Single-Use Bioreactor Interface for Single-Use Probes

Abstract

An optical interface with a housing such as a bioreactor bag, and a bioreactor bag having the optical interface. In some embodiments, the optical principle of Raman technology is used to provide several embedded single-use optical interfaces in the housing and, if necessary, modify the associated Raman probe. In some embodiments, the optical interface includes a single-use cap for reducing the ambient light intensity that can be harmful to Raman signals. The devices according to the embodiments disclosed herein enable avoidance of aseptic breakage that can typically occur due to the insertion of a penetrable reusable sensor through a sterile connection into the housing.

Description

Technical Field

[0001] This application claims the benefit of U.S. Provisional Patent Application No. 63 / 347,683, filed on June 1, 2022, the entire content of which is incorporated herein by reference in its entirety.

Background Art

[0002] Single-use disposable bioreactors are often vessels selected for biomolecule processing. Bags are often used as such single-use containers for receiving and maintaining fluids such as biological fluids. Bags can include single-layer or multi-layer flexible walls formed from polymer compositions such as polyethylene including ultra-high molecular weight polyethylene (UHMWPE), ultra-low density polyethylene (ULDPE), linear low density polyethylene (LLDPE), low density polyethylene (LDPE), medium density polyethylene (MDPE); polypropylene (PP); ethylene vinyl alcohol (EVOH); polyvinyl chloride (PVC); polyvinyl acetate (PVA); ethylene vinyl acetate copolymer (EVA copolymer); thermoplastic elastomer (TPE), and / or blends or alloys of any of the foregoing materials, as well as various other thermoplastic materials and additives known to those skilled in the art. The bag must be able to maintain a biologically active environment, for example, an environment in which cells can be grown in a cell culture situation. The bag may be a two-dimensional (2D) or "pillow" bag, or a three-dimensional (3D) bag. The bag may include one or more inlets, one or more outlets, and optionally other features such as a sterile vent, sparger, and ports for sensing the liquid within the container for parameters known to those skilled in the art such as conductivity, pH, temperature, dissolved gases such as oxygen and carbon dioxide.

[0003] Conventional monitoring strategies for mammalian cell cultures lack real-time measurements and rely on daily or other periodic manual sampling and feeding, which increases the risk of contamination and batch failure. Raman analyzers such as the ProCellics(TM) Raman Analyzer with Bio4C(TM) PAT Raman software available from Merck KGaA enable the inline and real-time monitoring, evaluation, and / or measurement of various parameters of bioreactor cultures. These parameters can include one or more important process parameters such as the concentrations of glucose, lactate, and ammonium, as well as important performance indicators such as total cell density and viable cell density. However, the sensing device, whether single-use or disposable or not, must not interfere detrimentally with the single-use device or compromise its integrity. Furthermore, the way the sensing device communicates with the contents of the bioreactor can pose problems at multiple levels, including sterility, component compatibility, and the method of attachment and removal.

[0004] Raman analyzers and equipment are mainly multi-purpose and require an autoclave process for sterilization, etc., before being incorporated into a bioreactor bag. It also requires a specific sterile interface for inserting the probe into the bag. SUMMARY OF THE INVENTION PROBLEMS TO BE SOLVED BY THE INVENTION

[0005] To improve the current system, it is desirable to provide a single-use analyzer optical interface with a bioreactor bag.

[0006] To further understand the nature of these and other objects of the present disclosure, reference should be made to the following description in conjunction with the accompanying drawings. MEANS FOR SOLVING THE PROBLEM

[0007] The problems of the prior art are addressed by the embodiments disclosed herein, which relate to an optical interface with a container or housing, such as a single-use bioreactor and / or a single-use bag of a mixer. In some embodiments, the optical principle of Raman technology is used to provide several embedded single-use optical interfaces to the bioreactor bag and, if necessary, modify the associated Raman probe. Preferably, the interface is gamma-sterilization compatible. In some embodiments, the optical interface includes a single-use cap for reducing the ambient light intensity that may be harmful to the Raman signal. The device according to the embodiments disclosed herein enables the avoidance of the breach of sterility that may typically occur due to the insertion of an invasive reusable sensor into the bioreactor through a sterile connection.

[0008] In some embodiments, the optical interface may include a single-use cap for reducing the ambient light intensity that may be harmful to the Raman signal.

[0009] In certain embodiments, the optical interface includes a sleeve configured to receive a probe, such as a sampling immersion tube of a Raman probe, and the sleeve has a light-reducer so that external light that may interfere with Raman signal generation is reduced or eliminated. In some embodiments, the sleeve cooperates with a barb that interfaces with the bioreactor and a clamp that secures the sleeve to the barb. In certain embodiments, the interface can be sterilized and keep the container or housing (e.g., the bag) closed even without the probe, so the interface does not leak even without the probe.

[0010] In some embodiments, the sleeve includes an optical window, such as a sapphire window or a quartz window. As a result, such a window may not be present in the probe itself.

[0011] Accordingly, an interface assembly for inserting a probe into a bioreactor is disclosed, the interface assembly comprising an elongate sleeve having a passage configured to receive the probe; and a clamp for fastening the sleeve to a fitting that can be disposed within a port of the bioreactor, the sleeve including a window. In some embodiments, the passage has a generally cylindrical cross-section. In some embodiments, the sleeve has a proximal end into which the probe is inserted and a distal end spaced from the proximal end, and a cap is disposed at the distal end. The cap may extend longitudinally from the window and may include one or more radial openings. In some embodiments, the interface further comprises a barb fitting having a barbed stem fixed to the clamp and a flange configured to be disposed within the internal volume of the bioreactor. In some embodiments, the window is a sapphire window or a quartz window.

[0012] In some embodiments, a bioreactor having an internal volume is disclosed, the bioreactor comprising a port providing access from a region external of the bioreactor to the internal volume, and a fitting attached to the port, the fitting being configured to receive an interface assembly configured to receive a probe through the port into the bioreactor volume, the interface assembly comprising an elongate sleeve having a passage configured to receive the probe, the sleeve including a window such as a sapphire window or a quartz window. The interface assembly may further comprise a clamp for fastening the sleeve to the fitting, and the fitting may be a barb fitting or a fitting threaded into the port.

[0013] Accordingly, an interface assembly configured to insert a probe into a housing such as a bioreactor or a bioreactor bag is disclosed, the interface assembly comprising an elongated sleeve having a passage configured to receive the probe; and a clamp for fastening the sleeve to a fitting that can be disposed within a port of the housing, the sleeve including an optical window. The sleeve may have a proximal end into which the probe is inserted and a distal end spaced from the proximal end, with a cap disposed at the distal end. The cap may extend longitudinally from the window and may have one or more radial openings. The probe may include an optical lens disposed adjacent to the window of the sleeve when assembled. The interface assembly may further comprise a barb joint having a barbed stem fixed to the clamp and a flange configured to be disposed within the internal volume of the housing. The housing may have a threaded port such that the interface assembly can be screwed into the threaded port. The elongated sleeve may be a single-use sleeve. The optical window may include sapphire.

[0014] Also disclosed is a housing having an internal volume, the housing comprising a port providing access from a region external of the housing to the internal volume, and a fitting attached to the port, the fitting being configured to receive an interface assembly configured to insert a probe through the port into the volume, the interface assembly comprising an elongated sleeve having a passage configured to receive the probe, the sleeve including an optical window. The interface assembly may further comprise a clamp for fastening the sleeve to the fitting, the clamp may be a barb joint or may be screwed onto the port. The optical window may include sapphire. The housing may be a bioreactor or a bioreactor bag. BRIEF DESCRIPTION OF THE DRAWINGS

[0015]

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DETAILED DESCRIPTION OF THE INVENTION

[0016] A more complete understanding of the components, processes, and devices disclosed in this specification can be obtained by referring to the accompanying drawings. The drawings are merely schematic diagrams based on the convenience and ease of explaining the present disclosure, and thus are not intended to show the relative sizes or dimensions of the devices or their components, and / or to define or limit the scope of the exemplary embodiments.

[0017] Certain terms are used in the following description for clarity, but these terms are intended to refer only to the specific structures of the embodiments selected for the description in the drawings and are not intended to define or limit the scope of the present disclosure. It should be understood that in the drawings and the following description, like numerals refer to components with like functions.

[0018] The singular forms "a", "an", and "the" include plural referents unless the context clearly dictates otherwise.

[0019] As used herein, various devices and components may be described as "comprising" other components. The terms "comprise(s)", "include(s)", "having", "has", "can", "contain(s)", and their variations as used herein are intended to be open-ended transitional phrases, terms, or words that do not exclude the possibility of additional components.

[0020] All ranges disclosed herein include the recited endpoints and can be combined independently (e.g., the range "from 2 inches to 10 inches" includes the endpoints, 2 inches and 10 inches, and all intermediate values).

[0021] As used herein, the language representing approximation may be applied to modify any quantitative expression that can vary without resulting in a change to the associated basic function. Thus, values modified by terms such as "about" and "substantially" may not be limited to their exact specified values. The modifier "about" should also be considered as disclosing a range defined by the absolute values of two endpoints. For example, the expression "about 2 to about 4" also discloses the range "2 to 4".

[0022] Note that many of the terms used herein are relative terms. For example, the terms "upper" and "lower" are relative to each other, i.e., an upper component is located at a higher altitude than a lower component, and should not be construed as requiring a particular orientation or position of the structure. As a further example, the terms "inner", "outer", "inward", and "outward" are relative to a center and should not be construed as requiring a particular orientation or position of the structure.

[0023] The terms "top" and "bottom" are relative to an absolute reference, i.e., the surface of the earth. In other words, a top position is always located at a higher altitude than a bottom position with respect to the surface of the earth.

[0024] The terms "horizontal" and "vertical" are used to indicate an absolute reference, i.e., a direction with respect to the earth's surface. However, these terms should not be construed as requiring that the structures be perfectly parallel or perfectly perpendicular to each other.

[0025] The terms "bioreactor", "bag", and "container" are generally used interchangeably within the present disclosure. As used herein, the terms bioreactor, bag, and container refer to any manufactured or designed device or system that supports a biologically active environment, such as a mixer bag, such as a bag for cell culture media preparation, antibody-drug conjugate (ADC) reactions, or other downstream processing steps that are or can be performed within the bulk. In some cases, a bioreactor is a vessel having an internal volume in which a cell culture process involving an organism or a biochemically active substance derived from such an organism is performed. A flexible bioreactor, bag, or container implies, for example, a flexible vessel that can contain biological fluids and can be folded, collapsed, and expanded, and / or otherwise manipulated. A single-use bioreactor, bag, or container is also typically flexible and is a vessel that is used once and discarded. In some embodiments, the bag, bioreactor, or container may be a two-dimensional (2D) or "pillow" bag, or alternatively a three-dimensional (3D) bag. The specific geometric shape of the container or bioreactor is not limited in any of the embodiments disclosed herein. In some embodiments, the container can include a rigid base that provides access points such as ports or vents. Any container described herein can include one or more inlets, one or more outlets, and optionally other features such as sterile vents, spargers, and ports for sensing the liquid within the container for parameters known to those skilled in the art, such as conductivity, pH, temperature, dissolved gases, such as oxygen and carbon dioxide. The container can be a bioreactor of a size sufficient to contain the fluids to be mixed, such as cells and culture media, a pilot scale, for example from 50 L, to a small or large production volume container, for example from 500 L to 3000 L or more.

[0026] The bioreactor can comprise a flexible and foldable bag having an inner surface and an outer surface. The inner surface bounds a compartment or space into which a liquid can be added. The bag can include a sidewall having a substantially circular or polygonal cross-section that extends between a first end and a second end opposite the first end when the bag is deployed. The first end terminates in a top end wall and the second end terminates in a bottom end wall. The bag can be constructed from a flexible waterproof material such as a polyethylene or other polymer sheet having a thickness in the range of about 0.1 mm to about 5 mm (more typically about 0.2 mm to about 2 mm). Other thicknesses can be used. The material can be composed of a single layer material or can include two or more layers that are either sealed together or separated to form a double wall container. When the layers are sealed together, the material can include a laminated or extruded material. A laminated material includes two or more separately formed layers that are later fixed to each other by an adhesive. The bag material includes a single integral sheet that includes two or more layers of different materials laminated together or separated by contacting layers that are coextruded all at once. In one embodiment, the material permits direct contact with living cells and can maintain a solution in a sterile state. In such an embodiment, the material can also be sterilizable, such as by ionizing radiation. Other examples of materials that can be used are disclosed in U.S. Patent No. 6,083,587, issued July 4, 2000, and U.S. Patent Application No. 10 / 044,636, filed October 19, 2001, which are incorporated herein by reference.

[0027] The terms "sterile" and "sterilized" are defined as a state free of contaminants and, particularly in the bioprocessing industry, as a state free of undesirable viruses, bacteria, germs, and other pathogens such as microorganisms. In this context, the terms "reduced bioburden" and "bioburden reduction" (e.g., by a non-sterilizing dose of gamma or X-ray radiation less than 25 kGy) can be used in place of specific embodiments that do not require a claim of sterility.

[0028] In certain embodiments, a bioreactor system is disclosed that comprises a bioreactor volume or chamber for containing a bioreactor culture and a probe capable of providing a Raman spectrum of the bioreactor culture within the bioreactor volume or chamber. In some embodiments, the probe interfaces with a bioreactor chamber having an optical interface described herein.

[0029] In certain embodiments, a computer can be provided in the bioreactor system that comprises a processor or processing unit that executes computer-readable instructions. The processor or processing unit can be a general-purpose computing device such as a microprocessor. Alternatively, it can be a dedicated processing device such as a programmable logic controller (PLC). Also, a storage element can be provided for storing instructions and providing a temporary storage device for use by the processor. The storage element can utilize any memory technology such as RAM, ROM, EEPROM, flash ROM, NVRAM, or any other suitable technology. An input device such as a touch screen, keyboard, or other suitable device can be provided.

[0030] In some embodiments, Raman spectroscopy can be performed in the visible, near-infrared, infrared, near-ultraviolet, or ultraviolet (UV) region. In some embodiments, surface-enhanced Raman spectroscopy (SERS) can be used. In some embodiments, resonance Raman spectroscopy, tip-enhanced Raman spectroscopy, polarization Raman spectroscopy, induced Raman spectroscopy, transmission Raman spectroscopy, spatially offset Raman spectroscopy, difference Raman spectroscopy, Fourier transform (FT) Raman spectroscopy, or hyper Raman spectroscopy can be used.

[0031] Referring now to FIGS. 1, 2A and 2B, an optical interface assembly 10 according to a particular embodiment is shown. In some embodiments, the optical interface assembly 10 includes a sleeve 12 and a clamp 14. The sleeve 12 may have a generally cylindrical configuration and an internal bore or passage configured (e.g., shaped and sized) to receive a probe such as a Raman probe. In the illustrated embodiment, the sleeve 12 includes an optional cap 15 disposed at the distal end of the sleeve 12, and the cap 15 may be integral with the sleeve 12. The cap 15 has one or more radial openings or through-holes 19 that terminate in front of the most distal end of the cap 15. The one or more radial openings can provide access to the contents of the bioreactor or expose an optical window to the contents of the bioreactor when in the assembled state. The cap 15 helps to manage stray light (e.g., reduce the ambient light intensity that can interfere with the Raman signal) and helps to protect the user from laser light that might leak out of the sleeve 12 in the absence of the cap. The optical window 16 can be disposed at or near the end of the passage as shown and can be an overmolded sapphire window (as those skilled in the art will know, overmolding is a process of adding an additional layer of material on top of an existing element). Typically, a first material, sometimes also referred to as a substrate, is partially or completely covered by a subsequent material (the overmolding material) during the manufacturing process. Those skilled in the art will understand that any material suitable for the application, such as sapphire or quartz, can be used for the optical window.

[0032] Preferably, the sleeve 12 is single-use, i.e., disposable; it is designed to be discarded after one use. Preferably, it comprises a gamma-compatible material, e.g., plastic or stainless steel, such as a plastic material having a low-intensity Raman signature, e.g., a plastic having a CxHy composition, e.g., polyethylene (PE), polypropylene (PP), polystyrene (PS), polystyrene sulfonate (PSS), polycarbonate (PC), polyamide PA6 / PA12, etc., to facilitate sterilization.

[0033] The outer diameter of the sleeve 12 is configured to receive a standard hose barb 17, such as a 1-inch barb. The barb 17 is configured to mate, dock, couple, or otherwise, typically mechanically, in a conventional manner, e.g., to integrate with a port of a bag, with a bioreactor bag 300 as shown in FIGS. 11 and 12, and includes a stem with a barb to be fixed to the clamp 14. Thus, the barb 17 has a first region 17A, e.g., a flange that is disposed (i.e., is inside) the bioreactor chamber or volume during use, and a second region 17B, e.g., a stem having one or more radial ridges or serrations and located outside the bag, e.g., on the probe insertion side. The flange surrounds the stem and projects radially outward from the stem. The barb 17 may include threads, such as a female thread, to fix or maintain the probe within the sleeve 12. Another embodiment is shown in FIGS. 13 and 14, where a PG13.5 or other thread on the bag 300 is used and the single-use sleeve 12 is screwed thereto. FIG. 14 shows the insertion of the tube into the bag 300 and the connection of the probe in this embodiment. The threaded member coupled to the bioreactor port may include a sapphire window and may include an optical lens.

[0034] In certain embodiments, the sleeve 12 is oriented to be inserted into barb 17 and secured to the barb 17 by a fastener such as clamp 14 (Figs. 2A, 2B). In some embodiments, clamp 14 includes a proximal circumferential radially inwardly projecting ridge 14B that abuts an annular ring 11 on sleeve 12 and a distal circumferential radially inwardly projecting ridge 14A that abuts a barb radial ridge 17C on the stem to lock the barb in place. The annular ring 11 functions as a stop to properly position or dispose the clamp 14 on the sleeve 12. An O-ring seal 18 or the like may be disposed on the sleeve 12 to ensure a liquid-tight connection between the barb 17 and the sleeve 12. The configuration of this assembly avoids aseptic breakage despite the insertion of the penetrating reusable sensor. In some embodiments, the probe or tube that may be introduced into the sleeve 12 may be threaded (e.g., PG13.5 or other thread configuration) or may be mechanically fastened to the sleeve 12.

[0035] Figs. 3, 4, 5A and 5B show one embodiment of a probe 20 shown as insertable into or inserted into sleeve 12. In the illustrated embodiment, the probe 20 includes a terminal optical lens 29 that is disposed adjacent to the optical window 16 of the sleeve 12 when in an assembled state.

[0036] Figures 6A, 6B, 7A, 7B and 8 illustrate embodiments where a barb-like fitting 17' is integrated with the sleeve 12'; that is, formed as a single integral element. Thus, in certain embodiments, the sleeve 12' having an overmolded optical window 16' (e.g., sapphire) and optionally one or more optical lenses on the sleeve 12' includes a barb-like structure 17' overmolded on the sleeve 12'. Similar to the barb 17, the barb-like structure 17' has a first region 17A', e.g., a flange that is disposed (i.e., inside) within a bioreactor chamber or volume during use, and a second region 17B', e.g., having one or more radial ridges or serrations, and a stem that is outside the bag, e.g., on the probe insertion side. The flange surrounds the stem and projects radially outward from the stem. The sleeve 12' has a generally cylindrical configuration and an internal bore or passage configured (e.g., shaped and dimensioned) to receive a probe such as a Raman probe. An optional end cap 15' may be present for stray light management (e.g., reducing ambient light intensity that may be harmful to the Raman signal) and laser safety and may be integrated with the sleeve 12'. Similar to the embodiment of FIG. 1, the flange functions as an interface with the bioreactor bag, e.g., by a port in the bioreactor.

[0037] In some embodiments, a probe 20 that can include an internal stainless steel tube can be coupled to the sleeve, such as by a male thread 21 that mates with a female thread within the sleeve, as soon as it is in an operable arrangement. One or more internal optical lenses 22 can be mounted to the internal tube, e.g., by a suitable lens holder. Alternatively, one or more optical lenses may be mounted to the sleeve 12', and the probe body 20 may be mounted directly to the sleeve 12' and there may be no internal tube.

[0038] Figures 9A, 9B, 9C, and 9D show another embodiment that omits the barb fitting by including a male thread 122 (e.g., of the PG13.5 / M18 thread type) on the proximal end of the single-use sleeve 12'' having the optical window 16'' that mates with the threads on the port of the bioreactor. A probe 20'' having an internal tube 200 with an optical lens can be inserted into the internal bore or passage of the sleeve 12'' (FIG. 9C). In some embodiments, the internal tube 200 can be omitted and the probe 20'' is mounted directly to the single-use sleeve 12'' (FIG. 10).

Claims

1. An interface assembly configured to insert a probe into a housing, comprising an elongated sleeve having a passage configured to receive the probe, and a clamp for fastening the sleeve to a fitting that can be disposed within a port of the housing, wherein the sleeve includes an optical window.

2. The interface assembly according to claim 1, wherein the sleeve has a proximal end into which the probe is inserted and a distal end spaced from the proximal end, and a cap is disposed at the distal end.

3. The interface assembly according to claim 2, wherein the cap extends longitudinally from the window.

4. The interface assembly according to claim 1, wherein the cap has one or more radially extending openings.

5. The interface assembly according to claim 1, wherein the probe includes an optical lens disposed adjacent to the window of the sleeve when assembled.

6. The interface assembly according to claim 1, further comprising a barb joint having a barbed stem for securing to the clamp and a flange configured to be disposed within the internal volume of the housing.

7. The interface assembly according to claim 1, wherein the housing has a threaded port and the interface assembly is threaded into the threaded port.

8. The interface assembly according to claim 1, wherein the elongated sleeve is for single use.

9. The interface assembly according to claim 1, wherein the optical window includes sapphire.

10. The interface assembly according to claim 1, wherein the housing is a bioreactor.

11. The interface assembly according to claim 1, wherein the housing is a bioreactor bag.

12. A housing having an internal volume, comprising a port providing access from an area external to the housing to the internal volume, and a fitting attached to the port, the fitting being configured to receive an interface assembly for inserting a probe into the volume through the port, the interface assembly comprising an elongate sleeve having a passage configured to receive the probe, the sleeve including an optical window.

13. The housing according to claim 12, wherein the interface assembly further comprises a clamp for fastening the sleeve to the fitting.

14. The housing according to claim 12, wherein the fitting is a barb fitting.

15. The housing according to claim 12, wherein the fitting is screwed into the port.

16. The housing according to claim 12, wherein the optical window comprises sapphire.

17. The housing according to claim 12, wherein the housing is a bioreactor.

18. The housing according to claim 12, wherein the housing is a bioreactor bag.

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