inner surface 102 of active RNase / DNase. The polymer film bag 42 can be connected to further polymer film bags 44, 46 via additional plasma gas lines (in particular hoses), so that their inner surfaces 104, 106 as well as the components connected thereto (e.g., hoses, valves, connectors, etc.) can also be freed of active RNase / DNase. The polymer film bags 42, 44, 46 can be connected in series as shown, or filled or discharged in parallel with plasma flow 26 via a suitable distributor.flow through. Here, too, the plasma flow 26 can be guided in a circuit. Likewise, the inner surface of so-called single-use assemblies can be cleaned with the device 100'. For this purpose, the plasma flow 26 is introduced into the single-use assembly and the inner surface is cleaned. Single-use assemblies typically comprise one or more polymer film bags 42, 44, 46 and at least one hose line that connects the polymer film bags. In addition, valves and / or the like can be part of the single-use assembly. By introducing the plasma flow 26, the inner surface of a substantially fully assembled single-use assembly can be cleaned, or individual parts (or subassemblies) of a single-use assembly can be cleaned.
[0109] The devices 100 and 100' can also be configured to perform a leak test on the at least one polymer film bag 42, 44, 46 or the decontamination container 50, respectively, using the plasma gas of the generated plasma flow 26 as the test gas. For example, if the pressure inside the polymer film bags 42, 44, 46 or the decontamination container 50 is kept constant for a predetermined test period, it can be determined that the polymer film bags / the decontamination container are leak-tight.
[0110] Figure 4 shows a schematic representation of a third device 100" for creating an RNase-free and / or DNase-free surface. The plasma flow is generated essentially as described with reference to Figure 1. However, as shown, the plasma gas line 126 is not connected to a decontamination container, but rather to a plasma gas nozzle 226. The plasma gas nozzle 226 is directed onto a polymer film 60, which is folded here. The folded film can be welded to form a polymer film bag using a polymer welding device 240 of the device 100". The plasma gas nozzle 226 directs the plasma flow 26 before and / or during welding onto the later inner surface of the polymer film bag. Thus, an RNase / DNase-free polymer film bag can be produced, which can be used, for example, in a bioreactor as a reaction space or for the storage / transport of RNA / DNA active orvaccines and / or their precursors and / or intermediates.
[0111] Figure 5 shows a schematic representation of a fourth device 100'" for creating an RNase-free and / or DNase-free surface. The plasma flow is generated essentially as described with reference to Figure 1. However, as shown, the plasma gas line 126 is not connected to a decontamination container, but rather to two plasma gas nozzles 222, 224. The first plasma gas nozzle 222 is directed at a polymer film 60, which is unwound here from an unwinding device 260 of the device 100'". The second plasma gas nozzle 224 is directed at a polymer film 62, which is unwound from an unwinding device 262. It is understood that the device 100'" is not limited to two plasma gas nozzles 222, 224 and two unwinding devices 260, 262. For example, multiple films (at least three, four, five, ...) can also be used.) can be used for polymer film bag production, for example, to produce three-dimensional polymer film bags. Depending on the number of films, a corresponding number of unwinding devices can be provided. Each of these unwinding devices can be assigned at least one plasma gas nozzle.
[0112] The first polymer film 60 and the second polymer film 62 can be welded to form a polymer film bag via a polymer welding device 240 of the device 100''. At suitable intervals (depending on the desired bag size), the polymer welding device 240 can weld the polymer films 60, 62 by means of a weld seam 47. Instead of two separate films 60, 62,
[0113] An extruded film tube can also be used for polymer film bag production.
[0114] The plasma gas nozzle 222 directs the plasma flow 26 downstream of the unwinding device 260 onto a surface of the film 60, which will later become an inner surface of the polymer film bag. Accordingly, the plasma gas nozzle 224 directs the plasma flow 26 downstream of the unwinding device 262 onto a surface of the film 62, which will later become an inner surface of the polymer film bag. Thus, an RNase / DNase-free polymer film bag can be produced, which can be used, for example, in a bioreactor as a reaction chamber or for the storage / transport of RNA / DNA active substances or vaccines and / or their precursors and / or intermediates.
[0115] Figure 6 shows a schematic representation of a fifth device 100"" for creating an RNase-free and / or DNase-free surface. This device 100"" is constructed essentially like the device 100' from Figure 3. The device 100"" is connected downstream of a polymer bag manufacturing device 250. Polymer film bags 42 produced by the polymer bag manufacturing device 250 can be subsequently connected to the device 100"". In this way, the inner surface of the polymer film bags 42 can be freed of active RNase / DNase. The device 100"" can be integrated into a system for producing polymer film bags, preferably inline, with the produced polymer film bags preferably being connected to the device 100"" in an automated manner.The device 100"" can also be configured to carry out a leak test of the produced polymer film bag 42, wherein the plasma gas of the generated plasma flow 26 is used as the test gas.
[0116] The polymer bag manufacturing device 250 can comprise at least one polymer welding device 240, as schematically illustrated in Figures 4 and 5. Figure 7 shows a schematic representation of a sixth device 100 for creating a nuclease-free surface. This device 100 comprises a gas inlet 212, a pressure control 214, a downstream flow controller 216, as well as a first plasma generator 112 and a downstream second plasma generator 114, from which the plasma gas line 126 (with plasma flow 26) leads to the decontamination container 50, in which a surface to be treated is arranged (not shown). An on / off switch 228 and a flow sensor 229 are connected downstream of the decontamination container 50. An ozone filter 230 is also connected to this to prevent ozone from escaping from the device 100.
[0117] Some of the embodiments are described in more detail than others with reference to the accompanying figures. However, other embodiments are also included in the present disclosure. The present disclosure should not be understood as being limited only to the embodiments explicitly set forth herein; rather, these embodiments are given as examples to convey the scope of the inventive subject matter to those skilled in the art. The present invention may, of course, be embodied otherwise than as set forth herein without thereby affecting essential features of the invention. The present embodiments are to be considered in all respects as illustrative and not restrictive, so that modifications of the described embodiments may also fall within the scope of protection.
[0118] List of reference symbols
[0119] 12 first (ozone-dominated) non-thermal plasma
[0120] 14 second (nitrogen-dominated) non-thermal plasma
[0121] 26 Plasma flow
[0122] 42 polymer film bags
[0123] 44 polymer film bags
[0124] 46 polymer film bags
[0125] 47 Weld seam
[0126] 50 decontamination containers
[0127] 60 polymer film
[0128] 62 polymer film
[0129] 100 device
[0130] 102 surfaces to be cleaned
[0131] 104 surfaces to be cleaned
[0132] 106 surfaces to be cleaned
[0133] 110 Plasma Generator
[0134] 112 plasma generator 114 plasma generator
[0135] 126 Plasma gas line
[0136] 127 Plasma gas line
[0137] 210 filters
[0138] 214 Pressure control
[0139] 216 Flow control
[0140] 212 Gas inlet
[0141] 222 Plasma gas nozzle
[0142] 224 Plasma gas nozzle
[0143] 226 Plasma gas nozzle
[0144] 228 On / Off switch
[0145] 229 Flow sensor
[0146] 230 ozone filters
[0147] 240 polymer welding device
[0148] 250 polymer bag making device
[0149] 260 unwinding device
[0150] 262 Unwinding device
[0151] 1000 procedures
[0152] 1100 Generation of a first non-thermal plasma under atmospheric pressure
[0153] 1200 Generation of a second non-thermal plasma under atmospheric pressure
[0154] 1300 Generating a plasma flow from the first non-thermal plasma and the second non-thermal plasma
[0155] 1400 Directing the plasma flow onto a surface to be cleaned
[0156] 1500 Inactivation of RNase and / or DNase
[0157] 1600 Removal of inactivated RNase and / or DNase
[0158] 1700 Redirecting the plasma flow onto a surface to be cleaned
[0159] The invention further relates to the following points:
[0160] Item 1. Method (1000) for creating an RNase-free and / or DNase-free surface, in particular an RNase-free and / or DNase-free polymer surface, the method comprising:
[0161] generating (1100) an ozone-dominated non-thermal plasma (10) under atmospheric pressure;
[0162] Generating (1200) a nitrogen-dominated non-thermal plasma (20) under atmospheric pressure;
[0163] Generating (1300) a plasma flow (26) from the ozone-dominated non-thermal plasma and the nitrogen-dominated non-thermal plasma; directing (1400) the plasma flow (26) onto a surface (102, 104) to be cleaned, and inactivating (1500) RNase and / or DNase on the surface (102, 104, 106) to be cleaned.
[0164] Item 2. The method (1000) of claim 1, further comprising removing (1600) the inactivated RNase and / or DNase from the surface to be cleaned (102, 104, 106), and optionally after removing (1600) the inactivated RNase and / or DNase from the surface to be cleaned (102, 104, 106), directing (1700) the plasma flow (26) onto the cleaned surface (102, 104).
[0165] Item 3. Method (1000) according to item 1 or 2, wherein the ozone-dominated non-thermal plasma (10) is generated by means of a dielectric barrier discharge or a corona discharge, and wherein the ozone-dominated non-thermal plasma is optionally generated at a power at a discharge area of below 0.5 W / cm 2 , especially below 0.4 W / cm 2 or below 0.3 W / cm 2 , or below 0.2 W / cm 2 is generated, and / or wherein the nitrogen-dominated non-thermal plasma (20) is generated by means of a dielectric barrier discharge or a corona discharge, and wherein the nitrogen-dominated non-thermal plasma (20) is optionally generated at a power at a discharge area of more than 0.1 W / cm 2 , especially more than 0.2 W / cm 2 , or more than 0.5 W / cm 2 , or more than 1.5 W / cm 2 , or more than 2 W / cm 2is generated, and / or wherein the generation (1100, 1200) of the ozone-dominated non-thermal plasma and the nitrogen-dominated non-thermal plasma takes place simultaneously, preferably in a common plasma generator (110).
[0166] Item 4. Method (1000) according to one of items 1 to 3, wherein the plasma flow (26) is mixed with an aerosol and then directed onto the surface to be cleaned (102, 104).
[0167] Item 5. Method (1000) according to one of items 1 to 4, wherein the surface to be cleaned comprises an inner surface of at least one volume, and wherein the at least one volume is formed in particular by a polymer film bag (42, 44, 46) or by a decontamination container (50), wherein the plasma flow (26) is guided into the volume at at least one inlet.
[0168] Item 6. Method (1000) according to one of items 1 to 5, wherein the method further comprises a leak test of the at least one volume, wherein a plasma gas of the generated plasma flow (26) is used as the test gas.
[0169] Item 7. The method (1000) according to any one of items 1 to 6, wherein the method further comprises unwinding a polymer film (60, 62) and / or extruding a polymer film (60, 62), and wherein the plasma flow (26) is directed onto at least one side of the extruded and / or unwound polymer film (60, 62).
[0170] Item 8. Method (1000) according to one of items 1 to 7, wherein the plasma flow (26) is directed onto the surface to be cleaned (102, 104, 106) for a period of at least 10 minutes, or at least 15 minutes, or at least 20 minutes. Item 9. Method (1000) according to one of items 1 to 8, wherein the method further comprises producing at least one volume, in particular at least one polymer film bag (42, 44, 46), wherein the volume is made from at least one polymer film (60, 62), and producing the at least one volume comprises polymer bonding and / or polymer welding, and wherein an RNase-free and / or DNase-free inner surface of the volume is created before and / or after the polymer bonding and / or polymer welding.
[0171] Point 10.Device (100) for creating an RNase-free and / or DNase-free surface, in particular an RNase-free and / or DNase-free polymer surface, wherein the device (100) is configured to carry out a method (1000) according to one of items 1 to 9, and comprises the following: at least one plasma reactor (110, 112) for generating an ozone-dominated non-thermal plasma (12) under atmospheric pressure; at least one plasma reactor (110, 114) for generating a nitrogen-dominated non-thermal plasma (14) under atmospheric pressure; at least one plasma gas line (126) which is connected to the plasma reactors (100, 112, 114) and is designed to direct a plasma flow (26) generated from the ozone-dominated non-thermal plasma and the nitrogen-dominated non-thermal plasma onto a surface (102, 104, 106) to be cleaned in order to inactivate RNase and / or DNase on the surface (102, 104, 106) to be cleaned.Item 11. Device (100) according to item 10, wherein the plasma reactor (110, 112) for generating an ozone-dominated non-thermal plasma (12) under atmospheric pressure and the plasma reactor (110, 114) for generating a nitrogen-dominated non-thermal plasma under atmospheric pressure are integrally designed.
[0172] Item 12. Device (100) according to item 10 or 11, wherein the plasma gas line (126) is configured to connect the plasma reactors (100, 112, 114) to at least one decontamination container (50), and / or to at least one volume, in particular at least one polymer film bag (42, 44, 46), and / or to at least one plasma gas nozzle (222, 224, 226), wherein the plasma gas nozzle (222, 224, 226) is optionally directed at at least one polymer film (60, 62), wherein the polymer film (60, 62) is an extruded film, a wound-off film and / or a film to be welded, and wherein the plasma gas nozzle (222, 224, 226) is further optionally configured to be movable.
[0173] Item 13. Device (100) according to one of items 10 to 12, wherein the device (100) forms a closed circuit for the plasma flow (26).
[0174] Item 14. Device (100) according to one of items 10 to 13, wherein the device (100) comprises a polymer welding device (240) which is designed to weld polymer film (60, 62) to form polymer film bags (42), and / or wherein the device (100) comprises an unwinding device (260, 262) for unwinding polymer film (60, 62), and / or wherein the device (100) comprises a decontamination container (50) which is connected to the at least one plasma gas line, and / or wherein the device (100) further comprises an aerosol generator which is designed to generate an aerosol, and wherein the device (100) is designed to mix the generated aerosol with the plasma flow (26) and to direct it onto a surface (102, 104, 106) to be cleaned.
[0175] Item 15. Polymer film bag (42, 44, 46) for use in a bioreactor, wherein the polymer film bag (42, 44, 46) is produced according to a method (1000) according to item 9, and whose inner surface is free of RNase and / or DNase.
Claims
Claims 1. A method (1000) for creating a nuclease-free surface, characterized by the steps: Generating (1100) a first non-thermal plasma (10) under atmospheric pressure, wherein the first non-thermal plasma (10) is generated at a discharge surface with a first power, Generating (1200) a second non-thermal plasma (20) under atmospheric pressure, wherein the second non-thermal plasma (20) is generated at a discharge surface with a second power, wherein the second power is higher than the first power, Generating (1300) a plasma flow (26) from the first non-thermal plasma and the second non-thermal plasma; Directing (1400) the plasma flow (26) onto a surface to be cleaned (102, 104), and Inactivation (1500) of nucleases on the surface to be cleaned (102, 104, 106).
2. The method (1000) of claim 1, further comprising removing (1600) the inactivated nucleases from the surface to be cleaned (102, 104, 106), and optionally after removing (1600) the inactivated nuclease from the surface to be cleaned (102, 104, 106), directing (1700) the plasma flow (26) onto the cleaned surface (102, 104).
3. Method (1000) according to claim 1 or 2, characterized in that the ozone-dominated non-thermal plasma (10) is generated by means of a dielectric barrier discharge or a corona discharge.
4. Method (1000) according to one of claims 1 to 3, characterized in that the first non-thermal plasma is generated at a power at the discharge surface of 0.2 W / cm 2 up to < 0.45 W / cm 2 , preferably 0.3 W / cm 2 up to < 0.45 W / cm 2 is generated, wherein the second non-thermal plasma (20) at a power at the discharge surface of > 0.45 W / cm 2up to 0.7 W / cm 2 , preferably > 0.45 W / cm 2 up to 0.6 W / cm 2 is generated.
5. Method according to one of claims 1 to 4, characterized in that the generation (1100, 1200) of the first non-thermal plasma and the generation of the second non-thermal plasma take place simultaneously.
6. Method (1000) according to one of claims 1 to 5, characterized in that an aerosol is further generated, wherein the plasma flow (26) is mixed with the aerosol and is then directed onto the surface to be cleaned (102, 104).
7. Method (1000) according to one of claims 1 to 6, characterized in that the surface to be cleaned is located in the interior of a hollow body.
8. The method (1000) according to claim 7, characterized in that the hollow body is a polymer film bag (42, 44, 46) or a decontamination container (50).
9. The method (1000) according to claim 7 or claim 8, wherein the method further comprises a leak test of the at least one hollow body, wherein a plasma gas of the generated plasma flow (26) is used as the test gas.
10. The method (1000) of any one of claims 1 to 9, wherein the method further comprises unwinding a polymer film (60, 62) or extruding a polymer film (60, 62), and wherein the plasma flow (26) is directed onto at least one side of the extruded or unwound polymer film (60, 62).
11. The method (1000) according to any one of claims 1 to 10, wherein the plasma flow (26) is directed onto the surface to be cleaned (102, 104, 106) for a period of at least 10 minutes, or at least 15 minutes, or at least 20 minutes.
12. The method (1000) according to any one of claims 7 to 11, wherein the method comprises Creating the hollow body comprises, wherein the hollow body is made from at least one polymer film (60, 62), and creating the at least one hollow body comprises polymer bonding or polymer welding, and wherein a nuclease-free inner surface of the volume is created before and after the polymer bonding or polymer welding.
13. The method (1000) according to any one of claims 1 to 7, characterized in that the surface is part of a pipette tip, a medical or pharmaceutical disposable plastic article, a disposable bioprocess container, a laboratory equipment, a centrifuge or a PCR unit.
14. A device (100) for creating a nuclease-free surface, the device (100) being adapted to carry out a method (1000) according to any one of claims 1 to 13, the device comprising: at least one plasma reactor (110, 112, 114) with at least one discharge surface on a discharge device, wherein a first non-thermal plasma (12) with a first power can be generated at the discharge surface under atmospheric pressure with the plasma reactor (110, 112, 114), characterized in that a second non-thermal plasma (14) with a second power can be generated at the discharge surface under atmospheric pressure with the plasma reactor (110, 112, 114), wherein the second power is higher than the first power; wherein at least one plasma gas line (126) is connected to the at least one plasma reactor (100, 112, 114) and is configured to direct a plasma flow (26) generated from the first non-thermal plasma and the second non-thermal plasma onto a surface (102, 104, 106) to be cleaned in order to inactivate nucleases on the surface (102, 104, 106) to be cleaned.
15. Device (100) according to claim 14, characterized in that the plasma reactor (110, 112, 114) with the discharge surface on the discharge device comprises a dielectric discharge device or a corona discharge device.
16. The device (100) according to claim 14 or claim 15, wherein the plasma reactor (110, 112, 114) is configured such that the first non-thermal plasma and the second non-thermal plasma can be generated simultaneously with a single plasma reactor.
17. Device (100) according to one of claims 14 to 16, characterized in that two plasma reactors (110, 112, 114) are provided, wherein the first non-thermal plasma can be generated with the first plasma reactor (110, 112) and the second non-thermal plasma can be generated with the second plasma reactor (110, 114).
18. Device (100) according to claim 14 or 17, wherein the plasma gas line (126) has a plasma gas nozzle (222, 224, 226) which can be connected to at least one decontamination container (50), or to at least one hollow body, in particular at least one polymer film bag (42, 44, 46).
19. Apparatus (100) according to any one of claims 14 to 18, further comprising an unwinding device (260, 262) for unwinding polymer film (60, 62) and a polymer welding device (240).
20. Device according to one of claims 14 to 19, wherein the device (100) comprises a decontamination container (50) which is connected to the at least one plasma gas line.
21. Device according to one of claims 14 to 20, wherein the device (100) further comprises an aerosol generator configured to generate an aerosol, and wherein the device (100) is configured to mix the generated aerosol with the plasma flow (26) and to direct it onto a surface (102, 104, 106) to be cleaned.
22. Polymer film bag (42, 44, 46) for use in a bioreactor, wherein the A polymer film bag (42, 44, 46) produced according to a method (1000) according to claim 12, and whose inner surface is free of nucleases.