Perfusion bioreactor with filtration system
Patent Information
- Authority / Receiving Office
- JP · JP
- Patent Type
- Patents
- Current Assignee / Owner
- GENZYME CORP
- Filing Date
- 2024-12-26
- Publication Date
- 2026-07-31
Smart Images
Figure 0007898504000001 
Figure 0007898504000002 
Figure 0007898504000003
Abstract
Description
Technical Field
[0001] The present disclosure generally relates to perfusion cell culture apparatuses and cell culture methods.
Background Art
[0002] In vitro cell culture is a complex process of growing cells under controlled conditions outside the natural environment of the cells. Culture conditions vary for each cell type and must be precisely controlled to ensure the correct cell phenotype and / or expression of the desired product.
[0003] Bioreactors provide a controlled environment for growing and maintaining cells in terms of temperature, pH, nutrients, gases, and other requirements as needed. To culture cells over a long period, fresh medium must be continuously supplied to the cells, and the spent medium must be removed at approximately the same rate. To extend the production duration of cell culture in a bioreactor, perfusion bioreactors have been developed that continuously perfuse cells into fresh medium and incorporate the desired product while retaining the cells in the bioreactor. Such bioreactors make it possible to grow cells at a higher cell concentration and maintain the cells for a longer period compared to bioreactors without an active perfusion system.
[0004] FIG. 1 shows a schematic view of a typical state-of-the-art perfusion bioreactor 100 known to those skilled in the art. The perfusion bioreactor includes a bioreactor vessel 110, an alternating tangential flow (ATF) filter 120, a harvest pump 130, and a continuous capture operation 160. The perfusion bioreactor 100 can operate by (i) attaching the ATF filter device 120 to the bioreactor vessel 110; (ii) filling the bioreactor vessel 110 with fresh medium; (iii) inoculating the bioreactor vessel 110 with cells; and (iv) perfusing fresh medium into the bioreactor vessel 110 and removing the spent medium through the harvest pump 130 in series with the ATF filter 120.
[0005] Under these conditions, cells can be cultured and maintained for longer periods. Continuously adding the medium provides the cells with the nutrients they need to grow, and removing used medium removes cellular waste and by-products from the system, preventing them from reaching harmful levels. An ATF filter allows for the removal of used medium from the bioreactor while retaining cells within the bioreactor. Other cell culture parameters are also controlled to improve performance and robustness, typically including temperature, dissolved oxygen, pH, pCO2, and cell density.
[0006] As shown in Figure 1, the perfusion bioreactor 100 can be integrated with a continuously operating capture operation 160. In this scenario, the spent media removed from the bioreactor vessel 110 contains the product of interest and is continuously fed to the continuous capture operation 160, where it is processed and purified in some way.
[0007] Standard perfusion bioreactors, such as the one shown in Figure 1, are intended to operate for long durations (at least 30-60 days). However, the robustness of the equipment can be limited. In particular, the ATF filter can fail, leading to process collapse or disruption. In some cases, the ATF filter can fail catastrophically, allowing cells to pass through the filter and enter the harvest pump. Such an event can have various adverse effects. Firstly, cells may quickly leach out of the bioreactor because they are no longer held in place by the ATF. This can lead to a decrease in the bioreactor's cell density and productivity. Secondly, in systems integrated with downstream continuous purification processes, cells can flow directly into the operation and damage the purification system. For example, cells can travel onto a column on a periodic continuous counter-chromatography (PCC) skid, leading to clogging, pressurization, and increased impurity intrusion. In such situations, the problem must be identified quickly, and the ATF must be replaced promptly to protect the bioreactor. Independently, it is likely that downstream operations will need to be stopped and cleaned, and disposable components may need to be replaced. A full response may result in several days of production loss, and in the worst case, it could lead to a complete shutdown. [Overview of the project] [Problems that the invention aims to solve]
[0008] Apparatus and methods for cell culture are provided that overcome one or more of the drawbacks known in the art. It has been found that it is possible to prepare a filtration system for a perfusion cell culture apparatus that automatically detects and responds to malfunctions of the ATF filter without stopping the cell culture operation. [Means for solving the problem]
[0009] In a first embodiment of the present disclosure, a perfusion cell culture apparatus is provided. The apparatus includes a bioreactor vessel, a first filtration assembly, a second filtration assembly, and a controller. The bioreactor vessel is configured to receive a liquid medium. The first filtration assembly is in fluid communication with the bioreactor vessel and includes a first filtration system, a first harvest pump, and a sensor. The first harvest pump is connected in series with the first filtration system and is configured to pump the liquid medium from the bioreactor vessel through the first filtration system. The sensor is configured to interact with the liquid medium within the first filtration assembly. A second filtration assembly is also in fluid communication with the bioreactor vessel and is configured to operate in parallel with the first filtration assembly. The second filtration assembly includes a second filtration system and a second harvest pump. The second harvest pump is connected in series with the second filtration system and is configured to pump the liquid medium from the bioreactor vessel through the second filtration system. The controller performs operations. The operation includes receiving information from a sensor indicating the operating status of the first filtration system. The operation further includes determining, based at least on the received information, whether the first filtration system is operational. The operation also includes, in response to the determination that the first filtration system is not operational, causing the first harvest pump to stop pumping the liquid medium through the first filtration system.
[0010] A second embodiment of the present disclosure provides a perfusion cell culture apparatus, which includes a bioreactor vessel, a filtration assembly, and a controller. The bioreactor vessel is configured to receive a liquid medium. The filtration assembly is in fluid communication with the bioreactor vessel and includes a filtration system, a harvest pump, and a sensor. The harvest pump is connected in series with the filtration system and is configured to pump the liquid medium from the bioreactor vessel through the filtration system. The sensor is configured to interact with the liquid medium within the filtration assembly. The controller performs an action, which includes receiving information from the sensor indicating the operational status of the filtration system. The action further includes determining, based at least on the received information, whether the filtration system is operational. In response to the determination that the filtration system is not operational, the action further includes causing the harvest pump to stop pumping the liquid medium through the filtration system.
[0011] In yet another embodiment of the present disclosure, a method is provided. This method involves a bioreactor container The method includes at least partially filling a container with cells and a liquid medium. The bioreactor container is in fluid communication with a first filtration system and a second filtration system. The second filtration system is connected in parallel with the first filtration system. The method further includes using a first harvest pump to pump the liquid medium from the bioreactor container through the first filtration system. The first harvest pump is connected in series with the first filtration system. The method also includes using a second harvest pump to pump the liquid medium from the bioreactor container through the second filtration system. The second harvest pump is connected in series with the first filtration system. The method further includes, as an additional, receiving information indicating the operating status of the first filtration system from a sensor configured to interact with the liquid medium. Furthermore, the method includes determining, at least based on the received information, whether the first filtration system is operational. Furthermore, this method includes, in response to a determination that the first filtration system is not in an operational state, causing the first harvest pump to stop pumping the liquid medium through the first filtration system.
[0012] Other aspects, embodiments, and implementations will become apparent from the following detailed description and claims, with reference to the accompanying drawings as appropriate. [Brief explanation of the drawing]
[0013] [Figure 1] This is a schematic diagram of a standard perfusion bioreactor configuration. [Figure 2] This is a schematic diagram of a perfusion cell culture apparatus having a dual filtration system and integrated capture according to one embodiment of the present disclosure. [Figure 3] This is a schematic diagram of a perfusion cell culture apparatus having a dual filtration system and integrated capture according to another embodiment of the present disclosure. [Figure 4] This is a schematic diagram of a perfusion cell culture apparatus configuration having a single filtration system according to another embodiment of the present disclosure. [Figure 5] This is a flowchart of a method according to one embodiment of the present disclosure. [Modes for carrying out the invention]
[0014] This disclosure generally relates to filtration systems and cell culture methods for cell culture apparatuses. The perfusion cell culture apparatus of this disclosure includes a bioreactor vessel and two or more filtration systems (e.g., alternating tangential flow filters) connected in parallel. Failure of any filter can be detected by an inline sensor, and an automated response system functions to isolate the malfunctioning filter by stopping the flow of liquid medium through the filter. The flow of medium through the remaining operational filters can be increased so that the rate of perfusion through the bioreactor remains relatively constant. The cell culture apparatus of this disclosure can prevent problems arising from filter failures in conventional perfusion bioreactors, thereby improving the long-term viability of cell cultures.
[0015] Exemplary methods, devices, and systems are disclosed. The terms “example” or “exemplary” are used in this disclosure to mean “acting as an example or illustration.” Any implementation or configuration disclosed as “exemplary” or “example” should not necessarily be construed as preferable or better than other implementations or configurations. Other implementations may be used and modified without departing from the scope of the subject matter presented in this disclosure.
[0016] Therefore, the exemplary implementations disclosed are not intended to be limiting. The components disclosed and illustrated in the figures can be arranged, replaced, combined, separated, and designed in a wide variety of different configurations contemplated in this disclosure.
[0017] Furthermore, unless otherwise indicated in the context, the configurations shown in each figure are meant to be used in combination with each other. Therefore, these diagrams should generally be viewed as components of one or more overall implementation forms, and it should be understood that not all configurations shown are necessarily required for each implementation form.
[0018] In an effort to provide technical content for this disclosure, the information in this chapter can broadly describe the various components of the disclosed implementation. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as those generally understood by those skilled in the art. Such information is provided solely for the benefit of the reader and does not expressly limit the subject matter of the claims. Furthermore, the components shown in the figures are for illustrative purposes only. Therefore, these examples should not be construed as limiting. To be understood, components may be added, deleted, or rearranged without departing from the scope of this disclosure.
[0019] I. Overview In certain applications, it is advantageous to provide a cell culture apparatus that automatically detects and responds to filtration system failures. Such a cell culture apparatus may include built-in redundancy so that a second and / or further filtration system can be used if a first filter fails (e.g., by rupture, clogging, contamination, or some other means). When a filter failure occurs, the automated response system can function to increase the flow rate through any remaining filters while stopping the flow of medium to the affected filter. This response can allow the perfusion of the liquid medium to continue at nearly the same rate without requiring immediate intervention by the cell culture apparatus operator. Such a system can improve cell viability and reduce the probability of failure in the perfusion bioreactor system.
[0020] The devices and methods of the present disclosure generally relate to perfusion cell culture devices having an improved tangential flow filtration system. The device can include a bioreactor vessel configured to contain a cell population and a liquid growth medium. Fresh liquid medium can be provided to the bioreactor vessel, and spent liquid medium can be removed through a series of filtration assemblies at substantially equal rates. Each filtration assembly can include a filtration system, a harvest pump, a sensor, and / or a guard filter.
[0021] During normal operation, spent liquid medium is removed through one or more of the filtration assemblies and pumped to a downstream capture operation. Sensors provided within each filtration assembly continuously monitor the operation of the filtration assembly and can determine that a malfunction (e.g., rupture, clogging, or some other failure mode) has occurred in one of the filtration systems.
[0022] When a malfunction is detected by one or more of the sensors, an automated response system acts to change the operating parameters of the device so that perfusion of the liquid medium continues without interruption. The automated response system is implemented by a controller in communication with the sensors and the harvest pumps. In response to determining that one or more of the filtration systems is not in an operable state, the controller can cause the harvest pump associated with the inoperable filter to stop pumping liquid medium to the affected filter. Additionally, the controller can cause the remaining operable filtration assemblies to process an increased amount of liquid medium, for example, by increasing the flow of liquid medium to the operable filtration systems to the associated harvest pumps. The controller can also cause the device to output a notification, such as an alarm, indicator light, or some other alert. The controller can also stop the filtration system (e.g., by causing the tangential flow pump of the filtration system to stop pumping liquid medium across the filter membrane.) In some scenarios The controller can also stop the downstream capture operation or activate a valve to stop the flow of the liquid medium and / or divert the liquid medium to a waste container. In yet another scenario, the controller can completely stop the flow in and out of the bioreactor vessel if a bioreactor sensor (e.g., capacitance, optical density, oxygen uptake rate, etc.) falls below a certain threshold. Other automated responses are contemplated.
[0023] II. Cell Culture Device A cell culture device according to one embodiment of the present disclosure is shown in FIG. 2. The cell culture device 200 includes a bioreactor vessel 210, a continuous capture operation 260, and at least a first filtration assembly 206 and a second filtration assembly 208 connected in parallel between at least one outlet of the bioreactor vessel 210 and the inlet of the continuous capture operation 260. The bioreactor vessel 210 of the cell culture device 200 is configured to receive a fresh liquid medium to maintain a cell population within the vessel. As used herein, the liquid medium can refer to a nutrient growth medium or a culture medium designed to support the growth of cells. Substantially equal flows of spent liquid medium are pumped from the bioreactor vessel 210 through the plurality of filtration assemblies 206, 208, thereby removing metabolic and protein waste materials while maintaining a constant volume of medium within the bioreactor vessel 210. To monitor the cell culture environment, a bioreactor sensor 215 can be positioned to interact with the liquid medium within the bioreactor vessel 210. Such a bioreactor sensor 215 can be configured to measure the temperature, dissolved oxygen, pH, pCO2, and / or cell density of the bioreactor vessel 210 and / or the liquid medium within the bioreactor vessel 210.
[0024] The cell culture apparatus 200 can be integrated with a continuously operating capture operation 260 configured to collect products (e.g., desired biological or therapeutic products) from a liquid medium. The capture operation 260 is in fluid communication with the bioreactor vessel 210 by at least a first filtration assembly 206. After passing through parallel filtration assemblies 206, 208, the capture operation 260 can receive the used liquid medium, process and / or purify the medium to incorporate the desired product of interest. In various embodiments, the capture operation 260 can incorporate the product of interest using various techniques. For example, in some examples, the capture operation 260 may include a continuous chromatography system including one or more chromatography columns. In certain examples, the capture operation 260 may include a periodic continuous counter-chromatography (PCC) skid or a simulated moving bed (SMB) skid. In other examples, the capture operation 260 may include a multi-chromatography (MCC) skid (i.e., a continuous chromatography system with multiple columns). Those skilled in the art can consider other purification methods.
[0025] Before reaching the continuous capture operation 260, the liquid medium flows through one or more filtration assemblies 206, 208 designed to retain cells within the bioreactor vessel 210. Each filtration assembly 206, 208 is in fluid communication with the bioreactor vessel 210 and can be connected to the outlet of the vessel 210 between the vessel 210 and the continuous capture operation 260. As shown in Figure 2, in some examples, two filtration assemblies can be connected in parallel to provide two parallel fluid paths. The first filtration assembly 206 includes a first filtration system 220, a first harvest pump 230, a first sensor 240, and a first guard filter 250. Similarly, the second filtration assembly 208 includes a second filtration system 222, a second harvest pump 232, a second sensor 242, and a second guard filter 252. The elements of each filtration assembly 206, 208 can be connected in series via one or more conduits, sterilized tubing, or other connecting means. As shown in Figure 2, the first harvest pump 230 can be connected downstream of the first filtration system 220, and the sensor 240 is located downstream of the first harvest pump 230. It can be positioned between the first harvest pump 230 and the guard filter 250 connected further downstream. In this context, “downstream” refers to the relative position or orientation of the elements with respect to the flow of the liquid medium through the filtration assemblies 206, 208, and “downstream” generally refers to the direction away from the bioreactor vessel 210 toward the capture operation 260. However, the elements of the first filtration assembly 206 can be connected in any number of configurations. A second filtration assembly 208 and / or further filtration assemblies can be positioned, and the second filtration assembly 208 and / or further filtration assemblies can have elements that are substantially the same as or completely different from those of the first filtration assembly 206.
[0026] In some examples, the second filtration assembly 208 is configured to operate in parallel (i.e., simultaneously) with the first filtration assembly 206. During normal operation of the cell culture apparatus 200, the flow rates of the liquid medium through the first filtration assembly 206 and the second filtration assembly 208 can be approximately equal. In other words, each filtration assembly 206, 208 can receive approximately equal flow rates of used liquid medium from the bioreactor vessel 210. However, in other embodiments, a single filtration assembly (e.g., the first filtration assembly 206) can operate at a time, and the second filtration assembly 208 and / or further filtration assemblies can operate as backups.
[0027] In some examples, a third or further filtration assembly can be connected in parallel to the first filtration assembly 206 and the second filtration assembly 208 to increase processing capacity, provide additional pre-filtration means in case of system failure, or provide some other benefit.
[0028] The first filtration assembly 206 and the second filtration assembly 208 each include a first filtration system 240 and a second filtration system 242, respectively. The first and second filtration systems 220 and 222 can be configured to allow the free passage of cellular waste and one or more desired products of interest while preventing cells in the bioreactor vessel 210 from entering the continuous capture operation 260. Hereinafter, the term “filtration system” is used to refer to filter-based means for retaining cells in the bioreactor vessel 210. In some examples, the first filtration system 220 and the second filtration system 222 may include filters that function by allowing a fluid (e.g., a liquid medium) to flow tangentially across a filter membrane. Such filtration systems 220 and 222 may include conventional tangential flow filtration (TFF) systems, cross-flow filters, and similar cell retention means. In another example, at least one of the first filtration system 220 and the second filtration system 222 includes an alternating tangential flow (ATF) filter. Such a filter typically includes a filter membrane and a tangential flow pump (e.g., a diaphragm pump) configured to guide fluid tangentially across the filter membrane. The tangential flow pump may include a diaphragm that operates to flow a liquid medium from the bioreactor vessel 210 across the surface of the filter membrane in a repeating back-and-forth pattern. Repeated perturbation of the fluid near the filter membrane can prevent cell accumulation and minimize contamination of the filter surface during operation. In some examples, the first tangential flow pump provided by the first filtration system can operate independently of the second tangential flow pump provided by the second filtration system. However, in other embodiments, two or more tangential flow pumps can be controlled uniformly, and their pumping operations can be fully synchronized or asynchronous.
[0029] Regarding the first filtration assembly 206 and the second filtration assembly 208, ATF and While the above has been described insofar as it includes other tangential flow-based filters, other filtration means can also be implemented to retain cells within the bioreactor vessel 210. For example, in other embodiments, the first filtration system 220 and the second filtration system 222 may include internal spin filters (ISFs), hollow fiber filters, porous membrane filters, depth filters, and / or other microfiltration or ultrafiltration systems.
[0030] The first filtration system 220 and the second filtration system 222 each include a first filter membrane and a second filter membrane, respectively. The pore size of the filter membrane can be selected to allow desired products of interest, cellular waste, and spent liquid media to flow freely through the membrane while the cells remain contained within the bioreactor vessel 210. The filter membranes of the filtration systems 220 and 222 can be made of any material having a suitable pore size, such as a porous polymer membrane. In some examples, the filter membrane consists of a series of selectively permeable hollow fibers organized in a parallel arrangement inside the housing or cartridge. The pore size of the hollow fiber membrane can be selected to achieve the desired permeability of the filtration systems 220 and 222. In some examples, the first filtration system 220 and the second filtration system 222 each include pore sizes ranging from about 500 kilodaltons to about 10 microns, or more preferably from about 0.1 microns to about 1 micron.
[0031] Each filtration assembly 206, 208 in the apparatus of the present disclosure includes harvest pumps 230, 232. The first filtration assembly 206 includes a first harvest pump 230 connected in series with a first filtration system 220. Similarly, the second filtration assembly 208 includes a second harvest pump 232 connected in series with a second filtration system 222. The first harvest pump 230 and the second harvest pump 232 are configured to pump the liquid medium from the bioreactor vessel 210 through the first filtration system 220 and the second filtration system 222, respectively, into the downstream capture operation 260. In various examples, the first harvest pump 230 and / or the second harvest pump 232 may include peristaltic pumps, bearingless pumps, diaphragm pumps, or centrifugal pumps. However, other pumps may also be used to maintain the sterile state of the liquid medium while drawing the medium through the filtration assemblies 206, 208.
[0032] The first harvest pump 230 pumps liquid medium from the bioreactor vessel 210 through the first filtration system 220 at a first flow rate. Similarly, the second harvest pump 232 pumps liquid medium from the bioreactor vessel 210 through the second filtration system 222 at a second flow rate. In some examples, the first flow rate can be approximately equal to the second flow rate. In other words, the first harvest pump 230 and the second harvest pump 232 can be configured to pump approximately equal amounts of liquid medium through their respective filtration systems 220 and 222, so that the filtration assembly processes equal amounts of spent liquid medium under normal operating conditions. However, in other embodiments, the first and second flow rates may be different. In certain examples, the first harvest pump 230 can operate independently, and the second harvest pump 232 (i.e., a second or further harvest pump connected to a second or further filtration device) can be configured to operate in response to a determination that the first harvest pump 230, the first filtration system 220, and / or the first filtration assembly 206 are not in an operational state.
[0033] The flow rate of the liquid medium through the filtration assemblies 206 and 208 can be made approximately equal to the flow rate of new medium entering the bioreactor vessel 210, so that a stable amount of liquid medium is maintained in the vessel 210 during perfusion. In other words, the sum of the first flow rate and the second flow rate can be made approximately equal to the flow rate of liquid medium entering the bioreactor vessel 210 (i.e., the flow rate of medium introduced into the vessel 210 by the supply operation).
[0034] The flow rates of the liquid medium through each filtration system 220, 222 (i.e., the first flow rate and the second flow rate) can be independently controlled by a controller 270 communicating with the first harvest pump 230 and the second harvest pump 232. The controller 270 can be configured to increase, decrease, or stop the flow of the liquid medium through the first filtration system 240 and / or the second harvest pump 232 depending on the operating conditions of the apparatus 200. For example, if one or more of the filtration systems 220, 222 fail, the power to the harvest pump of the affected assembly can be cut off, and the flow rate of the liquid medium through the remaining filtration assemblies can be increased to compensate for the malfunctioning filtration assembly.
[0035] Perfusion bioreactor systems are typically configured to operate over longer periods, but equipment failures are anticipated. In particular, filtration systems 220 and 222 are susceptible to malfunction, which can cause problems in the downstream capture operation 260, increasing impurities in the harvest and potentially leading to complete shutdown. To prevent cells that have breached filtration systems 220 and 222 from interfering with the continuous capture operation 260, guard filters 250 and 252 can be connected in series with the first filtration system 220 and / or the second filtration system 222. Such guard filters 250 and 252 can be configured to allow free diffusion of nutrients, metabolites, and the liquid medium while preventing cells and cell fragments from advancing downstream to the capture operation 260. As shown in Figure 2, the guard filter 250 can be connected in series with the first filtration system 220 and / or the first filtration assembly 206. In some examples, the guard filter 250 is connected downstream of the first filtration system 220 and / or sensor 240. The second filtration assembly 208 may also include a guard filter (i.e., a second guard filter 252) connected in series with the second filtration system 222 downstream of the filtration system 222. However, in other embodiments (as shown in Figure 3), multiple filtration assemblies may be brought together into a single shared guard filter located, for example, at the inlet of the continuous capture operation 260. Other locations and configurations of the guard filter 250 are also possible.
[0036] The guard filters 250 and 252 can be made of any material having a suitable porosity, such as porous polymer membranes and / or hollow fiber membranes. In some cases, the guard filter 250 has a pore size of about 500 kilodaltons to about 10 microns, or more preferably about 0.1 microns to about 1 micron. The guard filter 250 can be configured to have a pore size approximately equal to that of the first filtration system 220 and / or the second filtration system 222 (i.e., to mimic the selective permeability of the filtration systems 220 and 222). However, in other cases, the guard filter 250 can have a pore size smaller or larger than that of the filtration systems 220 and 222, thereby enabling the selective filtration of additional metabolic products.
[0037] Depending on the circumstances, at least one sensor 240 may be provided within the filtration assemblies 206, 208 for monitoring the operation of one or more filtration systems 220, 222 and / or detecting filter failures. In particular, the first filtration assembly 206 may include a sensor 240. In some examples, the second filtration assembly 208 may include an additional sensor (e.g., a second sensor 242). The sensor 240 may be located along the conduits of the filtration assemblies 206, 208 and may be configured to interact with the liquid medium within the filtration assemblies 206, 208, i.e., to detect various aspects of the liquid medium passing through the filtration assemblies 206, 208. As shown in Figure 2, the sensor 240 may be located downstream of the first filtration system 220 and upstream of the guard filter 250. Depending on the circumstances, the sensor 240 may be a pressure sensor, or more This may include a piezoelectric resistance pressure sensor. Sensor 240 may be configured to detect pressure in the first filtration assembly 206 (e.g., pressure of the liquid medium in the first filtration assembly 206) which can indicate clogging of the guard filter 250 and / or rupture in the first filtration system 220 upstream. In various examples, sensor 240 may be configured to detect pressure upstream or downstream of the first guard filter 250, pressure upstream or downstream of the first filtration system 220, and / or membrane pressure of the first filtration system 220 or the guard filter 250. In other examples, sensor 240 may be an optical sensor (e.g., an optical density probe), and sensor 240 may be configured to determine the cell density in the liquid medium flowing through the first filtration assembly 206. In further examples, sensor 240 may include a flow meter that can operate to measure the flow rate of the liquid medium through the filtration systems 220, 222 and / or filtration assemblies 206, 208. In further examples, sensor 240 may include a capacitive sensor, a Raman sensor, or an FTIR (Fourier Transform Infrared) sensor. In another example, sensor 240 may be integrated into a continuous acquisition operation (for example, as a pressure sensor upstream of the acquisition column). Other sensor types and applications are also possible.
[0038] While the sensors 240 and 242 in Figure 2 are shown as being contained within the first filtration assembly 206 and the second filtration assembly 208 (i.e., located in the conduits between the filtration systems 220 and 222 and the guard filters 250 and 252), the sensors can be placed in various locations. For example, in some examples, the bioreactor sensor 215 can be configured to interact with the liquid medium within the bioreactor vessel 210, so that the bioreactor sensor 215 can monitor changes in the pressure, temperature, pH, dissolved oxygen, pCO2, cell density, or other properties of the liquid medium within the vessel 210.
[0039] To mitigate potential problems arising from filter failures, the cell culture apparatus 200 may include an automated response system. This response system can be used to detect filtration system failures (e.g., filter membrane rupture), isolate the malfunctioning filtration system, and / or adjust the operating parameters of the apparatus 200 in order to maintain relatively stable cell perfusion. The automated response system can be implemented by a controller 270 communicating with at least the harvest pumps 230, 232 and sensors 240, 242. As described herein, the controller 270 may include at least one processor configured to perform actions (e.g., actions stored as program instructions in the controller's data storage). However, in other examples, the controller 270 may include a comparator or another simplified control system.
[0040] The controller 270 can be configured to receive information from sensors 240, 242 indicating the operating status of the first filtration system 220 and / or the second filtration system 222. Such information may include characteristics of the liquid medium flowing through the first filtration assembly 206 and / or the second filtration assembly 208. For example, this information may include the fluid pressure of the liquid medium in the filtration assemblies 206, 208, the cell density of the liquid medium, the flow rate of the liquid medium, the presence of a subject in the liquid medium, or any other information. In another example, as previously mentioned, the bioreactor sensor 215 may be configured to collect information about the liquid medium in the bioreactor container 210, such as cell density or other characteristics.
[0041] The controller 270 can then determine, based on information received from at least sensors 240 and 242, whether the first filtration system 220 and / or the second filtration system 222 are operational. In this context, the term “operational” means, as intended, that the liquid medium containing cellular waste and products of interest is selectively filtered. This term is used to refer to a filtration system that functions to retain cells within the bioreactor vessel 210 while allowing them to pass through. Conversely, filtration systems 220, 222 that are not operational may be unable to pass a desired or expected flow rate of liquid medium (i.e., indicating a clogged or contaminated filter membrane) or unable to retain cells (i.e., indicating a ruptured filter). Determining whether filtration systems 220, 222 are operational may include determining whether the information received is within the expected range of values or above or below a predetermined threshold.
[0042] In a specific example, the first filtration system 220 may have ruptured, thereby allowing cells to break through the first filtration system 220 and accumulate on the guard filter 250. The controller 270 can then receive information from the sensor 240 indicating that the pressure in the liquid medium has increased due to the cells clogging the guard filter 250. Determining whether the first filtration system 220 is operational at this point may include determining whether the pressure in the first filtration assembly 206 (i.e., the pressure of the liquid medium in the first filtration assembly 206) is greater than a threshold. In a similar example, one or more of the sensors 240 may include optical sensors (e.g., optical density probes), and determining whether the filtration system is operational may include determining that the cell density in the liquid medium in the first filtration assembly 206 is above a threshold level, indicating that cells have broken through the first filtration system 220. In yet another example, the bioreactor sensor 215 could be an optical sensor, and determining whether the filtration systems 220, 222 are operational could include determining whether the cell density of the liquid medium in the bioreactor container 210 is below a threshold level.
[0043] Other failure modes are also anticipated. For example, the first filtration system 220 may become inoperable due to a failure of the first tangential flow pump and / or fouling of the filter membrane (for example, if the first filtration system 220 is an ATF or other tangential flow-based filter). In such a situation, cells may accumulate on the filter membrane, blocking or reducing the flow of liquid medium through the first filtration system 220. In such an example, the sensor 240 may be configured to measure the membrane pressure of the first filtration system 220. Determining whether the first filtration system 220 is operational may include determining that the membrane pressure of the filtration system 220 is above a threshold. In a further example, the sensor 240 may include a flow meter, and determining whether the first filtration system 220 is operational may include determining that the flow of liquid medium through the first filtration system 220 (and / or the first filtration assembly 206) is below a threshold level. Those skilled in the art can consider other failure modes, sensor parameters, and determinants.
[0044] After detecting one or more filtration system failures, the automated response system (i.e., the controller 270) can function to isolate the malfunctioning filtration assemblies 206, 208 by preventing the liquid medium from flowing through the affected filtration systems 220, 222. In other words, in response to a determination that the first filtration system 220 is not operational, the controller 270 can cause the first harvest pump 230 to stop pumping the liquid medium through the first filtration system 220, for example by cutting off power to the harvest pump 230. The controller 270 can further configure the controller 270 to shut down the first filtration system 220 by causing the tangential flow pump of the filtration system 220 to stop pumping the liquid medium across the filter membrane (i.e., if the filtration system 220 contains ATF). In other examples, the controller 270 can activate a relief valve connected in series with the first filtration system 220 or the second filtration system 222. The system can be configured to divert the liquid medium to the waste collection system. In yet another scenario, the controller 270 can completely stop the flow in and out of the bioreactor container 210 if the information from the bioreactor sensor 215 (for example, the capacitance, optical density, oxygen absorption rate, or other aspect of the liquid medium in the bioreactor container 210) exceeds or falls below a certain threshold.
[0045] In addition or alternatively, the first filtration assembly 206 may include one or more first isolation valves 265 connected in series with the first filtration system 220. The first isolation valves 265 may be configured to control the flow of liquid medium through the first filtration system 220. In response to a determination that the first filtration system 220 is not operational, the controller 270 may be configured to operate the first isolation valves 265 to close. A second isolation valve 267 may be further connected in series with the second filtration system 222, and the second isolation valve 267 may be configured to control the flow of liquid medium through the second filtration system 222. As shown in Figure 2, the isolation valves 265, 267 may optionally be connected in series between the first filtration assembly 206 and / or the second filtration assembly 208 and the continuous capture operation 260. However, one or more such isolation valves 265, 267 can be located anywhere before, after, or within the first filtration assembly 206 or the second filtration assembly 208. For example, the first isolation valve 265 and / or the second isolation valve 267 can be located between the bioreactor vessel 210 and the respective filtration assemblies 206, 208, downstream of the filtration systems 220, 222, downstream of the guard filters 250, 252, or connected to the inlet of the continuous capture operation 260. When a filter malfunction is detected, the isolation valves 265, 267 can be activated to prevent the flow of liquid medium and cellular products through the affected filtration system 220, 222, thereby preventing cells and contaminants from reaching the continuous capture operation 260 and facilitating the replacement of any faulty equipment.
[0046] In addition to isolating the failed filtration systems 220 and 222, the controller 270 can also be configured to change various operating parameters of the cell culture apparatus 200 after a filter failure. As previously stated, a stable flow rate of liquid medium should be supplied to the bioreactor vessel 210, and used medium containing waste can be continuously removed at approximately equal flow rates to promote cell growth in the bioreactor vessel 210. After detecting a failure in the first filtration system 220, the controller 270 can, in addition, be configured to increase the flow rate of liquid medium through the remaining operational second filtration system 222 so that a stable perfusion of liquid medium is maintained in the bioreactor vessel 210. The increased flow rate through the remaining operational filtration system (e.g., the second filtration system 222) should be approximately equal to the total pre-failure flow rate through the original filtration systems 220 and 222 (e.g., the sum of the first flow rate through the first filtration system 220 and the second flow rate through the second filtration system 222). In one example, the controller 270 may, in response to a determination that the first filtration system 220 is not operational, enable the second harvest pump 232 to operate to increase the flow rate of the liquid medium through the second filtration system 242. In some examples, enabling the second harvest pump 232 to increase the flow rate of the liquid medium through the second filtration system 242 may include doubling the flow rate.
[0047] Depending on the configuration, the cell culture apparatus 200 may include multiple filtration assemblies operating simultaneously and connected in parallel (i.e., at least a first filtration assembly 206 and a second filtration assembly 208). For example, in some embodiments, the cell culture apparatus 200 may include three, four, six, or more filtration assemblies to achieve a desired perfusion rate. Filtration systems 220, 228 of filtration assemblies 206, 208 When one or more of the two systems fail, the controller 270 can be activated to adjust the flow rate through any number of remaining operational filter assemblies to maintain a stable perfusion of the liquid medium in the system. For example, in response to determining that the first filtration system 220 is not operational, the controller 270 can increase the flow rate of the liquid medium through one or more harvest pumps associated with one or more further filtration assemblies.
[0048] In some cases, it is desirable to notify the operator of the cell culture apparatus 200 when a filter malfunction is detected. For example, in response to a determination that the first filtration system 220 is not in an operational state, the controller 270 can be configured to output a notification. Such a notification may include a visual alert, such as flashing light, colored light, a visual message, or text or graphic information on a display. In addition or alternatively, the notification may include an auditory alert, such as an alarm sound, buzzer sound, bell sound, auditory message, or some other auditory component. In yet another example, the notification may include a text message, a phone call, or an email sent to a recipient associated with the cell culture apparatus.
[0049] The cell culture apparatus 200 shown in Figure 2 includes two filtration assemblies 206, 208, each containing separate filtration systems 220, 222, sensors 240, 242, and guard filters 250, 252; however, simpler cell culture apparatuses can also be included within the scope of this disclosure. For example, Figure 3 shows a cell culture apparatus 300 that includes dual filtration systems 320, 322, each containing a shared sensor 340, guard filter 350, and a capture operation 360. The first filtration system 320 is connected in series with the first harvest pump 330, and the second filtration system 322 is connected in series with the second harvest pump 332. Such a configuration can enable the sensor 350 to detect an overall failure of the cell culture apparatus 300. For example, if a rupture occurs in the first filtration system 320 or the second filtration system 322, cells from the bioreactor vessel 310 may immediately flow through the filtration systems 320, 322 and collect on the guard filter 350. Clogging of the guard filter 350 causes an increase in the pressure of the liquid medium upstream of the guard filter 350, which can be detected by a pressure sensor (e.g., sensor 340). However, because the downstream sensor 340 is shared, such a system cannot distinguish between a fault in the first filtration system 320 and a fault in the second filtration system 322. In such an exemplary system, an automated response system could function to alert the operator of the cell culture apparatus 300, for example by outputting a notification, to divert the liquid medium to the waste collection system or to stop the continuous capture operation. Other automated responses are also conceivable.
[0050] The bioreactor container 310, the first and second filtration systems 320 and 322, the first and second harvest pumps 330 and 332, the sensor 340, the guard filter 350, and the capture operation 360 shown in Figure 3 can be configured in the same manner as the corresponding components described above in relation to Figure 2.
[0051] In some other examples, a cell culture apparatus may rely on a single filtration assembly for handling used liquid media. Figure 4 shows an example of such a cell culture apparatus 400, where a bioreactor vessel 410 is fluidly communicated to a capture operation 460 by a single filtration system 420, a harvest pump 430, a sensor 440, and a guard filter 450. A controller 470 can be communicatively connected to at least the sensor 440 and the harvest pump 430. In such an example, a filter malfunction (e.g., rupture, clogging, or other defect of the filtration system 420) could render the entire filtration mechanism of the cell culture apparatus 400 inoperable. Filtration system 42 In response to the determination that 0 is not in an operational state, the controller 470 can enable the capture operation 460 to stop collecting products from the liquid medium.
[0052] In situations where all filtration assemblies of the cell culture apparatus are not operational (for example, when only one filtration system 420 fails, or when all of multiple filtration systems fail), it is advantageous to divert the flow of liquid medium to the waste collection system 480. To provide such a scenario, one or more filtration assemblies may include a relief valve 485 connected in series with the filtration system 420. The relief valve 485 can be configured to divert the flow of liquid medium to the waste collection system 480. The relief valve 485 may be a three-way valve, but in other examples, the relief valve 485 may include one or more two-way valves. Such a relief valve 485 may preferably be connected downstream of the harvest pump 430 and upstream of the guard filter 450, but various positions can be considered. In response to a determination that the filtration system 420 is not operational, the controller 470 may be configured to open the relief valve 485, thereby diverting the liquid medium to the waste.
[0053] By diverting the liquid medium to the waste collection system 480, a temporary continuation of perfusion through the cell culture system 400 can be permitted. However, as more cells are eventually pumped to the waste, the cell density in the bioreactor vessel 410 may drop to an unacceptable level. To prevent an undesirable loss of cell density, sensors (e.g., sensor 440 or additional bioreactor sensors) can be configured to interact with the liquid medium in the bioreactor vessel 410 and / or monitor the cell density in the vessel 410. In response to a determination that the cell density falls below a threshold, the controller 470 can be configured to completely stop perfusion by, for example, closing the relief valve 485, closing the isolation valve, causing one or more harvest pumps 430 to stop pumping the liquid medium through one or more filtration systems 420, or by some other means.
[0054] The bioreactor vessel 410, filtration system 420, harvest pump 430, sensor 440, guard filter 450, capture operation 460, and controller 470 shown in Figure 4 can be configured similarly to the corresponding components described above in relation to Figure 2.
[0055] III. Cell culture method Figure 5 is a flowchart of Method 500 for operating a cell culture apparatus, such as one of the apparatuses described herein, shown in Figure 2, Figure 3, or Figure 4. For illustrative purposes, the cell culture apparatus operating in Method 500 includes: (i) a bioreactor vessel; (ii) a first filtration system in fluid communication with the bioreactor vessel; (ii) a second filtration system in fluid communication with the bioreactor vessel; (iii) a first harvest pump connected in series with the first filtration system; (iv) a second harvest pump connected in series with the second filtration system; and (v) a sensor configured to interact with a liquid medium flowing through at least the first filtration system.
[0056] Block 501 of Method 500 includes at least partially filling a bioreactor vessel with cells and a liquid medium. The bioreactor vessel is in fluid communication with a first filtration system and a second filtration system connected in parallel to the first filtration system. At least partially filling the bioreactor vessel with cells and a liquid medium can include providing cells as a monolayer on an artificial substrate (i.e., adherent culture). In other examples, at least partially filling the bioreactor vessel with cells and a liquid medium can include providing cells freely suspended in the medium (i.e., suspension culture). The liquid medium for perfusion can be provided by a supply operation configured to continuously introduce new liquid medium into the bioreactor vessel. The liquid medium from the supply operation can be provided at a predetermined flow rate adjusted to the nutritional needs of the cells in the bioreactor vessel.
[0057] Block 502 of Method 500 includes pumping a liquid medium from a bioreactor vessel through a first filtration system using a first harvest pump. Block 503 of Method 500 includes pumping a liquid medium from a bioreactor vessel through a second filtration system using a second harvest pump. The first harvest pump can be connected in series with the first tangential flow. Similarly, the second harvest pump can be connected in series with the second filtration system.
[0058] Pumping the liquid medium through the first filtration system may include pumping the liquid medium at a first flow rate. Pumping the liquid medium through the second filtration system may include pumping the liquid medium at a second flow rate. In some examples, the first flow rate may be substantially equal to the second flow rate under normal operating conditions. However, in other examples, the flow rate of the liquid medium through the first filtration system may be different from the flow rate of the liquid medium through the second filtration system. In addition or alternatively, the sum of the first and second flow rates may be approximately equal to the flow rate of the medium entering the bioreactor vessel by a supply operation (e.g., the supply operation described above in relation to block 501 of method 500).
[0059] Block 504 of Method 500 includes receiving information indicating the operating state of the first filtration system from a sensor configured to interact with a liquid medium flowing through at least the first filtration system. Such a sensor may be located within a filtration assembly including the first filtration system, the first harvest pump, and / or other elements. Additionally or alternatively, the sensor may be located within a conduit connected in series with the first filtration system and / or the first harvest pump.
[0060] In some examples, the sensor is a pressure sensor (e.g., a piezoelectric pressure sensor). Information indicating the operating state of the first filtration system may include information about the pressure of the liquid medium flowing through the first filtration system. More specifically, the information may include the pressure of the liquid medium upstream of the first filtration system, the pressure of the liquid medium downstream of the first filtration system, or the membrane differential pressure of the liquid medium in the first filtration system. In some examples, the cell culture apparatus includes a guard filter connected in series with the first filtration system downstream of the first filtration system. In such examples, the information may include the pressure of the liquid medium upstream of the guard filter. Additionally or alternatively, the sensor may include a capacitance sensor, a Raman probe, an FTIR probe, or an optical density probe. In such examples, the information may include information about the cell density of cells in the liquid medium (e.g., the liquid medium downstream of the first filtration system).
[0061] Block 505 of Method 500 includes determining whether the first filtration system is operational based on at least the information received. When the first filtration system is not operational, the filter may not be able to allow the liquid medium to pass through at the desired or expected flow rate (i.e., indicating clogging or contamination of the filter membrane) or may not be able to hold cells (i.e., indicating that the filter has burst). Determining whether the filtration system is operational may include determining whether the information received from the sensor is within the expected range of values or is above or below a predetermined threshold. In some examples, the sensor is a pressure sensor, and determining whether the first filtration system is operational includes determining whether the pressure of the liquid medium is above a predetermined threshold. In some cases, determining whether the first filtration system is operational may include Determining whether the first filtration system is operational may include determining that the membrane pressure of the first filtration system exceeds a predetermined threshold. In a further example, determining whether the first filtration system is operational may include determining that the cell density of the liquid medium (e.g., the liquid medium in the bioreactor vessel or the liquid medium flowing through the first filtration system) is outside a predetermined threshold range.
[0062] Block 506 of Method 500 includes, in response to a determination that the first filtration system is not operational, causing the first harvest pump to stop pumping the liquid medium through the first filtration system. Such a response can effectively stop the flow of the liquid medium through the first filtration system. Additionally or alternatively, an isolation valve may be connected in series with the first filtration system, and the isolation valve may be configured to control the flow of the liquid medium through the first filtration system. In such an example, Method 500 may include closing the isolation valve in response to a determination that the first filtration system is not operational. Other responses to filter malfunctions are also expected.
[0063] In some examples, Method 500 may further include adjusting the flow rate of one or more remaining operational filters. For example, Method 500 may further include increasing the flow rate of the liquid medium through the second filtration system in response to a determination that the first filtration system is not operational. Increasing the flow rate of the liquid medium through the second filtration system can allow the perfusion of the liquid medium through the bioreactor vessel to continue at a stable rate after filter failure. In other words, the adjusted flow rate of the liquid medium through the second filtration system can be made approximately equal to the flow rate of the liquid medium through the operational first and second filtration systems. In some examples, increasing the flow rate of the liquid medium through the second filtration system in the second harvest pump may include doubling the flow rate of the liquid medium through the second filtration system.
[0064] In some examples, Method 500 further includes outputting a notification in response to a determination that the first filtration system is not in an operational state. The notification may include a visual alert (e.g., flashing light, colored light, visual message, character or graphic information on a display) and / or an auditory alert (e.g., an alarm sound, buzzer sound, bell sound, auditory message, or any other auditory component). In further examples, Method 500 may include diverting the liquid medium to a waste collection system, stopping a downstream capture operation, or affecting other aspects of the operation of the cell culture apparatus.
[0065] The exemplary method 500 shown in Figure 5 is meant to be illustrative and non-limiting. The blocks and steps described herein can be performed sequentially or in parallel. Furthermore, various blocks and steps can be performed in a different order than those described herein, and some blocks and steps can be deleted, omitted, and / or repeated. Additional or alternative elements of the method and additional or alternative components of the system are also contemplated.
[0066] While the apparatus and methods disclosed herein have been described in terms of various embodiments, it will be understood that variations and modifications will be conceivable to those skilled in the art. Therefore, the appended claims are intended to encompass all such equivalent variations within the scope of the claimed apparatus and methods. Additionally, the item names used herein are for organizational purposes only and should not be construed as limiting the subject matter described.
[0067] All references cited in this application are expressly incorporated herein by reference.
Claims
1. A perfusion cell culture device: (a) A bioreactor container configured to receive a liquid medium containing cell cultures; (b) A first filtration assembly that is in fluid communication with the bioreactor vessel, the: (i) First filtration system; (ii) A first harvest pump connected in series with the first filtration system and configured to pump the liquid medium from the bioreactor container through the first filtration system; and (iii) a first filtration assembly comprising a sensor configured to interact with the liquid medium within the first filtration assembly; (c) A second filtration assembly configured to operate in parallel with the first filtration assembly, which is in fluid communication with the bioreactor vessel: (i) a second filtration system; and (ii) A second filtration assembly including a second harvest pump connected in series with the second filtration system and configured to pump the liquid medium from the bioreactor vessel through the second filtration system; (d) A controller: (i) From the sensor, temperature, dissolved oxygen, pH, pCO2 in the first filtration system 2 , receiving information indicating changes in capacitance, optical density, and / or oxygen absorption rate; (ii) Based on the information received from the sensor, the temperature, dissolved oxygen, pH, and pCO2 in the first filtration system 2 , determining changes in capacitance, optical density, and / or oxygen absorption rate; and (iii) Temperature, dissolved oxygen, pH, pCO2 in the first filtration system 2 , in response to a determination of changes in capacitance, optical density, and / or oxygen absorption rate, to cause the first harvest pump to stop pumping the liquid medium through the first filtration system; and (iv) In response to the determination of changes in temperature, dissolved oxygen, pH, pCO2, capacitance, optical density, and / or oxygen absorption rate in the first filtration system, the second harvest pump increases the flow of the liquid medium through the second filtration system. A controller that performs an action including The perfusion cell culture apparatus, including the above.
2. The perfusion cell culture apparatus according to claim 1, wherein at least one of the first filtration system and the second filtration system includes an alternating tangential flow filter.
3. The perfusion cell culture apparatus according to claim 1, wherein at least one of the first filtration system and the second filtration system includes a tangential flow filter.
4. The perfusion cell culture apparatus according to claim 1, wherein each of the first filtration system and the second filtration system includes a filter membrane having a pore size of about 0.1 microns to about 1 micron.
5. The sensor measures temperature, dissolved oxygen, pH, and pCO2. 2 , including capacitance, optical density, and / or oxygen absorption rate sensors, Temperature, dissolved oxygen, pH, pCO2 in the first filtration system 2 Determining changes in capacitance, optical density, and / or oxygen absorption rate is possible in the first filtration assembly, which is determined by the temperature, dissolved oxygen, pH, and pCO2. 2 This includes determining whether the capacitance, optical density, and / or oxygen absorption rate are outside a predetermined threshold range. The perfusion cell culture apparatus according to claim 1.
6. The sensor measures temperature, dissolved oxygen, pH, and pCO2 in the first filtration system. 2 The perfusion cell culture apparatus according to claim 1, which detects an increase in capacitance, optical density, and / or oxygen absorption rate.
7. The sensor measures temperature, dissolved oxygen, pH, and pCO2 in the first filtration system. 2 The perfusion cell culture apparatus according to claim 1, which detects a decrease in capacitance, optical density, and / or oxygen absorption rate.
8. The perfusion cell culture apparatus according to claim 1, wherein increasing the flow of liquid medium through the second filtration system to the second harvest pump comprises doubling the flow rate by approximately twofold.
9. The controller is: Temperature, dissolved oxygen, pH, pCO 2 , capacitance, optical density, and / or the perfusion cell culture device according to claim 1, further configured to output a notification in response to a determination of a change in the oxygen absorption rate.
10. The perfusion cell culture apparatus according to claim 9, wherein the notification includes a visual alert or an auditory alert.
11. The system further includes an isolation valve connected in series with the first filtration assembly, the valve being configured to control the flow of a liquid medium through the first filtration assembly, and the controller is: Temperature, dissolved oxygen, pH, pCO2 in the first filtration system 2 The perfusion cell culture apparatus according to claim 1, further configured to close the isolation valve in response to a determination of a change in capacitance, optical density, and / or oxygen absorption rate.
12. The perfusion cell culture apparatus according to claim 1, wherein the first filtration assembly further includes a guard filter connected in series with the first filtration system, the guard filter being connected downstream of the first filtration system and the sensor.
13. The perfusion cell culture apparatus according to claim 12, wherein the guard filter has a pore size of approximately 0.1 microns to approximately 1 micron.
14. The perfusion cell culture apparatus according to claim 1, further comprising a capture operation configured to collect products from a liquid medium, wherein the capture operation is in fluid communication with a bioreactor vessel by at least a first filtration assembly, and the capture operation comprises multi-column chromatography (MCC).
15. A method for culturing cells: (a) a step of at least partially filling a bioreactor vessel with cells and a liquid medium, wherein the bioreactor vessel is in fluid communication with a first filtration system and a second filtration system connected in parallel with the first filtration system; (b) A step of pumping a liquid medium from a bioreactor vessel through a first filtration system using a first harvest pump, wherein the first harvest pump is connected in series with the first filtration system; (c) A step of pumping a liquid medium from a bioreactor vessel through a second filtration system using a second harvest pump, wherein the second harvest pump is connected in series with the second filtration system; (d) From a sensor configured to interact with a liquid medium, the cell density, temperature, dissolved oxygen, pH, and pCO2 of the first filtration system are measured. 2 , a process of receiving information indicating changes in capacitance, optical density, and / or oxygen absorption rate; (e) Based on information received from at least the sensors, the cell density, temperature, dissolved oxygen, pH, and pCO2 in the first filtration system 2 , a step of determining changes in capacitance, optical density, and / or oxygen absorption rate; (f) Cell density, temperature, dissolved oxygen, pH, pCO2 in the first filtration system 2 electrostatic A step of causing a first harvest pump to stop pumping the liquid medium through a first filtration system in response to a determination of changes in volume, optical density, and / or oxygen absorption rate; (g) Cell density, temperature, dissolved oxygen, pH, pCO2 in the first filtration system 2 electrostatic The process involves, in response to the determination of changes in volume, optical density, and / or oxygen absorption rate, increasing the flow rate of the liquid medium through the second filtration system in the second harvest pump. The method, including the method described above.
16. The method according to claim 15, wherein at least one of the first filtration system and the second filtration system includes an alternating tangential flow filter.
17. The sensor measures cell density, temperature, dissolved oxygen, pH, and pCO2. 2 The method according to claim 15, comprising a capacitance, optical density, and / or oxygen absorption rate sensor.
18. The information received included the cell density, temperature, dissolved oxygen, pH, and pCO2 of the liquid medium downstream of the first filtration system. 2 The method according to claim 17, comprising information relating to capacitance, optical density, and / or oxygen absorption rate.
19. Cell density, temperature, dissolved oxygen, pH, pCO2 of the first filtration system 2 The steps for determining changes in capacitance, optical density, and / or oxygen absorption rate include the cell density, temperature, dissolved oxygen, pH, and pCO2 of the liquid medium. 2 The method according to claim 17, comprising determining that capacitance, optical density, and / or oxygen absorption rate exceed predetermined thresholds.
20. The method according to claim 15, wherein the step of pumping a liquid medium from a bioreactor vessel through a first filtration system includes pumping the liquid medium at a first flow rate, and the step of pumping a liquid medium from a bioreactor vessel through a second filtration system includes pumping the liquid medium at a second flow rate, the first flow rate being substantially equal to the second flow rate.
21. The method according to claim 15, wherein the step of increasing the flow rate of the liquid medium through the second filtration system to the second harvest pump includes doubling the flow rate of the liquid medium through the second filtration system.
22. Cell density, temperature, dissolved oxygen, pH, pCO2 in the first filtration system 2 The process involves outputting a notification in response to the determination of changes in capacitance, optical density, and / or oxygen absorption rate: The method according to claim 15, further comprising: