EP4341683A1 - Procédé et dispositif de surveillance de contenu de réacteurs mixtes - Google Patents

Procédé et dispositif de surveillance de contenu de réacteurs mixtes

Info

Publication number
EP4341683A1
EP4341683A1 EP23727812.2A EP23727812A EP4341683A1 EP 4341683 A1 EP4341683 A1 EP 4341683A1 EP 23727812 A EP23727812 A EP 23727812A EP 4341683 A1 EP4341683 A1 EP 4341683A1
Authority
EP
European Patent Office
Prior art keywords
sensor
reactor
measuring arrangement
signal
sensors
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Pending
Application number
EP23727812.2A
Other languages
German (de)
English (en)
Inventor
Konrad Herzog
Julian SIDRAM
Felix Kortmann
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Aquila Biolabs GmbH
Original Assignee
Aquila Biolabs GmbH
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Priority claimed from DE102022002116.1A external-priority patent/DE102022002116B4/de
Application filed by Aquila Biolabs GmbH filed Critical Aquila Biolabs GmbH
Publication of EP4341683A1 publication Critical patent/EP4341683A1/fr
Pending legal-status Critical Current

Links

Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
    • G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
    • C12M41/30—Means for regulation, monitoring, measurement or control, e.g. flow regulation of concentration
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M27/00—Means for mixing, agitating or circulating fluids in the vessel
    • C12M27/02—Stirrer or mobile mixing elements
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
    • C12M41/26—Means for regulation, monitoring, measurement or control, e.g. flow regulation of pH
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N31/00—Investigating or analysing non-biological materials by the use of the chemical methods specified in the subgroup; Apparatus specially adapted for such methods
    • G01N31/22—Investigating or analysing non-biological materials by the use of the chemical methods specified in the subgroup; Apparatus specially adapted for such methods using chemical indicators
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
    • G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
    • G01N2021/7769—Measurement method of reaction-produced change in sensor
    • G01N2021/7786—Fluorescence
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
    • G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
    • G01N2021/7793—Sensor comprising plural indicators
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
    • G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
    • G01N2021/7796—Special mountings, packaging of indicators
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/84—Systems specially adapted for particular applications
    • G01N2021/8411—Application to online plant, process monitoring
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64—Fluorescence; Phosphorescence
    • G01N21/6408—Fluorescence; Phosphorescence with measurement of decay time, time resolved fluorescence

Definitions

  • the invention relates to a method and a device for monitoring the contents of mixed reactors. It is particularly applicable for monitoring the contents of mixed reactors with high demands on sterility and purity as well as in applications with reactors of complex geometry or small size and the associated limited accessibility to measuring devices. Applications of the invention can therefore be found, for example, in the process monitoring of cell cultures or chemical reactions, of processing, purification and formulation processes of pharmaceutical, biological or chemical products as well as in the monitoring of storage processes.
  • Mixed reactors are used in many areas of the chemical, biological and biotechnological industry and research, in particular for carrying out chemical or biochemical reactions and syntheses, for cultivating living cells, for processing, purifying and formulating pharmaceutical, biological or chemical products and for storing a wide range of components such processes.
  • the contents of mixed reactors are monitored in particular for the purposes of research, process development and optimization, process control and regulation as well as for characterization and quality assurance, with various properties of the reactor contents or their correlates being recorded using suitable sensors and measuring arrangements and often recorded over the duration of the process.
  • sensors that contain at least one sensory component, which is part of at least one sensor located inside the reactor the property of the reactor contents to be monitored reacts and, depending on this and possibly via intermediate steps of other sensory components, generates a signal that can be detected by a suitable measuring arrangement.
  • a variety of approaches are known to the person skilled in the art, in particular, but not exclusively, the sensory use of the dependence of optically detectable properties (e.g.
  • the combination of suitable sensors and measuring arrangements therefore fundamentally enables the contents of mixed reactors to be monitored, with the properties to be monitored being converted by suitable sensors inside the reactor into a signal that can be detected electronically by the measuring arrangement.
  • the reactor contents consist of several phases (for example liquid or liquid-like reaction mixture and gas phase above it in the headspace of the reactor), monitoring the properties of individual phases is often important in the application (for example oxygen concentration in the reaction mixture, oxygen concentration in the gas phase of the headspace).
  • the above-mentioned review articles describe the use of optically detectable sensory components, with at least one of their optical properties changing depending on the property of the reactor contents to be monitored (e.g. pH or oxygen concentration).
  • the optical detection is carried out using suitable measuring arrangements, which usually have at least one light source and at least one light detector.
  • suitable measuring arrangements which usually have at least one light source and at least one light detector.
  • the two overview articles reveal different sensor formats and the associated concepts and designs of the optical measurement arrangements.
  • Layer-like sensor formats are disclosed which are applied to the inside of the reactor wall, to the inside of optical windows of immersion probes, or to optical fibers and other totally reflective light guides in contact with the reactor contents.
  • the associated measuring arrangement is in optical contact with the sensors and the sensory components they contain.
  • micro- and nanoparticulate sensor formats are disclosed.
  • these particles are embedded in polymers and are stationary, comparable to the aforementioned layer-like sensors applied to the inside of the reactor, with the associated optical measuring arrangement also stationary.
  • these particles are added to the reactor contents and either concentrated locally in the optical field of view of the measuring arrangement by magnetic interactions, or added in such a high concentration that such magnetic concentration is not necessary to achieve sufficiently strong optical signals.
  • the article shows a serious disadvantage of sensors installed stationary in the reactor for applications in which the sensor is not permanently in contact with the reaction mixture.
  • the sensor reacts to both properties of the reaction mixture and to properties of the gas phase in the headspace of the reactor, resulting in a mixed signal that deviates from the actual signal of the respective phase.
  • DE10101576A1 discloses an optical sensor for determining at least one parameter in a sample, which has an indicator material with a short decay time that responds to the parameter and a reference material that does not respond to the parameter contains a long decay time and is used to record a measurement signal indicating the parameter to be determined based on the jointly recorded luminescence responses of the indicator and reference material.
  • the indicator and reference materials are immobilized on a common support and in contact with the reactor contents.
  • the carrier can be a film, a cassette through which the sample flows, or a flat or fiber-like light guide. In the case of several sensors, the sensors can be arranged side by side on the carrier, separated from one another, or they can also be mixed together.
  • EP1397672A1 discloses a device for detecting oxygen, comprising a carrier with a plurality of wells for receiving samples, with stationary oxygen sensors located on the inside of the wells being provided, comprising particles which contain a luminescent, oxygen-erasable dye and a gas-permeable and essentially water-impermeable first matrix contain, and a substantially water-permeable second matrix, the particles being dispersed in the second matrix.
  • the device is designed as a microtiter plate and as a culture plate for cultivating cells.
  • EP3821230A1, DE102018105174A1, EP2321052A1 and US2008171383A1 disclose devices and methods for monitoring the contents of various reactor types (channels, bags, stirred tanks), the sensors being mounted in a stationary manner on a sensor carrier and this sensor carrier being fastened in the reactor wall in such a way that the sensor is connected to the Reactor contents come into contact.
  • the optical properties to be recorded of the sensors installed in a stationary manner in the reactor wall are recorded via suitable stationary optical measuring arrangements or via correspondingly stationarily mounted optical fibers as components of suitable optical measuring arrangements.
  • the methods and devices known from the prior art for monitoring the contents of mixed reactors disclose sensors which, via suitable sensory components, convert at least one property of the reactor contents to be monitored into a signal that can be detected electronically by the measuring arrangement.
  • the vast majority of these methods and devices use devices that are permanently connected to the inside of the reactor wall or the reactor internals Sensors in combination with stationary measuring arrangements inside or outside the reactor to record the sensor signal.
  • Some disadvantages of stationary sensors can be avoided by using micro- and nanoparticles in the reaction mixture, which is known from the prior art. These are either freely and evenly distributed in the reactor contents or reaction mixture or they are magnetically focused in the detection area of the measuring arrangement for signal detection and therefore appear to the measuring arrangement like a stationary sensor in both cases. In comparison to the sensor that is fixed in place on the inside of the reactor wall, there is a clear phase assignment of the sensor particles However, this does not lead to the incorrect detection of mixed signals of the properties to be monitored from different phases of the reactor contents described above. Furthermore, the homogeneous distribution of the sensor particles in the reactor contents or reaction mixture, as well as the magnetic focusing of the sensor particles in front of the measuring arrangement, simplifies both the introduction of the sensors into the reactor contents and the positioning and orientation of the measuring arrangement.
  • DE102004017039A1 discloses a method and a device for recording process parameters of reaction liquids in several microreactors, which are continuous at least until the end of the reaction in all microreactors be shaken.
  • the process parameters in the microreactors are recorded during the reaction using at least one sensor lens as part of the measuring arrangement.
  • the sensor optics can be moved via a positioning unit.
  • the reaction liquids in the microreactors have at least one chemical sensor material and the chemical sensor material is attached to at least one inner surface of the microreactor.
  • each sensor optic is not moved, at least while the process parameters are being recorded, so that the shaken microreactors move relative to each sensor optic.
  • embodiments are also disclosed in which a flash lamp is used as the radiation source of the sensor optics, the pulse frequency of which is coordinated with the shaking movement in such a way that the flashes of light always hit the microreactor at the same point during the shaking movement.
  • An embodiment is also disclosed in which many measuring points are recorded using a high flash and detection frequency, which is above the shaking frequency of the shaker, in order to prevent the measurement signal from beating.
  • the overall disclosed combination of movable sensor optics as a component of a measuring arrangement, chemical sensors fixed in place on the inner wall of the reactor and a pulsed radiation source of the sensor optics synchronized to the shaking movement basically allows the detection of several stationary sensors, even using identical optical sensory components and at the same time Miniaturization of the reactor.
  • the object on which the invention is based is achieved by a method according to claim 1 and a device according to claim 8; Preferred embodiments result from the subclaims and the description.
  • a reactor is a container that can be used in particular to cultivate organisms or to carry out chemical and biochemical reaction processes. Further areas of application of reactors include biocatalytic processes using organisms and/or biomolecules as well as other chemical and/or physical processes, whereby the term process includes all types of conversion, separation, combination, mixing, size change and storage of, in particular, chemical substances and organisms , particles, solutions, emulsions and foams.
  • Reactors within the meaning of the invention include, in particular, stirred tank fermenters and reactors, bubble column fermenters, shake flasks, T-flasks, microtiter plates, deep well plates, shaking barrels, fermentation bags, multi-purpose tubes and cell culture dishes. Reactors can be closed or open to their environment.
  • the reactor contents include all matter that is located within the outer shell of the reactor. In the case of a reactor that is open to the environment, the reactor contents are limited by the inner wall of the reactor and by the hypothetical closure surface, which represents the transition between the inside of the reactor and the environment.
  • the reactor contents are composed of one or more phases, which are in particular designed as fluids (gases, liquids) or as phase mixtures with a fluidic character (foams, emulsions, suspensions, powder beds).
  • the reactor contents are formed by two phases, by the reaction mixture in which the majority of the process to be carried out takes place, and by the headspace above, which is usually in the form of a gas phase.
  • each phase of the reactor contents can be a pure substance or a mixture of substances.
  • the reactor contents and therefore in particular the reaction mixture and the headspace have physical, chemical, biological or other properties that can be the target of monitoring.
  • the sensor-based monitoring of one or more properties of the reactor contents, the reaction mixture, the headspace or their components can be carried out by detecting and determining the property itself or by detecting correlates of the property.
  • correlates are any phenomena, processes, signals, properties or environmental conditions of suitable sensors, sensory components or converting components that correlate with the property to be monitored.
  • Properties within the meaning of the invention are qualitative or quantitative quantities that are suitable for describing the state of a thing, in particular but not exclusively substance and particle concentrations, environmental parameters such as temperature and pressure, optical parameters such as luminescence lifetime, emission, absorption or scattering intensity and -wave lengths, other physical parameters such as emissivity, impedance, electrical capacity, inductance and conductivity, biological parameters such as expression rates, metabolic pathway activities, division rates or viability and many more.
  • the monitoring of properties of the reactor contents or its components can also be carried out via the individual or joint detection of several correlates and their combination with suitable mathematical calculation and evaluation methods or with other algorithms.
  • Mixing the reactor contents refers to any method of influencing the reactor contents in such a way that at least two states of the distribution of the reactor contents and its components in the reactor recorded at different times are not the same.
  • Common mixing processes use in particular mechanical, thermal or thermodynamic processes, in particular but not exclusively shaking processes, stirring processes, pumping processes and diffusion processes.
  • Sensors in the sense of the invention are devices that are suitable for converting at least one property of the reactor contents or at least one of its components into at least one signal, which can be detected by suitable measuring arrangements if necessary by means of excitation.
  • sensors in the sense of the invention include at least one sensory component which has at least one property or a corresponding correlate, which can be detected by the measuring arrangement and is influenced by the property of the reactor contents to be monitored or at least one of its components.
  • the influence by the at least one property to be monitored can occur directly or indirectly, in particular through interaction chains with various converting components or with other properties or components of the reactor contents.
  • properties or correlates are recorded by measuring arrangements via at least one signal of at least one sensory component, which in some cases has to be stimulated by the measuring arrangement.
  • Sensory components within the meaning of the invention are in particular, but not exclusively, dyes, fluorophores, luminophores, nano or microparticles made of metals or semiconductors and other structures that can generate or influence optical or other electromagnetic signals, as well as combinations of the aforementioned or similar components with one another and systems that contain at least one of the aforementioned components.
  • Sensors within the meaning of the invention can also contain one or more converting components.
  • a converting component in the sense of the invention has at least one property or a corresponding correlate, which is determined by the property of the reactor contents to be monitored or at least one of its components is influenced directly or indirectly (via another converting component), although it cannot itself be detected by a measuring arrangement, but directly or indirectly (via another converting component) at least one property and thus at least one signal at least one is in the environment can influence the sensory component located in the converting component.
  • Converting components in the sense of the invention can be linked via their interactions with one another or with sensory components to form interaction chains of any length in order to convert changes in at least one property of the reactor contents or at least one of its components into a signal that can be detected by at least one measuring arrangement.
  • Converting components in the sense of the invention advantageously have a high specificity with respect to the property to be monitored or the interacting other converting components or sensory components and are used in particular to monitor substance concentrations.
  • Converting components within the meaning of the invention are in particular enzymes, catalysts, nucleic acids, aptamers, ribozymes, antibodies and other selectively binding proteins or structures, as well as combinations of the aforementioned or similar components with one another and systems that contain at least one of the aforementioned components.
  • the sensory component influenced by at least one converting component is preferably in spatial proximity to the converting component influencing it, in particular in order to ensure an effective interaction between the two, for example through local changes in substance concentrations (in particular oxygen or proton concentration) or through energy To enable electron or proton transfer processes (in particular quenching, FRET, PET, electroluminescence, reactions).
  • Sensors within the meaning of the invention can further contain markers that generate at least one marker signal, so that in particular the location, position and identity of the respective sensor can be determined by suitable measuring arrangements via the detection of the at least one marker signal.
  • Sensors within the meaning of the invention can have at least one sensor matrix, which, as a structural component, provides mechanical stability to maintain the sensor-internal arrangement of sensory components, converting Components and markers, can embed them in the sensor, can separate them from each other and can thus segment the sensor.
  • a sensor matrix can also be used to give the sensor a specific shape and maintain it.
  • sensor matrices in the sense of the invention can be used to set selective mass transport, for example through defined pore sizes, polarity or surface charge.
  • Sensor matrix in the sense of the invention includes, in particular but not exclusively, all types of polymers, hydrogels, membranes and amorphous, semi-crystalline or crystalline solids. Several identical or different sensor matrices can be used in one sensor.
  • a measuring arrangement within the meaning of the invention is any device or combination of devices that is suitable for detecting at least one signal of at least one sensory component or at least one marker signal of at least one marker.
  • every property that can be detected by a measuring arrangement or its correlate of a sensory component or a marker is a signal or marker signal in the sense of the invention.
  • a measuring arrangement comprises at least one signal detector. If the signal or marker signal to be detected needs to be excited, a suitable measuring arrangement also includes at least one signal exciter.
  • Signals to be excited within the meaning of the invention are, in particular, but not exclusively, optical signals such as luminescence, absorption and scattering intensities, degrees of luminescence polarization, luminescence lifetimes and impedance signals such as capacity, inductance or conductivity.
  • Signals can be recorded depending on frequency or time. Signals can be acquired by modulating the signal excitation. Derived signals (e.g. spectra, ratiometric signals, decay curves, Bode plots, time series) can be formed from individual recorded signals (e.g. intensities).
  • suitable electrical circuits or computers with suitable software and algorithms can be used, which are part of the corresponding measuring arrangement within the meaning of the invention.
  • a computer is any electronic device that can store data (especially arithmetic and logical) and process it based on programmable rules.
  • Computers in the sense of the invention are particularly, but not exclusively, considered to be computers Microcontrollers, microprocessors, system-on-a-chip computers (SoC), PCs and servers.
  • SoC system-on-a-chip computers
  • Signals that can be detected by suitable measuring arrangements are detected in the detection area of the measuring arrangement, the detection area resulting from the detection area of at least one signal detector and, in the case of a required signal excitation, the excitation area of at least one signal exciter.
  • a measuring arrangement detects all signals that can be detected by it in its detection range, i.e. not only the signals of sensory components or the marker signals of markers, but also background and environmental signals, which can be eliminated if necessary using suitable methods.
  • measuring arrangements in the sense of the invention also detect signals, in particular environmental and background signals, when no sensor is currently located in their detection area.
  • the dwell time in the sense of the invention is the time in which a specific sensory component of a sensor is in the detection range of a measuring arrangement that can detect its signal.
  • the acquisition time in the sense of the invention is the time required by a measuring arrangement to capture a specific signal exactly once.
  • a signal detector in the sense of the invention is any device or combination of devices that is suitable for detecting and digitizing at least one signal of at least one sensory component or at least one marker signal of at least one marker.
  • Signal detectors in the sense of the invention therefore include in particular but not exclusively electrodes, antennas, photodiodes, phototransistors, photoresistors, CCD and CMOS arrays, optical elements such as filters, gratings, lenses, fibers and diaphragms, but also amplifier and signal shaping circuits, analogue Digital converters, computers and combinations of all of the aforementioned elements.
  • a signal exciter in the context of the invention is any device or combination of devices that is suitable for stimulating at least one signal of at least one sensory component or at least one marker signal of at least one marker.
  • Signal exciters in the sense of the invention therefore include in particular but not exclusively electrodes, antennas, LEDs, flash lamps, lasers, optical elements such as filters, gratings, lenses, fibers and diaphragms, but also driver and modulation circuits, digital-to-analog converters, computers and combinations all of the aforementioned elements.
  • the addition of at least one sensor according to the invention into a reactor is any targeted movement of at least one sensor from outside the reactor into the interior of the reactor, in particular into the reactor contents or at least one of its components, such as into a reaction mixture or the head space.
  • a storage medium within the meaning of the invention is any substance or any mixture of substances with which at least one sensor according to the invention is specifically brought into contact or held during its storage or other preservation before being added to a reactor in order to specifically target certain properties or behaviors of the at least one sensor influence or maintain.
  • Storage media according to the invention are in particular but not exclusively air, defined gas mixtures, protective gases, aqueous solutions, buffer solutions, salt solutions, mixtures of organic and aqueous components, emulsions, foams and also powders.
  • Fins in the sense of the invention can be components of sensors according to the invention.
  • a fin in the sense of the invention is any device that is suitable for specifically influencing the movement behavior of sensors according to the invention in the reactor contents or at least one of its components or the flow behavior of fluids inside the reactor on sensors according to the invention.
  • Fins in the sense of the invention are therefore particularly but not exclusively fluid-mechanical structural elements, such as fins, notches, gaps, grooves and perforations, but also buoyancy elements such as weights, cavities, foams or other closed-porous structures.
  • An agitator in the sense of the invention is any device in or on the reactor that can either be set in motion itself or set the reactor in motion in order to achieve a mixing movement, in particular convective mass transport reactor contents, or in at least one of its components, for example in the reaction mixture or in the headspace.
  • the problem is solved for monitoring the contents of mixed reactors, the reactor contents having at least one property to be monitored and the at least one property to be monitored influencing at least one signal of at least one sensory component and the at least one signal of the at least one sensory component being influenced by at least one Measuring arrangement is detected by a method in which at least one sensor, which contains the at least one sensory component, is not stationary and moves in the reactor, so that the at least one sensor is not permanently in the detection range of the at least one measuring arrangement and at least one detection of the at least one signal of the at least one sensory component takes place through the at least one measuring arrangement, while the at least one sensor is in the detection range of the at least one measuring arrangement.
  • At least one signal of the at least one sensory component which was detected while the at least one sensor was in the detection range of the at least one measuring arrangement, is used to monitor or determine the at least one property of the reactor contents to be monitored. According to the invention, this monitoring can also be carried out by detecting correlates of the property to be monitored.
  • the sensors according to the invention are not evenly distributed in the reactor contents or reaction mixture, so that the sequential or parallel detection of several sensors with different sensory components is made possible by the same or by several suitable measuring arrangements, without the sensors according to the invention influencing or interfering with each other with regard to their detection by suitable measuring arrangements.
  • the method according to the invention allows for non-uniform movement in the reactor Distributed sensors enable more cost-effective monitoring of larger reactors, since larger quantities of sensors do not have to be added to the reactor contents to achieve detectable measurement signals, as is required for micro- and nanoparticle sensors.
  • the inventive use of non-uniformly distributed sensors that move in the reactor enables a sequential detection of the signals from the sensory components located in the sensors, so that, in contrast to the stationary sensors known from the prior art, the detection of different signals from different sensors and sensor components for monitoring various properties of the reactor contents is possible with a reduced number of measuring arrangements, and such monitoring can also be carried out on miniaturized reactors.
  • the method according to the invention also simplifies the use of sensors in the reactor, since the sensors according to the invention can simply be added while the reactor is being filled and do not have to be previously applied to the inner walls or other internals of the reactor, as is the case with stationary sensors .
  • the volume of at least one detection area of at least one measuring arrangement overlapping the reactor contents is smaller than the volume of the reactor contents.
  • the volume of each detection area of each individual measuring arrangement that overlaps with the reactor contents is smaller than the volume of the reactor contents, but in particular so small that the state of the method according to the invention can be achieved for every sensor used and detectable by the relevant measuring arrangement in which the respective sensor is not in the detection range of the relevant measuring arrangement.
  • the detection range of each individual measuring arrangement is less than or equal to the volume of the reactor contents or reaction mixture minus the volume of all sensors that can be detected by the respective measuring arrangement, which are currently in the reactor or reaction mixture and enter the detection range of the respective measuring arrangement through movement could.
  • the detection range of each individual measuring arrangement is at least so large that the smallest sensor to be detected by the respective measuring arrangement can still be detected by it.
  • At least one detection area of at least one measuring arrangement is smaller than at least one of the sensors that can be detected by the measuring arrangement.
  • each detection area of each measuring arrangement is smaller than the smallest sensor that can be detected in the corresponding detection area.
  • the detection range of each measuring arrangement is smaller than the smallest contiguous sensor area within a sensor that can be detected in the corresponding detection range and contains the same type of sensory components.
  • the at least one signal or several signals or the entirety of the signals of a sensory component that can be detected by at least one suitable measuring arrangement can be detected even if the sensor does not completely fill the detection area of the at least one measuring arrangement.
  • signals according to the invention are in particular, but not exclusively, ratiometric signals as well as lifespans and decay times.
  • each sensor is at least as large as or larger than the detection range of the at least one measuring arrangement, which detects the at least one signal of the at least one sensory component contained in the sensor.
  • the detection time that a measuring arrangement requires to detect at least one signal of a sensory component is shorter than the dwell time of the sensor containing the sensory component in the detection area of the measuring arrangement.
  • the detection time that a measuring arrangement requires to detect at least one signal of a sensory component is shorter than the residence time of the segment of a sensor containing the sensory component in the detection area of the measuring arrangement. This applies particularly advantageously to sensors that contain several sensory components arranged in segments.
  • a sensor according to the invention or several sensors according to the invention are passively moved by the movement of the reactor contents or reaction mixture that occurs as part of the mixing of the reactor and thereby reach the detection area of at least one measuring arrangement and also out of the detection area of at least one measuring arrangement.
  • shaken reactors for example shake flasks, bags, culture plates, microtiter plates, T-flasks
  • stirred tank reactors for example shake flasks, bags, culture plates, microtiter plates, T-flasks
  • bubble column reactors for example shake flasks, bags, culture plates, microtiter plates, T-flasks
  • At least one sensor according to the invention contains at least one marker, which can be detected by at least one measuring arrangement via at least one marker signal.
  • the marker uses its marker signal to trigger the start of the detection of at least one signal of at least one sensory component by at least one measuring arrangement.
  • sensors that are used to monitor different properties are also equipped with markers that have different marker signals, in order in particular, but not exclusively, to trigger different detection methods of the measuring arrangement or to uniquely identify the different sensors in each case to be able to assign the characteristic of the reactor contents monitored by them.
  • At least one property that can be detected by suitable measuring arrangements is at least a sensory component in a sensor is used as a marker.
  • the luminescence intensity of a luminophore whose luminescence lifetime is recorded as a signal can be recorded as a marker signal, so that a suitable measuring arrangement always records the luminescence lifetime when a luminescence marker signal is present.
  • a measuring arrangement can detect different signals from different sensory components and/or different marker signals from different markers. This is done in particular by combining various suitable signal detectors and, where necessary, signal exciters.
  • At least one measuring arrangement is attached to the reactor in a stationary manner, so that there is no relative movement between the reactor and the measuring arrangement. In other embodiments of the invention, a relative movement takes place between the reactor and at least one measuring arrangement.
  • At least one sensor according to the invention is added to the reactor when the reactor is filled. In some embodiments of the invention, at least one sensor according to the invention is added to the reactor before the start of the process to be carried out in the reactor, for example before the start of the cultivation or incubation of cells or before the start of a chemical reaction. In other embodiments of the invention, at least one sensor according to the invention is added to the reactor before the reactor is packaged or sterilized or during the process already running in the reactor. In versions that require high sterility or purity, the addition takes place from a sterile addition system for sensors according to the invention, which are then also sterile.
  • the sensors according to the invention are stored in a storage medium before being added to the reactor, which ensures the durability of the sensors and keeps the sensors ready for use.
  • each sensor can be adapted to the shape and type of mixing of a reactor or its reactor contents.
  • sensors in the form of ring segments or shapes derived therefrom can enable optimal movement of the sensors in the reactor to match the radii of the reactors used or orbital movements.
  • spherical or ellipsoidal sensors can be used advantageously.
  • the sensors according to the invention have shaped elements that promote optimal alignment or movement of the sensors in the reactor contents or reaction mixture.
  • shaped elements are in particular, but not exclusively, wing-, fin- or fin-like structures.
  • At least one sensory component or at least one converting component or at least one sensor matrix which contains at least one sensory component or at least one converting component, is set back or inward relative to the outer contour of the sensor in such a way that in the event of a collision or other contact between the sensor in question and the reactor (in particular the reactor wall or reactor internals), there is no direct contact between the reactor and the at least one sensory component, converting component or sensor matrix placed back or inwards.
  • a sensor has a geometry that specifically hinders or prevents certain forms of movement in the reactor.
  • this is achieved by corners or edges in the geometry of the sensor, so that rolling movements are hindered and a correspondingly designed sensor can be used under a wide variety of mixing conditions (in particular with regard to shaking conditions, fill level of the reactor and viscosity of the reaction mixture ) preferably moves with the reaction mixture instead of rolling or sliding over the reactor floor.
  • the corners and edges are arranged and combined with suitable transitions in such a way that a Overall, an advantageous flow condition results, which also prevents undesirable rolling or sliding movements, stabilizes the position of the sensor in the reaction mixture and supports the movement of the sensor with the reaction mixture.
  • the density of sensors according to the invention can be adapted to the density of the reactor contents or the reaction mixture in accordance with the monitoring objective in order to monitor properties of the reactor contents or reaction mixture in certain regions, for example at the bottom or on the surface of the reaction mixture.
  • sensors of the same type and characteristics can be used in a reactor. This can be done in particular, but not exclusively, in large reactors in order to shorten the time between the detection of at least one signal of one or more specific sensory components by more frequent entry of corresponding sensors with these same sensory components into the detection range of suitable measuring arrangements and thus a closer monitoring of at least one property of the reactor contents.
  • Sensors according to the invention can include one or more converting components that do not emit a signal that can be detected by measuring arrangements, but that interact with at least one sensory component and thus influence at least one property that can be detected by a measuring arrangement and thus at least one signal of the sensory component.
  • different sensors according to the invention each with different converting components, contain the same sensory component as well as different markers in order to reduce the number or complexity of the required measurement arrangements.
  • multiple different signals from one or more sensory components or one or more combinations of at least one sensory component and at least one converting component are used to monitor or determine a property of the reactor contents determine.
  • sensors according to the invention are used which are divided into segments, with different segments each containing different combinations of at least one marker or at least one sensory component and none, one or more converting components.
  • the segments are in a defined order and a defined distance, so that only one marker is required to trigger the sequential detection of the signals of the sensory components from the respective segments of the sensor.
  • each segment is larger than the detection range of the sensor.
  • at least one marker is located between adjacent segments or on at least one outside of a sensor according to the invention.
  • the movement of sensors according to the invention in the reactor contents or reaction mixture occurs periodically or approximately periodically, so that signals from the respective sensory components can be detected at regular intervals by suitable measuring arrangements.
  • the detection of signals and marker signals can advantageously be synchronized with the movement periodicity, in particular but not exclusively on the basis of previously detected signals and periods calculated therefrom or on the basis of the detection of periodic movement signals, for example accelerations, rotation rates or positions or distributions or movement behavior the reactor contents or reaction mixture.
  • the detection of at least one signal of at least one sensory component or at least one marker signal occurs periodically.
  • the detection can advantageously take place at a frequency that is higher than the mixing frequency. Examples of mixing frequencies are stirrer speeds, shaking or Rocking frequencies and volume flows.
  • the detection of at least one signal of at least one sensory component or at least one marker signal is carried out non-periodically, in particular partially or completely randomized, in order to avoid beats, oscillations, aliasing and other acquisition time-dependent phenomena of at least one detected signal of at least one sensory component or at least of a marker signal to minimize or eliminate.
  • the reactor has devices that influence the movement of sensors according to the invention, in particular in order to influence, increase or reduce the probability of presence of sensors according to the invention in certain areas of the reactor or reactor contents.
  • flow elements, baffles, flaps, fins, grooves, channels, nets, or targeted fluid flows are used in particular, but not exclusively, to influence the movement of sensors according to the invention.
  • several signals from sensory components and/or marker signals from one or more sensors are recorded simultaneously or overlapping in time by several different measuring arrangements or by several different signal detectors or signal exciter-signal detector pairs on the same measuring arrangement.
  • several recorded signals are calculated using suitable algorithms and software on computers to form new signals or correlates, which are then used to monitor at least one property of the reactor contents or reaction mixture.
  • various converting components or sensory components or combinations of both are mixed together or otherwise embedded in close proximity to one another in a common segment of a sensor.
  • such mixtures are made from various converting components or sensory components or combinations of both are recorded together by a suitable measuring arrangement, for example in the case of self-referenced sensors as part of dual-lifetime referencing or to compensate for ambient temperature or other environmental properties and parameters.
  • the measuring arrangement is located outside the reactor and its detection area penetrates the reactor wall and part of the reactor contents.
  • the measuring arrangement is designed as an immersion probe and is therefore located inside the reactor.
  • the measuring arrangement is located completely within the reactor and moves with or against the reactor contents.
  • the at least one sensory component and its signal that can be detected by at least one measuring arrangement can be influenced directly or indirectly by the at least one property to be monitored, in particular also by interaction chains with converting components or with other properties or components of the reactor contents.
  • the same property of the reactor contents or reaction mixture to be monitored is recorded redundantly by several different combinations of sensory components and, if necessary, converting components and, if necessary, different measuring arrangements. This redundancy advantageously increases the robustness and quality of the monitoring of the property to be monitored.
  • At least one sensor according to the invention is constructed in layers.
  • at least one sensor according to the invention has a core-shell structure, wherein a core consisting of at least one sensor matrix functions as a carrier of at least one active shell, the active shell having at least one sensory component or at least one converting component or at least one Includes markers or combinations of the aforementioned.
  • such a core can also have at least one sensory component or at least one converting component or at least one marker or combinations thereof include the aforementioned or even be constructed in layers.
  • At least one sensor according to the invention is constructed in such a way that, during the residence time of the sensor in the detection range of at least one measuring arrangement, at least one sensory component or at least one converting component or at least one marker or a combination of the aforementioned is carried out largely or completely independently of its orientation the measuring arrangement can be detected or excited or excited and detected.
  • the sensor is therefore designed to be rotationally symmetrical.
  • At least one sensor matrix or at least one of its components or at least one component embedded by it is prepared in such a way that background signals from the environment, in particular but not exclusively ambient light, electromagnetic fields or other radiation, are shielded.
  • Examples of such embodiments may include, in particular but not exclusively, optically opaque or selectively permeable layers or dyes, scattering or absorption components and materials with suitable permeability or permittivity.
  • At least one sensor matrix is arranged such that it amplifies at least one signal of at least one sensory component or at least one marker signal or at least one signal excitation, in particular but not exclusively by means of scattering or reflection layers or scattering or reflection particles introduced into at least one sensor matrix .
  • Figure 1 a schematic representation of the method according to the invention using two sensors 5 with different sensory components 7.
  • FIG 2 a schematic representation of a sensor 5 according to the invention and a device according to the invention for carrying out the method according to the invention using the example of a T-flask as a reactor 1 for cultivating cells.
  • FIG. 3 a schematic representation of a sensor 5 according to the invention for carrying out the method according to the invention using the example of a shake flask as a reactor 1 for cultivating cells with an addition system 18 for sensors 5.
  • FIG 4 a schematic representation of carrying out the method according to the invention on a shake flask as a reactor 1 for cultivating cells using the sensor 5 according to the invention shown in Figure 3.
  • FIG. 5 a schematic representation of two sensors 5 according to the invention and a device according to the invention for carrying out the method according to the invention on a stirred tank reactor as reactor 1.
  • Figure 6 a schematic representation of a sensor 5 according to the invention in a core-shell design with adapted geometry for use in orbitally shaken systems.
  • Figure 1 shows a schematic representation of the method according to the invention using two sensors 5 with different sensory components 7.
  • the method according to the invention is carried out on a reactor 1, the reactor contents 2 of which are mixed as a result of at least one mixing movement 25.
  • the two sensors 5A and 5B are used to monitor two properties of the reactor contents 2, with each property of the reactor contents 2 influencing a sensory component 7A or 7B in such a way that a signal 8A or 8B emanating from the respective sensory component 7A or 7B passes through the measuring arrangement 6 can be detected.
  • the method according to the invention carried out is characterized in that the sensors 5A and 5B used are not stationary and, as a result of the mixing movement 25, carry out a movement 10A or 10B in the reactor 1, so that the sensors 5A and 5B used are not permanently in the detection area 9 of the measuring arrangement 6, but from time to time, in the case of periodic movements 10 also periodically, stay in the detection area 9 of the measuring arrangement 6 for a certain dwell time 24 and can be detected by this with regard to the signals 8 of their sensory components 7.
  • 23 signals 8 from the detection area 9 are detected by the measuring arrangement 6 at regular intervals over a specific detection time.
  • the embodiment of the method according to the invention shown in Figure 1 uses detection times 23 that are shorter than the dwell time 24 of the respective sensor 5A or 5B. This, as well as in combination with a periodically repeated detection of signals 8 by the measuring arrangement 6, ensures, even without the use of a marker 13, that according to the invention the detection of the respective signals 8A and 8B of the two sensory components 7A and 7B required for monitoring the reactor contents 2 is carried out by the Measuring arrangement 6 takes place while the respective sensor 5A or 5B is in the detection area 9 of the measuring arrangement 6.
  • the measuring arrangement 6 shown is designed in such a way that it can detect different signals 8, but in particular the signals 8A and 8B of the two sensory components 7A and 7B.
  • Figure 1 shows in the upper and middle area four snapshots I, II, III and IV from a continuous mixing movement 25 of the reactor contents 2 in the reactor 1, which over time 21 the non-stationary sensors 5A and 5B detect a movement 10 in the reactor 1 can be carried out.
  • the course of the intensities 22 of the signals 8A and 8B of the two sensory components 7A and 7B recorded by the measuring arrangement 6 is shown schematically over time 21, the times of the four snapshots being given the respective Roman numerals I, II , III and IV are shown.
  • sensor 5B is located in the detection area 9 of the measuring arrangement 6, so that the intensity 22 of the signal 8B is so high that it is used to determine the property of the reactor contents 2 to be monitored by sensor 5B with the sensory component 7B.
  • the signal 8B suitable for monitoring can be identified safely and robustly.
  • the situation of the sensor 5A in the detection area 9 of the measuring arrangement 6 shown in FIGS. 1, III, which here too is used to monitor the property of the sensor to be monitored Reactor contents 2 causes a suitable signal 8A.
  • the respective intensity 22 of the signal 8A or 8B is used as a marker signal 14 for the respective sensor 5A or 5B.
  • FIG. 2 shows a schematic representation of a sensor 5 according to the invention and a device according to the invention for carrying out the method according to the invention using the example of a T-flask as a reactor 1 for cultivating cells.
  • Shown as a side section (top left) and cross section (top right) is a capsule-shaped sensor 5 which contains a sensory component 7 which is enveloped by a converting component 11.
  • the sensory component 7 and the converting component 11 are embedded in a sensor matrix 12.
  • the sensory component 7 generates at least one signal 8 in the detection area 9 of a suitable measuring arrangement 6, which is dependent on at least one property of the reactor contents 2.
  • Sensory components 7 according to the invention which can generate signals 8 according to the invention depending on these properties, are in particular, but not exclusively, dyes and luminophores that generate optical signals 8, but also metallic or semiconductor-derived nano- or microparticulate structures, bioconjugates or systems combined with gels , which can generate capacitive signals 8, more complex, in particular frequency-dependent, impedance signals 8 or magnetic signals 8.
  • any properties of the reactor contents 2 can be monitored according to the invention by using at least one suitable converting component 11 in combination with at least one sensory component 7.
  • the at least one property of the reactor contents 2 to be monitored interacts with at least one converting component 11 and thereby influences at least one property of the sensory component 7 that can be detected by a measuring arrangement 6.
  • Typical examples of such converting components 11 are in particular, but not exclusively, enzymes, antibodies, ribozymes , catalysts, Nucleic acids and their derivatives as well as aptamers, whereby the interaction between the property to be monitored (for example a substance concentration) with at least one converting component n takes place in particular through specific association and dissociation processes or specific, often catalytically influenced, reactions.
  • Such interactions according to the invention between at least one property of the reactor contents 2 to be monitored and at least one converting component 11 influence locally, in particular in the immediate vicinity of the converting component 11, at least one property of the reactor contents 2, which in turn at least one signal 8 of at least one in the interaction area of the converting component 11 sensory component 7 is influenced.
  • the embodiment of the sensor 5 shown in FIG not exclusively oxygen or pH-dependent luminophores or luminophore systems are used as sensory components 7 in combination with oxygen or proton-consuming or producing enzymes and enzyme reactions as converting components 11.
  • Specific binding antibodies or aptamers can also be used as a converting component 11 together with metallic or semiconducting particles or particle systems as a sensory component 7, in particular if the latter has a suitable surface plasmon resonance for optical detection or suitable magnetic, capacitive or resistive properties for, in particular frequency-dependent, Have impedance detection.
  • the sensor matrix 12 shown in Figure 2 not only serves to embed the converting component 11 and the sensory component 7, but also gives the sensor 5 according to the invention mechanical stability. Furthermore, the sensor matrix 12 can also be designed as a partially or completely material-selective structure in order to increase the specificity or sensitivity of the at least one converting component 11 or the at least one sensory component 7.
  • Such versions of the sensor matrix 12 include in particular, but not exclusively, hydrogels and membranes with defined pore size and distribution or with defined polarity, hydrophobicity, oxygen or proton permeability.
  • Figure 2 illustrates the method according to the invention using the example of a T-flask as a reactor 1, the reactor contents 2 of which are formed by a cell culture suspension as a reaction mixture 3 and, above it, by a gas-filled headspace 4.
  • a sensor 5 in the embodiment shown above in Figure 2.
  • the T-flask shown in Figure 2 as a reactor 1 is subjected to a mixing movement 25 according to the invention by shaking or weighing, which also causes a movement 10 of the sensor 5 via the resulting movement of the reaction mixture 3 and reactor 1, so that it moves into the detection area 9 according to the invention the measuring arrangement 6 moves in and out again.
  • Figure 3 shows a schematic representation of a sensor 5 according to the invention for carrying out the method according to the invention using the example of a shake flask as a reactor 1 for cultivating cells with an addition system 18 for sensors 5.
  • the one in Figure 3 shows a schematic representation of a sensor 5 according to the invention for carrying out the method according to the invention using the example of a shake flask as a reactor 1 for cultivating cells with an addition system 18 for sensors 5.
  • the one in Figure 3 shows a schematic representation of a sensor 5 according to the invention for carrying out the method according to the invention using the example of a shake flask as a reactor 1 for cultivating cells with an addition system 18 for sensors 5.
  • the sensor 5 shown in FIG. 3 is designed in an arc shape in order to be able to carry out a uniform movement 10 with as little collision as possible along the round geometry of the wall of the shaking flask as a reactor 1.
  • Markers 13A and 13B are embedded in the sensor matrix 12 in the front and rear areas of the sensor 5. Between the areas with markers 13A and 13B there are several areas with different sensory components 7A and 7B as well as converting component 11, the components 7A and 7B or their combinations (7A enveloped by 11) being designed as particles which are embedded in the sensor matrix 12 are. There is a maximum of one type of particle in each area of the sensor 5 shown in FIG. 3. An area of the sensor 5 shown contains particles of the sensory component 7A, which, for example, generates an oxygen-dependent signal 8A.
  • An area of the sensor 5 shown contains particles of the sensory component 7B, which, for example, generates a pH-dependent signal 8B.
  • An area of the sensor 5 shown contains combined particles from the sensory component 7A, which are each coated with a converting component 11 and, for example, generate an oxygen-dependent signal 8A, the local oxygen concentration in the area of influence of the covered sensory component 7A being controlled by an oxidoreductase (e.g. for glucose , lactate, glutamine) as an enveloping converting component 11 is influenced.
  • an oxidoreductase e.g. for glucose , lactate, glutamine
  • the individual areas of the sensor 5 are segmented so that the different signals 8 can be assigned locally as part of the detection, or even afterwards, and thus a distinction can be made, for example, between the oxygen concentration and the glucose concentration in the reactor contents 2 determined under local oxygen consumption.
  • the areas of the sensor 5 that contain sensory components 7 can be distinguished regardless of the orientation of the sensor 5.
  • the lower section of Figure 3 shows a schematic representation of a shake flask as a reactor 1 before the start of the mixing according to the invention as a result of a mixing movement 25.
  • the reactor 1 comprises a reactor content 2, which is formed by a cell culture suspension as a reaction mixture 3 and, above it, by a gas-filled headspace 4 becomes.
  • an addition system 18 for sensors 5 is shown, which is divided into individual segments, each segment containing at least one sensor 5 and a storage medium 19.
  • the sensors 5 and the storage medium 19 are sterile.
  • the addition 20 of at least one sensor 5 to the reactor 1 takes place as part of the method according to the invention.
  • FIG. 4 shows a schematic representation of how to carry out the method according to the invention on a shake flask as a reactor 1 for cultivating cells using the sensor 5 according to the invention shown in Figure 3.
  • a sensor 5 in the embodiment shown above in FIG. 3, so that for the purpose of clarity, a numerical designation of the sensor components is omitted and reference is made to FIG. 3.
  • a measuring arrangement 6 which comprises at least one signal exciter 16 and at least one signal detector 15.
  • the measuring arrangement 6 comprises several signal exciters 16 and signal detectors 15 working in parallel, so that the various markers 13A and 13B as well as the sensory components 7A and 7B and the sensory component 7A covered with a converting component 11 are detected by a single measuring arrangement 6 can.
  • the various markers 13A and 13B as well as the sensory components 7A and 7B and the sensory component 7A covered with a converting component 11 are detected by a single measuring arrangement 6 can.
  • only one signal exciter 16 and one signal detector 15 are shown in FIG.
  • At least one combination of at least one signal exciter 16 and at least one signal detector 15 spans at least one detection area 9 of the measuring arrangement 6, which penetrates through the wall of the reactor 1 into the reactor contents 2, so that there the signal excitation 17 of at least one sensor 5 according to the invention over the at least a sensory component 7 contained in it can take place and its signal 8 or a marker signal 14 can be detected by at least one signal detector 15 of the measuring arrangement 6.
  • the shaking flask shown in Figure 4 as a reactor 1 is subjected to a mixing movement 25 according to the invention by orbital shaking, which also causes a movement 10 of the sensor 5 via the resulting periodic rotational movement of the reaction mixture 3, so that according to the invention it moves into the detection area 9 of the measuring arrangement 6 also moved out again.
  • the reaction mixture 3 is moving straight into the detection area 9 of the measuring arrangement 6. Since there is no sensor 5 in the detection area 9, neither is a valid one Marker signal 14 nor a valid signal 8 of a sensory component 7 is detected by the measuring arrangement 6. If the reaction mixture 3 moves further, the sensor 5 reaches the detection area 9 of the measuring arrangement 6.
  • the area of the sensor 5 with marker 13A reaches the detection area 9, so that based on the measurement arrangement 6 the orientation of the sensor 5 can be determined using the signal excitation 17MA detected marker signal 14A.
  • the signal exciters 16 and signal detectors 15 are now activated for the following sensory components 7A and 7B, so that at the moment the respective sensory component 7 is present in the detection area 9 of the measuring arrangement 6, a valid signal 8 can be detected in order to determine the property associated with the invention of the reaction mixture 3 to monitor.
  • the measuring arrangement 6 detects the signals 8 of the sensory components 7 one after the other, initially with Signal excitation 17A the signal 8A of the sensory component 7A ( Figure 4, II), then with signal excitation 17B the signal 8B of the sensory component 7B ( Figure 4, III), then with signal excitation 17A the signal 8A of the sensory component 7A enveloped with converting component 11 ( Figure 4, IV). Finally, the marker signal 14B of the marker 13B is detected using the signal excitation 17MB (FIG. 4, IV), which also validates the positioning of the sensor 5 during its movement 10 through the detection area 9.
  • the sensor 5 then moves out of the detection area 9 of the measuring arrangement 6 ( Figure 4, V-VI).
  • the movement 10 of the sensor 5 is also subject to a certain periodicity.
  • the detection of the individual signals 8 and marker signals 14 can be synchronized in some embodiments of the invention.
  • the use of several different markers 13 with correspondingly different marker signals 14 in different segments of a sensor 5 allows a clear assignment of different signals 8 to their properties of the reactor contents 2 or reaction mixture 3 to be monitored, even if the position changes as a result of the movement 10 of a sensor 5 and orientation of the sensor 5 in the detection area 9.
  • FIG. 5 shows a schematic representation of two sensors 5 according to the invention and a device according to the invention for carrying out the method according to the invention on a stirred tank reactor as reactor 1.
  • the sensors 5A and 5B shown contain sensory components 7 designed as particles, which are surrounded by a converting component 11 and in the Sensor matrix 12 are embedded.
  • Sensor 5A also contains markers 13A in two areas, the marker signal 14A of which differs from the marker signal 14B of the marker 13B contained in sensor 5B, so that the sensors 5A and 5B can be distinguished based on their marker signals 14A and 14B.
  • the sensors 5A and 5B each include a fin 26 with a density that is higher than that of the reaction mixture 3 and a fin 27 with a density that is lower than that of the reaction mixture 3.
  • these fins 26 and 27 are used in particular for fluid dynamic stabilization and positioning of the sensors 5 are used. Furthermore, by appropriately dimensioning the fins 26 and 27, the average density of a sensor 5 can be set and the probability of the sensor 5 being located in certain areas of the reactor contents 2 or the reaction mixture 3 can be specifically influenced.
  • the sensor 5A shown at the top left in Figure 5 is equipped with a large fin 27A of low density and a small fin 26A of high density, so that the average density of the sensor 5A is lower than the density of the reaction mixture 3 of the reactor 1 shown on the right in Figure 5. Therefore, even during turbulent mixing, sensor 5A remains predominantly in the upper part of the reaction mixture 3 and can therefore be used specifically to monitor properties of this upper region of the reaction mixture 3.
  • the sensor 5B shown at the bottom left in Figure 5 is equipped with a small fin 27B of low density and a large fin 26B of high density, so that the average density of the sensor 5B is higher than the density of the reaction mixture 3 of the reactor 1 shown on the right in Figure 5.
  • sensor 5B remains predominantly in the lower part of the reaction mixture 3 and can therefore be used specifically to monitor properties of this lower region of the reaction mixture 3.
  • FIG. 5 an embodiment of the invention is shown schematically on a stirred tank reactor as reactor 1 with the sensors 5A and 5B shown on the left in Figure 5.
  • a stirred tank reactor is shown as a reactor 1, the reactor contents 2 of which are formed by a cell culture suspension as a reaction mixture 3 and, above it, by a gas-filled headspace 4.
  • the reaction mixture 3 there are two sensors 5A and 5B in the embodiment shown on the left in FIG. 5, with the sensors 5A predominantly in the upper region and the sensors 5B predominantly in the lower region of the reaction mixture 3.
  • a measuring arrangement 6 for both the upper and lower region of the reaction mixture 3, each of which comprises at least one signal exciter 16 and at least one signal detector 15.
  • each measuring arrangement 6 comprises several signal exciters 16 and signal detectors 15 working in parallel, so that the various markers 13A and 13B as well as the sensory component 7A covered with a converting component 11 can be detected by a single measuring arrangement 6.
  • signal exciter 16 and signal detector 15 For the purpose of clarity, only one signal exciter 16 and signal detector 15 per measuring arrangement 6 is shown.
  • At least one combination of at least one signal exciter 16 and at least one signal detector 15 spans at least one detection area 9 of the respective measuring arrangement 6, which penetrates through the wall of the reactor 1 into the reaction mixture 3, so that there the signal excitation 17 of at least one sensor 5 according to the invention is transmitted via the at least one sensory component 7 contained in it can take place and its signal 8 or a marker signal 14 can be detected by at least one signal detector 15 of the measuring arrangement 6.
  • the reactor 1 shown has an agitator 28 and is thereby exposed to a mixing movement 25 according to the invention, which also causes a movement 10 of the sensors 5A and 5B via the resulting movement of the reaction mixture 3, so that, according to the invention, these move into the detection area 9 of the respective measuring arrangement 6 move in and out again.
  • Figure 6 shows a schematic representation of a sensor 5 according to the invention in a core-shell design with adapted geometry for use in orbitally shaken systems.
  • the sensor matrix 12 forms the mechanically stable core of the sensor 5 and acts as a carrier for the marker 13 and the sensory component 7.
  • Figure 6 shows a longitudinal section of a sensor 5 according to the invention at the top and two cross sections at the bottom, on the left a cross section through the central area of the sensor 5 and on the right is a cross section through one of the edge caps of the sensor 5.
  • the sensory component 7 is set back inwards in such a way that even when the sensor 5 comes into contact with the reactor 1, it does not touch the latter and is therefore largely protected from mechanical damage.
  • the two end caps coated with marker 13 stand out clearly and thus form the main points of contact of the sensor 5 with the reactor 1.
  • the two end caps of the sensor 5 are angular, hexagonal in the embodiment shown in Figure 6.
  • the sensor 5 is adapted to the flow conditions in the reaction mixture 3.
  • the sensor 5 is constructed rotationally symmetrical and has a length-diameter ratio of more than one, preferably at least two.
  • the embodiment of the sensor 5 according to the invention shown in Figure 6 has a smooth transition of the hexagonal edge structures of the two end caps carrying the marker 13 into a fluidically more advantageous edge-free geometry, here a spherical segment.
  • the depth of and the transition into the reset of the sensory component 7 can also be advantageously chosen so that that in the application there is an optimal compromise between the avoidance of direct contact between the sensory component 7 or other mechanically sensitive components of the sensor 5 and the reactor 1 as well as a flow to the sensor 5 that is optimal for the application, in particular but not exclusively through the use of suitable Geometries to avoid flow stalls and eddies behind the flow end caps of the sensor 5.
  • the depth of the reset of the sensory component 7 is also chosen to be just so small that mechanical protection of sensitive components of the sensor 5 by avoiding contact with the reactor 1 as well Fluidic optimization is ensured but at the same time as little reaction mixture 3 as possible is between the sensory component 7 and the at least one suitable measuring arrangement 6 during the detection of at least one signal 8 of at least one sensory component 7.
  • This is important for applications in which, in particular but not exclusively, the turbidity or absorption of the reaction mixture 3 changes over time, for example due to cell growth or the formation or dissolution of substrates, products, by-products, metabolites, foams, particles, emulsions or others phase mixtures.
  • the sensor 5 according to the invention shown in FIG Carriers of another luminophore act as marker 13.
  • the sensory component 7 and the marker 13 can also be embedded in one or more suitable sensor matrices 12 and applied to the core as the primary sensor matrix 12 by means of this.
  • Converting component that interacts with at least one sensory component 7 or at least one converting component

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Abstract

L'invention concerne un procédé et un dispositif de surveillance de contenu de réacteurs mixtes. L'invention est particulièrement applicable à la surveillance de contenu de réacteurs mixtes dont les exigences en matière de stérilité et de pureté sont élevées, ainsi qu'à des applications dans lesquelles la géométrie des réacteurs est complexe ou leur taille est réduite, ce qui limite l'accès aux appareils de mesure. L'invention peut par conséquent être appliquée, par exemple, à la surveillance des cultures cellulaires ou des réactions chimiques, des processus de traitement, de purification et de formulation de produits pharmaceutiques, biologiques ou chimiques, ainsi qu'à la surveillance des processus de stockage. Le problème soulevé par la présente invention est de fournir un procédé permettant de surveiller le contenu de réacteurs mixtes à l'aide de capteurs situés à l'intérieur du réacteur, de manière robuste et facile à manipuler, tout en offrant une bonne extensibilité pour les petits et les grands volumes de réacteurs et la possibilité de détecter une pluralité de propriétés de contenu de réacteur. Le problème de la surveillance de contenu de réacteurs mixtes est résolu selon l'invention, au moins une propriété de contenu du réacteur pouvant être surveillée et ladite au moins une propriété influençant au moins un signal d'au moins un composant de capteur et ledit au moins un signal dudit au moins un composant de capteur étant détecté par au moins un dispositif de mesure, par un procédé dans lequel au moins un capteur, qui contient ledit au moins un composant de capteur, n'est pas fixe et se déplace dans le réacteur de sorte que ledit au moins un capteur ne se trouve pas en permanence dans la zone de détection dudit au moins un dispositif de mesure et que ledit au moins un signal dudit au moins un composant de capteur est au moins détecté par ledit au moins un dispositif de mesure pendant que ledit au moins un capteur se trouve dans la zone de détection dudit au moins un dispositif de mesure.
EP23727812.2A 2022-06-13 2023-05-12 Procédé et dispositif de surveillance de contenu de réacteurs mixtes Pending EP4341683A1 (fr)

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
DE102022002116.1A DE102022002116B4 (de) 2022-06-13 2022-06-13 Verfahren und Vorrichtung zur Überwachung des Inhalts durchmischter Reaktoren
US202263433229P 2022-12-16 2022-12-16
DE102023111143 2023-04-28
PCT/EP2023/062790 WO2023241869A1 (fr) 2022-06-13 2023-05-12 Procédé et dispositif de surveillance de contenu de réacteurs mixtes

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EP4341683A1 true EP4341683A1 (fr) 2024-03-27

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DE10101576B4 (de) 2001-01-15 2016-02-18 Presens Precision Sensing Gmbh Optischer Sensor und Sensorfeld
US20040171094A1 (en) 2001-06-18 2004-09-02 Ingo Klimant Oxygen sensors disposed on a microtiter plate
DE102004017039A1 (de) 2004-04-02 2005-11-03 Rwth Aachen Verfahren und Vorrichtung zur Erfassung von Prozessparametern von Reaktionsflüssigkeiten in mehreren geschüttelten Mikroreaktoren
US7824902B2 (en) 2006-11-03 2010-11-02 Mark Selker Optical interface for disposable bioreactors
WO2010017519A1 (fr) 2008-08-08 2010-02-11 Broadley-James Corporation Dispositif pour exposer un capteur à une population de culture cellulaire dans une cuve de bioréacteur
DE102014001284B3 (de) * 2014-02-01 2015-01-22 aquila biolabs GmbH Verfahren, Vorrichtung und System zur automatisierten Bestimmung optischer Dichten oder der Veränderung optischer Dichten von Reaktionsgemischen in geschüttelten Reaktoren
DE102014107837B4 (de) * 2014-06-04 2021-09-02 Presens Precision Sensing Gmbh Optischer Sensor zum quantitativen Nachweis eines Analyten in einer Probe und Verfahren zur Herstellung des Sensors
DE102017012186B4 (de) * 2017-02-17 2018-10-18 aquila biolabs GmbH Verfahren und Vorrichtung zur Abstimmung optischer Messungen an kontinuierlich durchmischten Reaktoren
DE102018105174B4 (de) 2018-03-07 2020-03-12 Presens Precision Sensing Gmbh Analyseeinheit
EP3821230B1 (fr) 2018-07-09 2024-03-27 PreSens Precision Sensing GmbH Système d'analyse d'un échantillon de fluide
US20220178824A1 (en) * 2020-12-08 2022-06-09 Imam Abdulrahman Bin Faisal University Method for detecting heavy metal pollutants using a fluorescent material from bacillus endophyticus and method for making

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US20240287435A1 (en) 2024-08-29

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