Production system and method for disposable high pathogenic microorganisms or inactivated vaccines

Abstract

The present application relates to the field of bioengineering, and particularly to a production system and method for disposable high-pathogenic microorganism or inactivated vaccine. The method comprises: providing a self-definable assembly bio-culture module and a corresponding management module; the bio-culture module comprises: a bio-reaction device, an intermediate device and a purification device connected through detachable pipelines, and a liquid leakage sensing line is arranged on the pipelines; when the bio-culture module is started, a first set composed of readings on the liquid leakage sensing line is collected; the management module judges the risk state of the bio-culture module through the first set and controls the safe operation thereof. The present application provides a disposable production method and system suitable for high-pathogenic viruses, bacteria and inactivated systems, which has the advantages of flexible assembly, automatic leakage warning, sterilization and the like, and the cumbersome processes (such as centrifugation, cleaning and the like) in the fixed production method are omitted, thereby effectively saving the research and development time and cost.

Description

Technical Field

[0001] This invention belongs to the field of bioengineering technology, specifically relating to a production system and method for single-use highly pathogenic microorganisms or inactivated vaccines. Background Technology

[0002] Currently, the cultivation of microorganisms or biological products such as vaccines typically employs fixed biological culture systems. However, these traditional fixed culture systems have certain drawbacks in practical applications, especially when used for small-scale microbial cultivation or pilot-scale processes in the early stages of vaccine development. Fixed culture systems suffer from relatively complex operation processes (involving tedious steps such as sample collection, centrifugation, waste disposal, and system cleaning), high application costs, and poor flexibility. For example, patent application CN201780082359.X discloses a system for producing cells and / or cell products. Another example is patent application CN201880076583.2, which discloses a system and method for preparing biomolecules (such as viral vaccines). Both of these applications attempt to reduce the tedious manual operations associated with fixed culture to some extent through automated control. However, for the early stages of vaccine development (or the early stages of microbial cultivation), the production process (such as the selected processing steps and the set processing sequence) may need to be adjusted according to the specific needs at the time. However, for stationary production systems, adjusting the system configuration in real time according to changes in the process is extremely difficult and costly. In other words, existing vaccine production systems and automation systems still have many limitations when applied to small-scale vaccine trials.

[0003] For example, patent application CN201980085256.8 discloses a method for purifying a biomolecule production system and a system suitable for purification. This system attempts to simplify cumbersome cleaning and disinfection steps by using disposable pipes or components. However, this production system still does not break away from the traditional fixed production model, and therefore, problems such as high application costs and poor flexibility in vaccine trials remain unresolved or unmitigated.

[0004] Furthermore, the disposable bioreactor involved in the bioculture system (or vaccine production system) is a novel type of bioreactor, typically made of disposable plastic or stainless steel. This novel bioreactor can generally be applied in fields such as cell culture, bacterial culture, virus culture, and the cultivation and extraction of effective substances such as monoclonal antibodies. However, existing bioreactors are usually used in large-scale cell or microbial cultures (such as commercial applications) and are difficult to meet the needs of small-scale experiments. For example, when researchers are in the early stages of vaccine development, they usually need to conduct multiple small-scale virus cultures in the laboratory first. However, existing biosafety reactors have certain shortcomings for small-scale virus culture in terms of cost, safety, and ease of operation. For example, Chinese invention patent CN102492607B discloses a disposable bioreactor system and method. The disposable bioreactor system includes: a disposable container with at least one inlet, at least one outlet, and at least one collection port, using sterile filters connected to all external openings to maintain the integrity of the sterile environment; a structure supporting the disposable container; one or more sensors for sensing one or more parameters of the biological material in the container; a heater for heating the contents of the container, the heater having a thermostat; and a mixing system integrated with the system to mix the biological material contained in the disposable container. This bioreactor system is relatively large in size, employing a large-capacity bioreactor and requiring a series of auxiliary equipment, resulting in high costs and greater demands on experimental operating space. Furthermore, there are significant differences between large-scale microbial or cell culture and micro-microbial or cell culture in practical applications. Therefore, even proportionally scaling down existing equipment is insufficient to meet the needs of micro-scale laboratory experiments. For example, Chinese invention patent application CN103945928A discloses a single-use mixing and bioreactor system, which will also face similar problems when applied to micro-scale experiments. To further adapt to the needs of micro-scale experiments, some small-scale reactor designs have been proposed in the prior art. For example, Chinese patent application CN102212474A discloses a small bioreactor comprising five parts: a cell culture tank, a temperature control kit, a liquid (gas) replenishment device, a cell optical density detection kit, and a culture control device. The cell culture tank is connected to the liquid replenishment device, and the temperature control kit, cell optical density detection kit, and culture control device work together to provide optimized growth conditions for cell culture. However, this type of small bioreactor requires steam sterilization after each reaction, which is cumbersome and costly for large-scale experiments.Furthermore, when this bioreactor is used to cultivate pathogenic microorganisms such as bacteria or viruses, it is difficult to ensure that the internal and external environments of the reactor are not disturbed (for example, pathogenic bacteria in the reactor may spill out and contaminate the experimental environment, or even threaten the safety of the operators).

[0005] Therefore, there is an urgent need for a vaccine production system that can be applied to small-scale microbial culture or vaccine pilot-scale processes. Summary of the Invention

[0006] The purpose of this invention is to provide a production system and method for single-use highly pathogenic microorganisms or inactivated vaccines.

[0007] This approach partially addresses or alleviates the aforementioned shortcomings of existing technologies, effectively improving microbial culture efficiency and reducing culture costs. Specifically, the following technical solution is adopted: a method for producing single-use highly pathogenic microorganisms or inactivated vaccines, including:

[0008] S001 provides a customizable bioculture module and a management module for controlling the operating status of the bioculture module; wherein, the bioculture module includes: a bioreactor for culturing microorganisms, an intermediate device for processing liquid containing microorganisms, a purification device for purifying liquid, and a collection device, wherein the intermediate device is connected to any one or both of the bioreactor and purification devices via at least one detachable pipe for collecting liquid from the bioreactor and / or purification device and processing the liquid, wherein at least one leakage sensing line is arranged along the length of the detachable pipe; S002 when the bioculture module is turned on, a first set consisting of at least one reading on at least one leakage sensing line is collected in real time; S003 a first risk state of the bioculture module is determined based on the first set, the first risk state including: leakage object, and / or leakage risk level; S004 the management module generates a corresponding control signal or instruction based on the risk state and sends it to the bioculture module; S005 the bioculture module responds to the control signal or instruction by selecting to continue production, suspend production, or stop production.

[0009] In some embodiments, S004 includes the step of: the management module determining whether the number of leaking objects exceeds a preset first threshold; if so, the management module sends a first control signal indicating a halt to production to the biological culture module, wherein,

[0010] The first control signal includes a stop signal indicating the cessation of production and a disinfection signal indicating the initiation of disinfection; otherwise, the management module sends a second control signal to the biological culture module indicating the suspension of production. The second control signal includes the object to be suspended; wherein the object to be suspended includes the leaking object and one or more detachable pipes and devices connected to the leaking object.

[0011] In some embodiments, the purification apparatus includes one or more of the following: an ultrafiltration deactivation device, an ultrafiltration liquid exchange device, a clarification filtration device, and a supernatant concentration device; the intermediate device includes one or more of the following: a chromatography device and a mixing device.

[0012] In some embodiments, the device is provided with a diaphragm pump or peristaltic pump for conveying liquid at the interface between the device and the detachable pipe.

[0013] In some embodiments, the reading includes: current value, and S003 includes the step of: when the current value of the detachable pipe is greater than a preset second threshold, a first leakage signal including first risk status information is generated, and the first leakage signal is associated with one or two devices connected to the detachable pipe; correspondingly, S004 includes the step of: 1) when the management module detects that the device is associated with two first leakage signals, the corresponding device and detachable pipe are identified as first leakage objects and a second control signal indicating suspension of production is generated; wherein, the second control signal includes: first leakage object information. Alternatively, in other embodiments, the following is also included: 3) When a first leakage signal is detected associated with a device, the corresponding device is identified as the second leakage object and the detachable pipe is identified as the first leakage object, and a first alarm signal is sent to the user; 4) The management module monitors whether the user issues a corresponding instruction within a first alarm time after the first alarm signal is issued; wherein the instruction includes: a first instruction indicating cancellation of the alarm, and / or, a second instruction indicating receipt of the alarm, and / or, a third instruction indicating correction of the alarm, and the third instruction includes: the corrected leakage object; if yes, the received instruction is sent to the biological culture module; if no, a second control signal indicating suspension of production is sent directly to the biological culture module.

[0014] In some embodiments, the detachable pipe includes: an internal pipe for conveying fluid, and an external pipe for protecting the internal pipe; wherein at least one first leakage sensing line is provided on the outer side of the internal pipe along its length, and at least one second leakage sensing line is provided on the outer side of the external pipe along its length; and the reading includes: the current value of the leakage sensing line, and correspondingly, step S003 includes: (i) determining that the internal pipe is leaking when the current value of the internal pipe is detected to be greater than a preset second threshold; (ii) generating a second leakage signal associated with the detachable pipe, the second leakage signal including: pipe information, and / or leakage risk level; and (iii) determining that the external pipe is leaking when the current value of the external pipe is detected to be greater than a preset second threshold; (iv) generating A third leakage signal associated with the pipeline is generated, the third leakage signal including: pipeline information, and / or leakage risk level; accordingly, S004 includes: when the management module detects that the detachable pipeline is associated with both the second and third leakage signals, the detachable pipeline is identified as a leakage object, and a first control sub-signal indicating a suspension of production is generated and sent to the biological culture module based on the leakage object; or, when the management module only detects that the detachable pipeline is associated with the second leakage signal, a second alarm signal is issued to the user; or, when the management module detects that the detachable pipeline is associated with only the third leakage signal, the leakage risk level of the external pipeline and device is determined based on the reading, and a second control sub-signal indicating a suspension of production is generated and sent to the biological culture module based on the leakage risk level.

[0015] In some embodiments, the concentration and filtration device includes: a first chamber for filtration, and a second chamber disposed on the outer layer of the first chamber. The first chamber contains a filter medium for filtering the liquid, and one end of the first chamber is connected to a bioreactor or a purification device, while the other end of the first chamber can be connected to other purification devices, intermediate devices, or collection devices. Leakage sensing units are respectively disposed on the outer walls of the first and second chambers, and the leakage sensing units are leakage sensors or leakage sensing lines. In some embodiments, the alarm signal includes one or more of the following forms: voice alarm, and / or SMS alarm, and / or signal light alarm.

[0016] In some embodiments, the step of determining the leakage risk level of pipe leakage and device leakage based on readings includes: determining the distribution characteristics of the leakage location of the internal pipe based on the readings, wherein the distribution characteristics include: continuous distribution connected to both sides of the pipe, and / or regional distribution disconnected from both sides of the pipe; when the distribution characteristic is detected as continuous distribution, the leakage risk level at the interface of the device on both sides of the pipe is determined to be Level 1, and the leakage risk level of the pipe is determined to be Level 2; when the distribution characteristic is detected as regional distribution, the leakage risk level of the pipe is determined to be Level 1, and the leakage risk level of the device on both sides of the pipe is determined to be Level 2; correspondingly, S004 further includes the step of: generating a third alarm signal based on the leakage risk level information and sending it to the user or a second control sub-signal to be sent to the biological culture module.

[0017] In some embodiments, the first risk state further includes: gas leakage information, and the production method further includes the steps of: S006 when the biological culture module is turned on, collecting the gas around the biological culture module; S007 detecting the gas to obtain a first index value of the gas; S008 determining whether the biological culture module has leaked gas based on the first index value, and generating a gas leakage signal if so; correspondingly, S004 includes: 6) when the management module detects that the biological reactor is a leaking object and simultaneously detects a gas leakage signal, sending a first control signal indicating that production should be stopped to the biological culture module; and 7) when the management module detects that the biological reactor is a leaking object but does not detect a gas leakage signal, sending a second control signal to the biological culture module to suspend production; and 8) when the management module detects a gas leakage signal but the biological reactor is not at risk of leaking liquid, sending a fifth alarm signal to the user to prompt the user to check the gas leakage signal.

[0018] In some embodiments, prior to S002, the steps further include: introducing liquid into the biological culture module such that the liquid flows sequentially through the bioreactor, the purification device, and the collection device; acquiring a second set consisting of at least one reading on at least one leakage sensing line; determining a third risk state of the biological culture module based on the second set, the third risk state including: a leakage object; generating and sending a fourth alarm signal to the user based on the third risk state.

[0019] In some embodiments, the bioreactor includes: a bioreactor bag for culturing microorganisms, a first pressure monitoring unit for monitoring the hydraulic pressure in the bioreactor bag, and a first leakage monitoring module correspondingly disposed at the bottom of the bioreactor bag. The method includes the steps of: S102 introducing a preset amount of gas or liquid into the bioreactor bag through a first inlet or a second inlet on the bioreactor bag for culturing microorganisms or cells; S104 monitoring the hydraulic pressure of the first pressure monitoring unit and the leakage value of the first leakage monitoring module in real time; S106 determining a second risk state of the bioreactor based on the hydraulic pressure and the leakage value, the risk state including: a safe state, and / or a pending state, and / or a leakage state; wherein, S106 includes: S61 determining the pressure drop risk of the bioreactor bag based on the change parameters of the hydraulic pressure; wherein, when When the changing parameter falls within the preset first pressure drop threshold, the pressure drop risk is level one; when the changing parameter falls within the preset second pressure drop threshold, the pressure drop risk is level two; otherwise, the pressure drop risk is level zero. S62 determines the leakage risk of the reaction bag based on the leakage value. When the leakage value falls within the preset first leakage threshold, the leakage risk is level one; when the leakage value falls within the preset second leakage threshold, the leakage risk is level two; otherwise, the leakage risk is level zero. S63 determines the second risk state based on the pressure drop risk in S61 and / or the leakage risk in S62. When the pressure drop risk is level two and / or the leakage risk is level two, the second risk state is a leakage state. When both the pressure drop risk and the leakage risk are level zero, the second risk state is a safe state; otherwise, the second risk state is a pending state.

[0020] This embodiment provides a bioreactor suitable for small-scale batch cultivation of highly pathogenic microorganisms. The bioreactor combines an internally circulating condensate guiding path with a limited combination of key parameters (such as internal hydraulic pressure and external leakage value) for monitoring, providing a highly sensitive and low-cost bioreactor module suitable for cultivating highly pathogenic microorganisms. In some embodiments, a condensate guiding zone is formed in the top region of the first space within the reaction bag to guide condensate to condense within the zone and flow back to the second space via the guiding path provided by the condensate guiding zone. Correspondingly, the method further includes the steps: S108 When a risk state of pending status or leakage status is detected, a corresponding warning signal is issued to the user; S110 In response to the feedback signal issued by the user, secondary monitoring of the pressure reduction risk and / or leakage risk of the bioreactor is performed, or the warning signal is received or corrected. This embodiment also provides a monitoring method that enables coordinated manual and automated monitoring for the selected key parameter group. This monitoring method utilizes limited dual-source data to simultaneously monitor and comprehensively evaluate the actual working state of the bioreactor, thereby reducing manual intervention. This method can also automatically correct or review comprehensive assessment results (such as risk status) to further improve the sensitivity and accuracy of the monitoring method (reducing the issuance of erroneous signals). Furthermore, by combining it with an internally circulating condensate water guiding function, it can reduce the interference of condensate water on pressure changes, thereby helping to improve the monitoring accuracy of pressure sensors (such as the first pressure monitoring unit).

[0021] A second aspect of the invention is that it also provides a production system for disposable highly pathogenic microorganisms or inactivated vaccines.

[0022] The system includes: a customizable bioculture module and a management module for controlling the operating status of the bioculture module; wherein the bioculture module includes: a bioreactor for culturing microorganisms, an intermediate device for processing liquid containing microorganisms, a purification device for purifying liquid, and a collection device, wherein the intermediate device is connected to any one or both of the bioreactor and purification devices via at least one detachable pipe for collecting liquid from the bioreactor and / or purification device and processing the liquid, wherein at least one leakage sensing line is arranged along the length of the detachable pipe; a leakage reading acquisition module is configured to acquire a first set of readings from at least one leakage sensing line in real time when the bioculture module is running; a leakage reading processing module is configured to determine a first risk state of the bioculture module based on the first set, the first risk state including: leakage object and / or leakage risk level; and a management module is configured to generate corresponding control signals or instructions based on the first risk state and send them to the bioculture module; wherein the bioculture module responds to the control signals or instructions by selecting to continue production, suspend production, or stop production.

[0023] In some embodiments, the reading includes a current value, and the leakage reading processing module is further configured to generate a first leakage signal including first risk status information when the current value of the detachable pipe is greater than a preset second threshold, and the first leakage signal is associated with one or two devices connected to the detachable pipe; correspondingly, the management module includes a first monitoring unit, which identifies the corresponding device and detachable pipe as first leakage objects and generates a second control signal indicating a production halt when the management module detects that the device is associated with two first leakage signals; wherein the second control signal includes first leakage object information; and / or, the second monitoring unit is configured to generate a first leakage signal when the current value of the detachable pipe is greater than a preset second threshold. When a first leakage signal is detected associated with a device, the corresponding device is identified as the second leakage object, and the detachable pipe is identified as the first leakage object, and a first alarm signal is sent to the user; the third monitoring unit is configured to monitor whether the user issues a corresponding instruction within a first alarm time after the first alarm signal is issued; wherein the instruction includes: a first instruction indicating cancellation of the alarm, and / or a second instruction indicating receipt of the alarm, and / or a third instruction indicating correction of the alarm, and the third instruction includes: the corrected leakage object; if yes, the received instruction is sent to the biological culture module; if no, a second control signal indicating suspension of production is sent directly to the biological culture module.

[0024] Beneficial Technical Effects: Unlike traditional fixed culture methods, this invention proposes a disposable vaccine production line that can be flexibly assembled and dynamically maintained. This disposable production line allows staff to flexibly assemble and replace the entire line during the early stages of process design and later stages of vaccine production (i.e., dynamic maintenance). For example, when staff need to verify or adjust the vaccine pilot process in the early stages of vaccine development, they can assemble the various devices or pipelines in a laboratory environment (specifically within a biosafety isolator) using a quick-disassembly method to freely configure the disposable production line. Furthermore, the multi-segment leakage signal monitoring method adopted in this invention can accurately monitor and locate risks to the production line, thereby assisting users in performing "dynamic maintenance" of the production line to ensure its stable operation.

[0025] The disposable production line provided in this invention is particularly suitable for small-scale microbial culture (such as 2-20L culture of highly pathogenic viruses and bacteria) or vaccine pilot-scale processes. Attached Figure Description

[0026] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. In all the drawings, similar elements or parts are generally identified by similar reference numerals. The elements or parts in the drawings are not necessarily drawn to scale. Obviously, the drawings described below are some embodiments of the present invention, and those skilled in the art can obtain other drawings based on these drawings without any creative effort.

[0027] Figure 1 This is a schematic diagram of the first structure of a disposable bioreactor in an exemplary embodiment of the present invention;

[0028] Figure 2 This is a schematic diagram of the second structure of a disposable bioreactor in an exemplary embodiment of the present invention;

[0029] Figure 3 This is a schematic diagram of the internal structure of a disposable bioreactor in an exemplary embodiment of the present invention;

[0030] Figure 4 This is a schematic diagram of the structure of the condensate guiding region in an exemplary embodiment of the present invention;

[0031] Figure 5 This is a schematic diagram of the top wall structure in another exemplary embodiment of the present invention;

[0032] Figure 6 This is a schematic diagram of the first structure of the support device in an example embodiment of the present invention;

[0033] Figure 7 This is a schematic diagram of the second structure of the support device in an example embodiment of the present invention;

[0034] Figure 8 This is a cross-sectional schematic diagram of the sensor quick-connect device in an exemplary embodiment of the present invention;

[0035] Figure 9 This is a schematic diagram of the sensor quick-connect device in an exemplary embodiment of the present invention;

[0036] Figure 10 This is a flowchart illustrating an exemplary embodiment of the automated control method of the present invention;

[0037] Figure 11 This is a flowchart illustrating an automated control method in another exemplary embodiment of the present invention;

[0038] Figure 12 This is a schematic diagram of the modules of an automated control system in an exemplary embodiment of the present invention;

[0039] Figure 13 This is a schematic diagram of a production or purification method in an exemplary embodiment of the present invention;

[0040] Figure 14 This is a schematic diagram of the configuration of a production system for inactivated bacteria in a specific embodiment of the present invention;

[0041] Figure 15a This is a schematic diagram of a leaking pipe in an exemplary embodiment of the present invention;

[0042] Figure 15b This is a first schematic diagram of the detachable pipe and device interface in an exemplary embodiment of the present invention;

[0043] Figure 15c This is a second schematic diagram of the detachable pipe and device interface in an exemplary embodiment of the present invention;

[0044] Figure 16 This is a schematic diagram of an apparatus module for a production method in an exemplary embodiment of the present invention.

[0045] Reference numerals: 1 is the reaction bag, 11 is the top wall, 12 is the annular guide rail, 12-1 is the annular groove, 13 is the first space, 14 is the second space; 2 is the stirring device, 21 is the stirring paddle, 22 is the stirring shaft, 23 is the heating unit; 3 is the working status monitoring module; 4 is the first inlet; 5 is the first outlet; 6 is the second outlet, 61 is the drying device, 62 is the filtering device; 7 is the supporting body, 71 is the leakage monitoring sensor, 72 is the base, 73 is the first side wall, 74 is the second side wall; 81 is the disposable sensor probe, 82 is the probe terminal, 83 is the power terminal, 84 is the electrode; L1 is the horizontal plane, L2 is the guide path, 91 is the detachable pipe, 91-1 is the first external interface, 91-2 is the first internal interface, 92 is the second interface, 92-1 is the second external interface, 92-2 is the second internal interface, 92-3 is the threaded structure. Detailed Implementation

[0046] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some, not all, of the embodiments of the present invention. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention without creative effort are within the scope of protection of the present invention.

[0047] In this document, the suffixes such as "module," "part," or "unit" used to denote elements are used only for the purpose of illustrative purposes and have no specific meaning in themselves. Therefore, "module," "part," or "unit" can be used interchangeably. In this document, the terms "upper," "lower," "inner," "outer," "front," "rear," and "one end" are used interchangeably.

[0048] The orientation or positional relationship indicated by terms such as "the other end" is based on the orientation or positional relationship shown in the accompanying drawings and is only for the convenience of describing the invention and simplifying the description, and does not indicate or imply that the device or element referred to must have a specific orientation, or be constructed and operated in a specific orientation, and therefore should not be construed as a limitation of the invention. Furthermore, the terms "first" and "second" are used for descriptive purposes only and should not be construed as indicating or implying relative importance. In this document, unless otherwise expressly specified and limited, the terms "installed," "equipped with," "connected," etc., should be interpreted broadly. For example, "connection" can be a fixed connection, a detachable connection, or an integral connection; it can be a mechanical connection, a direct connection, or an indirect connection through an intermediate medium; it can be a connection within two elements. Those skilled in the art can understand the specific meaning of the above terms in this invention based on the specific circumstances. In this document, "and / or" includes any and all combinations of one or more of the listed related items. In this document, "a plurality of" means two or more, that is, it includes two, three, four, five, etc. It should be noted that, in this document, the terms "comprising," "including," or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such a process, method, article, or apparatus. Without further limitation, an element defined by the phrase "comprising one..." does not exclude the presence of other identical elements in the process, method, article, or apparatus that includes that element. As used in this specification, the term "about" typically means + / -5% of the value, more typically + / -4%, more typically + / -3%, more typically + / -2%, even more typically + / -1%, even more typically + / -0.5% of the value. In this specification, certain embodiments may be disclosed in a range-bound format. It should be understood that such "range-bound" descriptions are merely for convenience and brevity and should not be construed as a rigid limitation of the scope of disclosure. Therefore, the description of a range should be considered as having specifically disclosed all possible subranges and the independent numeric values ​​within those ranges. For example, range The description should be considered as having specifically disclosed subranges such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6, etc., as well as individual numbers within this range, such as 1, 2, 3, 4, 5, and 6. The above rules apply regardless of the breadth of the range.

[0049] In this article, "communication connection" can refer to a direct communication connection between two or more parties, or it can be established through a third party (e.g.,

[0050] Indirect communication connection is achieved through a central control module or other central control system. In this paper, the "leakage sensing line" can also be called a "leakage sensing rope" or "leakage detection line," which consists of a sensing cable for detecting liquid leakage and a controller with a location alarm. Of course, depending on the actual application requirements, the biological culture module in this paper can use either a location-based or non-location-based leakage sensing line. In this paper, "reading" can refer to the current value or current change value at one or more points (or one or more segments) in the leakage sensing line.

[0051] "Suspension of production" refers to suspending one or more devices within the entire bioculture module (i.e., bioculture system). This suspension can be a complete halt to the bioculture module or a suspension of only some devices. For example, in some embodiments, when a leakage risk is detected in the purification device, the operation of all devices in the bioculture module can be suspended until the faulty device is replaced, at which point the bioculture module can be restarted. Alternatively, in other embodiments, when a leakage risk is detected in the purification device, only the purification device can be suspended and replaced, while the bioreactor (e.g., bioreactor) continues to cultivate microorganisms. In this document, "index value" refers to the concentration of one or more gaseous components in a gas (such as collected waste gas or air).

[0052] The term "indicator value" can also refer to the difference in the change of one or more gaseous components within a gas over a certain period of time. For example, for some gas-producing microorganisms, such as Clostridium perfringens, the corresponding indicator value is the concentration of carbon dioxide or the increase in carbon dioxide over a certain period of time; similarly, if the microorganisms being cultured are hydrogen-producing bacteria or methanogens, the corresponding indicator values ​​are the concentration of hydrogen and the concentration of methane, respectively. In this article, "inactivated vaccine" refers to various vaccines made by culturing pathogenic microorganisms and then inactivating or reducing their virulence to varying degrees. For example, "inactivated vaccine" can refer to an inactivated vaccine made by killing pathogenic microorganisms, rendering them non-pathogenic while retaining their antigenicity, or it can refer to a live attenuated vaccine made by reducing the virulence of pathogenic microorganisms, enabling them to induce an ideal immune response without producing clinical symptoms.

[0053] like Figures 1-5As shown, in a first aspect, the present invention provides a disposable biosafety bioreactor suitable for large-scale, miniaturized microbial or cell culture in a laboratory. The bioreactor includes: a reaction bag 1, the internal chambers of which sequentially include a first space 13 and a second space 14; wherein, when the bioreactor is in operation, the first space 13 is used to contain gases (e.g., air, oxygen, carbon dioxide, etc.), and the second space 14 is used to contain liquids (e.g., cell culture medium, water, etc.); a condensate guiding zone is also formed in the top region of the first space to guide condensate to condense in the condensate guiding zone and flow back to the second space through the guiding path provided by the condensate guiding zone; and a working status monitoring module 3 for monitoring the working status parameters of the bioreactor, including: liquid pressure, foam generation, dissolved oxygen rate, CO2 concentration, pH value, and turbidity; correspondingly, the working status monitoring module includes monitoring units: a first pressure monitoring unit for monitoring hydraulic pressure, a foam monitoring unit for monitoring foam generation, a dissolved oxygen rate monitoring unit for monitoring dissolved oxygen rate, a carbon dioxide monitoring unit for monitoring CO2 concentration, a pH monitoring unit for monitoring pH value, and a turbidity monitoring unit for monitoring turbidity;

[0054] The monitoring ends of each monitoring unit (e.g., the monitoring ends of sensors) are set on the wall of the second space to ensure accurate monitoring of various working state parameters of the liquid. The first space is provided with at least one first inlet 4 for introducing a first gas (e.g., compressed air, carbon dioxide, nitrogen, oxygen, etc.) and at least one second inlet for introducing a first liquid (e.g., cell culture medium, cultured bacteria, or water, etc.). Both the first and second inlets are located outside the condensate guiding area to prevent the first or second inlet from being blocked or contaminated by condensate during the reaction process. When the bioreactor is in operation (e.g., ... Figure 1 (as shown in the placement state), the opening of the first inlet is higher than that of the second inlet (or, in some other embodiments, the position of the second inlet may be unrestricted) to prevent water or other liquid substances discharged through the second inlet from flowing through the first inlet and causing blockage or contamination; the lower end region of the second space is also provided with at least one first outlet 5 for liquid sampling.

[0055] In other embodiments, taking into account the specific growth characteristics of microorganisms, some or all of the first inlet or the second inlet may be located on the wall of the second space 14, so that some gas or liquid samples can be directly introduced into the culture medium in the second space.

[0056] In some embodiments, the reaction bag is a disposable biological reaction bag, and the disposable biological reaction bag includes, from the inside out: a contact layer for direct contact with microorganisms or cell cultures, a barrier layer for isolating gases, and a structural layer for providing mechanical stability.

[0057] Specifically, in some embodiments, the volume of the reaction bag is preferably set to 1.5L, 2L, 2.5L or 5L.

[0058] Specifically, in some embodiments, the bioreactor bag is a disposable bag composed of three layers of plastic material: a first layer made of polyethylene terephthalate or LDPE to provide mechanical stability; a middle layer made of PVA or PVC to act as a gas barrier; and finally, a contact layer that comes into contact with the cell culture made of PVA or PP.

[0059] To ensure stable microbial or cell culture within a small-capacity bioreactor, it is essential to guarantee the stable operation of other structures associated with the bioreactor (such as a first inlet, a second inlet, etc.). The volume (or capacity) of the second space is typically smaller than that of the first space. Preferably, the second space occupies approximately 20%–50% of the internal chamber volume of the bioreactor.

[0060] For example, in some embodiments, when the bioreactor bag is 5L, the second space is approximately 2L. During the experiment,

[0061] It is necessary to ensure that the total amount of liquid added does not exceed the volume of the second space.

[0062] For example, in some embodiments, a boundary line is provided between the first space and the second space to determine whether the liquid addition is excessive. It is understood that the use of the terms "first space" and "second space" in this invention is merely a distinction between the internal chambers of the reaction bag from a spatial function perspective. Therefore, the first space and the second space can actually be unobstructed, interconnected spaces, such as... Figure 1 and Figure 3 As shown.

[0063] In some embodiments, such as Figure 4As shown, the top wall 11 inside the condensate guiding area is configured as an inclined surface, and an annular guide rail 12 is provided at the edge of the condensate guiding area. The annular guide rail 12 provides a guiding path L2 for the condensate to condense and flow back. For example, in some embodiments, the top of the first space (or only the condensate guiding area) is configured as an inclined surface with a certain angle. Specifically, the top of the first space can be configured as an inverted cone. For example, in some embodiments, an annular guide rail is provided at the edge of the condensate guiding area. When the air humidity in the first space is high, water droplets can preferentially condense on the inclined surface (i.e., the top wall 11). The water droplets flow downward along the inclined surface and collect inside the guide rail. As the collected water droplets gradually increase in size, they flow back downward into the second space under the action of gravity.

[0064] In some embodiments, the wall of the reaction bag inside the condensate guiding zone is configured as a slope, and the inclination angle α (or slope) of the slope is preferably set to 1°-5°. The inclination angle refers to the angle between the wall of the condensate guiding zone and the horizontal plane L1 when the bioreactor is placed vertically. Preferably, the inclination angle of the slope is 2-3°.

[0065] In some embodiments, such as Figure 5 As shown, the annular guide rail is designed with a gradually decreasing curvature from its first end to its second end (i.e., the condensate outlet end). Specifically, the curvature of each point on the guide path of the annular guide rail 12 gradually decreases along the direction from its first end to its second end, so that the second end of the guide path is lower than the first end, that is, the condensate outlet end is at the lowest position.

[0066] In some embodiments, such as Figure 3 As shown, the bioreactor also includes: a stirring device 2, which includes a stirring shaft 22 and a stirring paddle 21 disposed at the end of the stirring shaft; a mounting hole for installing the stirring device 2 is provided in the central area of ​​the condensate guiding zone, and the stirring shaft 22 is fixedly installed on the reaction bag through the mounting hole, while the stirring paddle 21 extends into the second space; wherein, a heating unit 23 (e.g., a constant temperature heating plate can be provided) is provided on the side of the stirring shaft 22 located or close to the condensate guiding zone to avoid the formation of condensate on the outside of the condensate guiding zone; the monitoring end of the working status monitoring module is higher than the horizontal height of the stirring paddle to reduce the interference of the stirring paddle on the working stability of the working status monitoring module. In this embodiment of the invention, in order to avoid the generation of excessive condensate in the first space due to excessively low temperature and excessively high humidity, so that the condensate may condense outside the condensate guiding zone or overflow from the condensate guiding zone, an electric heating plate is provided on the stirring shaft 22 near the condensate guiding zone and away from the liquid surface. For example, the electric heating plate can activate the heating function when the humidity is too high or the temperature is too low.

[0067] Understandably, since stirring devices are usually made of metal, when the heating function of the stirring device is not turned on,

[0068] A certain amount of condensate typically condenses at the top of the stirring shaft. Part of this condensate flows back to the second space along the stirring shaft, while the other part flows back to the second space along a guiding path under the influence of the condensate guiding zone. In some embodiments, the stirring device can be a magnetic stirring device. In this embodiment of the invention, to control the volume and application cost of the miniaturized bioreactor, an internal circulation condensate recirculation scheme (without external drainage pipes) is provided. This scheme avoids clogging or even contamination of the inlets and outlets by condensate during the reaction process, and further ensures the internal stability of the bioreactor (reducing liquid loss).

[0069] In some embodiments, such as Figure 4 As shown, the annular guide rail 12 has an annular groove 12-1 for providing a guide path L2, and a condensate outlet is provided on the annular groove 12-1. The bottom surface of the annular groove is inclined so that the condensate outlet is the lowest horizontal point of the annular groove, thereby guiding the condensate accumulated in the annular groove to flow out through the condensate outlet; wherein, the condensate outlet is arranged in a direction away from the first inlet and the second inlet. In this embodiment of the invention, the low outlet design of the inclined groove can further guide and promote the formation of condensate in a limited area (i.e., the condensate guiding area) and avoid condensate overflow due to excessive condensation. Especially when culturing highly pathogenic viruses or bacteria, the concentrated formation and reflux of condensate can ensure the stable progress of the experiment, and at the same time reduce the possibility of highly pathogenic viruses or bacteria overflowing through the pipe and contaminating the external environment.

[0070] In some embodiments, a first inlet for introducing gas (i.e., gas inlet) is provided with a first control valve (i.e., air inlet valve), a second inlet for introducing liquid (i.e., liquid inlet) is provided with a second control valve (i.e., liquid inlet valve), and a second pressure monitoring unit (i.e., air pressure monitoring unit) for monitoring the internal air pressure of the first space is provided in the first space, and the first pressure monitoring unit (i.e., hydraulic monitoring unit) is communicatively connected to the second control valve, and the second pressure monitoring unit is communicatively connected to both the first control valve and the second control valve.

[0071] The following describes part of the workflow of the air pressure monitoring unit, hydraulic monitoring unit, air inlet valve, and liquid inlet valve:

[0072] When the hydraulic monitoring unit detects that the hydraulic pressure exceeds a preset first pressure threshold, it sends a first shut-off signal to the inlet valve to indicate that the liquid inlet should be stopped. The inlet valve responds to the first shut-off signal and keeps the liquid inlet closed. When the pneumatic monitoring unit detects that the pneumatic pressure exceeds a preset second pressure threshold, it sends a second shut-off signal to the air inlet valve and the liquid inlet valve to indicate that the sample should be stopped. The air inlet valve and the liquid inlet valve keep the gas and liquid inlets closed based on the second shut-off signal.

[0073] In some embodiments, when hydraulic or pneumatic pressure is detected to exceed a preset pressure threshold, a warning signal can be sent directly to the user, allowing the user to intervene and determine whether to close or open each inlet.

[0074] To ensure the safety and reliability of miniaturized experiments (e.g., when the reaction bag capacity is only 0.5L, 1L, or 2L), this embodiment of the invention synchronously monitors air pressure and hydraulic pressure to avoid experimental failure or even reaction bag rupture due to excessive liquid or gas addition. Furthermore, in some embodiments, it also includes a first leakage detection module (e.g., a leakage sensor) corresponding to a location at the bottom of the reaction bag for monitoring whether the reaction bag leaks. Furthermore, in some embodiments, it also includes a safety disinfection module, which includes:

[0075] The invention includes a recovery unit and a sterilization unit connected to the reaction bag. The recovery unit comprises a recovery bag and a recovery pipe for connecting the recovery bag and the reaction bag, wherein a third control valve (i.e., a recovery valve) is installed in the recovery pipe. The sterilization unit comprises a chamber for storing sterilizing substances and a sterilization pipe for connecting the chamber and the reaction bag, wherein a fourth control valve (i.e., a sterilization valve) is installed in the sterilization pipe. Both the third and fourth control valves are communicatively connected to a pressure monitoring sensor and a first leakage monitoring module. In this embodiment of the invention, to control the amount of sterilizing substances used and to reduce the adverse effects of the sterilization process on the environment surrounding the reaction bag (e.g., the support device for supporting the reaction bag, or the biosafety cabinet where the reaction bag is located), a recovery-then-sterilization mode is adopted.

[0076] The following describes the preferred disinfection process for the safety disinfection module: Step 21: The safety disinfection module (such as a recovery valve or a disinfection valve) receives an opening signal (e.g., when leakage is detected in the reaction bag, such as when a leakage risk status is detected, or when a disinfection command is input by the user); Step 22: The recovery valve opens and draws the liquid in the second space into the recovery bag; After the liquid is completely recovered, or after a certain preset recovery time (e.g., 10 seconds),

[0077] Step 23: Close the recovery valve and open the sterilization valve. The sterilization valve will allow the sterilization substance in the chamber to enter the reaction bag.

[0078] In some embodiments, the recycling bag contains a disinfectant for sterilizing the liquid fluid. In some embodiments, the disinfectant is a second liquid or a second gas, such as hydrogen peroxide gas. In some embodiments, the recycling bag may employ a multi-layer design similar to or the same as that of a disposable bioreactor bag.

[0079] In some embodiments, after the entire bioreactor operation is completed, the safety disinfection module can disinfect the bioreactor in the same or similar manner as steps 21-23.

[0080] In this embodiment of the invention, to prevent further leakage of the liquid (containing highly hazardous microorganisms) inside the reaction bag during the filling of the disinfectant, which could even worsen the damage to the reaction bag, the liquid is recovered before filling with the disinfectant, and then disinfectant gas or liquid is introduced to centrally disinfect the reaction bag.

[0081] In some embodiments, a disinfectant substance may be introduced into the reaction bag through the first outlet.

[0082] In some embodiments, the device further includes a safety sampling device comprising: a sampling tube connected to the first outlet 5, a fifth control valve disposed within the sampling tube, and a plurality of sampling ports arranged axially on the sampling tube, wherein a disposable sampling bag is sealed to the sampling end of each sampling port, and the open end of the disposable sampling bag is made of thermoplastic plastic. After sampling is completed, the open end of the sampling bag can be heat-treated by a clamping heater, causing the sampling bag to automatically seal under heat treatment and detach from the sampling end of the sampling port for removal by the user.

[0083] In some embodiments, the reaction bag is connected to four gas inlets. Of course, the number and location of the gas inlets can be adjusted according to actual needs. In some embodiments, the control valves in the gas inlets are backflow preventers. In some embodiments, a second outlet 6 (i.e., an exhaust port) is also provided on the wall of the first space. The exhaust port is connected to an exhaust pipe, and a filter device (e.g., an H14 HEPA filter) is connected inside the exhaust pipe to filter aerosols inside the reaction bag and prevent pollution to the external environment. When the reaction bag is used in a bioreactor, the exhaust pipe can also be connected to the exhaust pipe of the bioreactor. In some embodiments, a drying device 61 is provided at the exhaust port. For example, a desiccant is provided inside the exhaust pipe to filter contaminants in the gas. Alternatively, in other embodiments, when an anhydrous disinfectant is used inside the filter device 62, the desiccant can be omitted.

[0084] Furthermore, in some embodiments, by monitoring key parameters of the liquid inside the reaction bag, such as pressure (which can be used to measure the content of the reaction liquid), pH, dissolved oxygen rate, CO2 concentration, turbidity, etc., it is determined whether it is necessary to replenish cell culture medium, water, acid or alkali solution, etc., into the reactor to complete the automatic replenishment of liquid during the culture process.

[0085] In some embodiments, any one of the control valves can be a one-way valve. In some embodiments, the control valve for controlling the liquid can be controlled by a peristaltic pump to control the amount or rate of liquid addition. In some embodiments, a sterilization unit, such as a 0.22-micron sterilization filter, is provided at the inlet for ventilation (e.g., oxygen) in the reaction bag.

[0086] It is understood that the disposable bioreactor in this embodiment of the invention can also be used to achieve continuous culture of microorganisms or cells. For example, taking virus culture as an example, when the number of viruses in the disposable bioreactor reaches a certain amount, a portion of the virus sample can be taken out from inside the reactor, and then new culture medium can be added to the reactor so that the remaining viruses can continue to grow inside the reactor, thereby achieving continuous virus culture.

[0087] In some embodiments, such as Figure 8-9 As shown, the sensor employs a biosafety-compliant quick-connect device to achieve sealing. This device includes a disposable sensor probe 81, a probe terminal 82, a power terminal 83, and an electrode 84. The probe terminal 82 contains positive and negative electrode terminals, which can be connected to the power terminal 83 via the electrode 84 to transmit current signals (specifically, sensing signals such as hydraulic pressure or dissolved oxygen levels) to the central control system (such as a central control module) for real-time monitoring of sensor parameters. In use, the probe terminal 82 is connected to the power terminal 83 to transmit electrical signals. In some embodiments, the support device has an interface for installing the power terminal 83. The interface of the support device is made of corrosion-resistant sealing material. The power terminal 83 can be inserted into the interface via a quick-connect or threaded connection, ensuring a seal between the power terminal 83 and the support device and preventing aerosol leakage. In some embodiments, for disposable bioreactors of 20L or less, placed in a biosafety cabinet / isolator, sampling and testing are performed directly within the biosafety cabinet / isolator by controlling a one-way sampling valve. In some embodiments, for reactors larger than 20L, the sampling pipeline is directly connected to the biosafety cabinet / isolator, and sampling and testing are performed by controlling the sampling check valve.

[0088] Alternatively, in some embodiments, the bioreactor may further include a central control module, which is communicatively connected to one or more control valves, one or more monitoring units or monitoring modules; the central control module includes a risk monitoring unit to enable real-time and accurate monitoring and maintenance of reaction safety.

[0089] Specifically, the risk monitoring unit is configured to execute the following process: Step 11: Real-time acquisition of monitoring values ​​from the first leakage monitoring module (such as the electrical signal from the leakage sensor); Step 12: The risk monitoring unit determines the leakage risk based on the leakage value; wherein, in order to accurately monitor the leakage situation and reduce the risk of misjudgment, the leakage risk is divided into level 0, level 1, and level 2 according to the magnitude of the leakage value; Next, safety warnings or safety disinfection measures are taken for the bioreactor according to the specific leakage risk level; Step 13: When the leakage risk is 1, the risk monitoring unit will determine whether the hydraulic pressure is abnormal; if so, it will send corresponding disinfection signals to the recovery valve and the disinfection valve; if not, it will continue to monitor the leakage risk and hydraulic pressure. If the hydraulic pressure monitoring value increases or the hydraulic pressure is abnormal, it will send corresponding disinfection signals to the recovery valve and the disinfection valve; Step 14: When the leakage risk is 2, it will directly send corresponding start signals to the recovery valve and the disinfection valve. In some embodiments, when a leakage risk is detected, a warning signal can also be sent to the staff, and the staff can manually decide whether to continue the experiment.

[0090] This invention provides a highly sensitive and relatively low-cost monitoring method (i.e., a dual-mechanism risk assessment method) by selecting two limited parameters for monitoring: external leakage and internal hydraulic pressure. Currently, the cultivation of highly infectious and pathogenic bacteria or viruses often requires high-level biosafety laboratories (e.g., P3 laboratories). This invention integrates key status monitoring, dual-mechanism risk assessment, and safety protection functions within a single small bioreactor. This allows the small bioreactor to maintain internal environmental stability during the cultivation of highly hazardous microorganisms through key parameter monitoring and an internal circulating condensate reflux design, while also enabling autonomous risk monitoring and disinfection based on low-cost dual-mechanism monitoring. Therefore, this small bioreactor can be directly applied to cell or microbial cultivation within a biosafety cabinet (BSC), thereby reducing the human and material costs associated with the cultivation of highly hazardous microorganisms.

[0091] Regarding the bioreactor in any embodiment of the present invention, the present invention also provides a monitoring method for using a bioreactor to achieve microbial culture. Preferably, in some embodiments, such as Figure 10As shown, the method includes the following steps: S102 Introducing a preset amount of gas or liquid into the reaction bag through a first inlet or a second inlet to culture microorganisms or cells; S104 Real-time monitoring of the hydraulic pressure of the first pressure monitoring unit and the leakage value of the first leakage monitoring module; S106 Determining the risk status of the bioreactor based on the hydraulic pressure and the leakage value, wherein the risk status includes: safe status, and / or pending status, and / or leakage status;

[0092] S106 includes: S61 determining the pressure drop risk of the reaction bag based on the hydraulic pressure change parameters (e.g., pressure drop value, or pressure drop rate); wherein, when the change parameter belongs to a preset first pressure drop threshold, the pressure drop risk is level one; when the change parameter belongs to a preset second pressure drop threshold, the pressure drop risk is level two; otherwise, the pressure drop risk is level zero; S62 determining the leakage risk of the reaction bag based on the leakage value (e.g., in some embodiments, the leakage value is the electrical signal value of a leakage sensor), wherein when the leakage... When the liquid level is within a preset first leakage threshold, the leakage risk is level one; when the leakage level is within a preset second leakage threshold, the leakage risk is level two; otherwise, the leakage risk is level zero. S63 determines the risk status based on the pressure drop risk in S61 and / or the leakage risk in S62; wherein, when the pressure drop risk is level two and / or the leakage risk is level two, the risk status is a leakage status; when both the pressure drop risk and the leakage risk are level zero, the risk status is a safe status; otherwise, the risk status is a pending status.

[0093] In some embodiments, a condensate guiding zone is further formed in the top region of the first space to guide condensate to condense in the condensate guiding zone and flow back to the second space through the guiding path provided by the condensate guiding zone. The method further includes the steps of: S108 issuing corresponding warning signals to the user when the risk state is detected to be a pending state or a leakage state; S110 performing secondary monitoring of the pressure reduction risk and / or leakage risk of the bioreactor in response to the feedback signal issued by the user, or receiving or correcting the warning signal.

[0094] In some embodiments, step S110 includes: when the pressure drop risk is level one and the leakage risk is level zero.

[0095] The condensate guide zone is heated in response to a first feedback signal from the user; when the pressure drop risk is detected to be reduced to zero during or after heating, the risk state is corrected to a safe state in response to a first correction signal from the user; otherwise, the risk state is corrected to a leakage state in response to a second correction signal from the user.

[0096] Specifically, in some embodiments, when a risk status is detected as pending, a warning signal needs to be sent to the user.

[0097] This serves as a reminder for users to promptly check the operating status of the bioreactor (e.g., whether there is a tendency or slight leakage, or whether there is excessive condensation). For example, in some embodiments, when a risk status is detected as pending, the user will receive a corresponding warning signal. At this time, the user can choose to heat the condensate guiding zone (preferably through a heating unit installed on the stirring shaft) to limit or reduce condensation formation, thereby determining whether there is excessive condensation in the current bioreactor. If so, the user can correct the risk status to a safe status when the pressure risk returns to a normal state (e.g., level zero).

[0098] It is understandable that, especially for bioreactors with a volume of only a few liters (e.g., 1L), the total amount of liquid inside the reactor is relatively limited, making hydraulic pressure relatively susceptible to multiple factors (such as stirring speed, stirring intensity, leakage, condensation, etc.). In this case, setting the sensor's monitoring difference (equivalent to the second pressure drop threshold) too high will make it difficult to detect problems immediately, while setting it too low may lead to frequent false alarms interfering with the normal operation of the experiment. In this embodiment of the invention, the formation or backflow of condensate is controlled or influenced by an internally circulating condensate guiding zone, so that even when the sensor is at a low monitoring difference (i.e., high sensitivity), excessive false alarms will not interfere with the normal operation of the experiment. In other words, this invention can improve the accuracy of leakage detection to a certain extent.

[0099] For example, in some embodiments, the monitoring difference can be set to around 5% or even lower. Specifically, when the difference in hydraulic pressure reaches or exceeds 5%, the bioreactor is considered to have a high risk of leakage.

[0100] Specifically, in some embodiments, when it is detected that the hydraulic pressure value recovers to the expected level (the expected level can be determined based on the actual amount of liquid added) within a preset second time period, and the risk of leakage remains unchanged, the risk state can be converted into a safe state. Alternatively, in some embodiments, when it is detected that the hydraulic pressure parameters do not change significantly within a preset third time period (e.g., still within the range of the first pressure drop threshold), a corresponding warning signal is issued to the user to alert them to the current risk of excessive condensate.

[0101] It is understood that each threshold in the embodiments of the present invention can be adaptively set by the user in combination with the actual situation (such as the volume of the reactor, the amount of liquid added, etc.).

[0102] In this embodiment of the invention, by real-time monitoring of a limited number of parameters, a low-cost and high-precision monitoring mode is provided for the working status (specifically, the risk status) of the bioreactor. On the one hand, this can reduce the manual monitoring pressure on staff to a certain extent, and on the other hand, it can avoid interference caused by over-monitoring (causing the experiment to be stopped due to error signals, resulting in a waste of human and material resources).

[0103] In some embodiments, prior to S102, the step further includes: S100 pre-testing the bioreactor;

[0104] S100 includes: introducing gas into the reaction bag through the first inlet while keeping the other inlets and outlets of the reaction bag closed; ending the gas supply and keeping the corresponding first inlet closed; collecting the first gas pressure change value of the reaction bag within a first time period through the first pressure monitoring unit, and determining whether the bioreactor passes the pre-test (i.e., whether the sealing performance is good) based on the first gas pressure change value; wherein, when the first gas pressure change value is within a preset safety threshold (specifically, when the gas pressure change is small or the change value is 0), the bioreactor passes the pre-test.

[0105] like Figure 6 , 7 As shown, to further improve operational safety during the cultivation of highly pathogenic microorganisms, the present invention also provides a support device for supporting the aforementioned bioreactor. For example... Figure 6As shown, it includes: a temperature-controlled support body 7 for supporting the bioreactor; the bioreactor includes: a reaction bag; the support body 7 includes: a sidewall structure formed by a first sidewall 73 and a second sidewall 74, a base 72 formed below the sidewall structure, and a top cover for sealing the top of the sidewall structure, wherein the sidewall structure and the base 72 provide support for the reaction bag; the top cover is provided with at least one third inlet for leading out at least one pipe connected to the first inlet and the second inlet in the reaction bag, the first and second inlets being used to add gas and liquid to the reaction bag, respectively; and for sealing the reaction bag. The heating module for heating includes: a heating cavity disposed on a first side wall and / or a second side wall, the heating cavity being used to contain a heat-conducting medium, and the heating cavity also having a medium inlet and a medium outlet for the heat-conducting medium to enter and exit; a temperature monitoring unit (e.g., a temperature sensor) for monitoring the temperature of the heat-conducting medium, and a heating unit communicatively connected to the temperature monitoring unit for heating the heat-conducting medium; and a first leakage monitoring module (e.g., a leakage monitoring sensor 71) disposed in the central region of the bottom, the first leakage monitoring module being used to monitor whether leakage occurs in the reaction bag and / or the heating cavity.

[0106] In some embodiments, for small bioreactor bags, water is used as the heat transfer medium to uniformly heat the circumferential walls of the small bioreactor bag. For example, in some embodiments, when the internal temperature of the reactor is detected by a temperature sensor to be lower than a preset culture temperature, an appropriate high-temperature heat transfer medium can be introduced into the heating chamber to heat the reactor. Alternatively, the heat transfer medium can be directly heated by a heating unit (e.g., an electric heating tube inserted into water, etc.).

[0107] In some embodiments, the bioreactor includes: a working status monitoring module for monitoring the working status parameters of the bioreactor, the working status parameters including: liquid hydraulic pressure, foam generation, dissolved oxygen rate, CO2 concentration, pH value, and turbidity; correspondingly, the support device further includes: a central control module, the central control module being used to communicate with the working status monitoring module, the first leakage monitoring module, and the heating module, and to control the operating status of the bioreactor and the support device according to the monitoring results communicated by the working status monitoring module, the first leakage monitoring module, and the heating module.

[0108] In some embodiments, the bioreactor further includes: a safety disinfection module communicatively connected to the central control module; the safety disinfection module includes: a recovery unit connected to the reaction bag and a disinfection unit, the recovery unit includes: a recovery bag and a recovery pipe for connecting the recovery bag and the reaction bag, the recovery pipe being provided with a third control valve, the disinfection unit includes: a chamber for storing disinfectant (e.g., hydrogen peroxide disinfectant), a first disinfection pipe for introducing the disinfectant into the interior of the reaction bag, and a second disinfection pipe for introducing the disinfectant between the support body and the reaction bag, both the first and second disinfection pipes being provided with a fourth control valve, wherein the chamber may be located on the second side wall.

[0109] Furthermore, in some embodiments, the safety disinfection module can also disinfect various pipelines in the reaction bag or support device. Especially when culturing highly pathogenic viruses, bacteria, and other microorganisms, the support device in this embodiment of the invention can monitor the working status of the disposable bioreactor in real time, and promptly activate the safety disinfection module to complete the safety disinfection upon detecting leakage signals.

[0110] In some embodiments, a viewing window is also provided on the second sidewall for observing the reaction conditions inside the reaction bag.

[0111] In some embodiments, a second leakage monitoring module is provided on the second sidewall, and when the reaction bag is installed inside the support body, the monitoring end of the second leakage monitoring module is in contact with the wall surface of the reaction bag. Therefore, when the reaction bag is slightly damaged (such as condensate overflowing from the mounting hole of the stirring device, or a small amount of leakage in the upper area of ​​the reaction bag), the leakage monitoring sensor provided on the second sidewall can supplement the monitoring of the stability of the reaction bag, further reducing the experimental risk. In some embodiments, the leakage monitoring module includes one or more of the following sensors: point leakage sensor, bracket probe leakage sensor, non-positioning leakage detector, and positioning leakage detector.

[0112] In some embodiments, the top cover and sidewall structure are connected by a threaded connection. For example, in some embodiments, the top cover and sidewall structure are connected by a quick-connect structure, and the areas where the top cover and sidewall structure contact each other are made of a corrosion-resistant material.

[0113] In some embodiments, the side wall of the support device is provided with two leak-proof disinfection ports (connected to the disinfection unit).

[0114] In the event of a leak, the cavity is circulated and disinfected through two leak-proof ports. These ports can be flexibly positioned on the side wall or base of the support device. For example, in some embodiments, one leak-proof port is an inlet and the other an outlet. Specifically, during disinfection, a certain amount of disinfectant (e.g., liquid disinfectant / disinfectant gas can be introduced or sprayed through the port) can be output into the support device through the inlet for a period of time, and then discharged through the outlet. Alternatively, in other embodiments, the input disinfectant can be recycled. Specifically, disinfectant is introduced at the inlet, and a peristaltic pump is used to recover the disinfectant at the outlet, and the recovered disinfectant is then reintroduced into the support device (e.g., added back into the support device through the inlet). In some embodiments, after disinfection is complete, the interior of the support device can be cleaned through the inlet and port (e.g., by washing with water) to extend the lifespan of the support device. It is understood that personnel can choose different disinfection methods according to different application needs (e.g., disinfection of new products, or disinfection after a reaction).

[0115] This invention also provides a monitoring method for a disposable biosafety bioreactor support device, comprising the following steps: S200 providing any support device provided by this invention, and a reaction bag; wherein, the reaction bag includes: a first pressure monitoring unit for measuring hydraulic pressure, the reaction bag being installed inside the support body of the support device, the wall of the reaction bag abutting or adjacent to at least one side wall of the support device to improve the heating efficiency of the heating module heating the reaction bag, and the bottom of the reaction bag contacting a first leakage monitoring module to facilitate leakage monitoring of the bottom of the reaction bag by the first leakage monitoring module; S202 introducing a preset amount of gas or liquid into the reaction bag through a first inlet or a second inlet on the reaction bag for the cultivation of microorganisms or cells; S204 real-time monitoring of the hydraulic pressure of the first pressure monitoring unit and the leakage value of the first leakage monitoring module through the central control module of the support device; S206 determining the risk status of the support device and the bioreactor based on the hydraulic pressure and the leakage value. In some embodiments, the risk state includes: a safe state, and / or a leakage state, and / or a pending state; wherein, S206 includes: determining the pressure reduction risk of the reaction bag based on the change parameters of the hydraulic pressure; wherein, when the change parameters belong to a preset first pressure reduction threshold, the pressure reduction risk is level one; when the change parameters belong to a preset second pressure reduction threshold, the pressure reduction risk is level two; otherwise, the pressure reduction risk is level zero; determining the leakage risk of the reaction bag based on the leakage value, wherein, when the leakage value belongs to a preset first leakage threshold, the leakage risk is level one. The leakage risk is classified as follows: when the leakage value falls within a preset second leakage threshold, the leakage risk is level two; otherwise, the leakage risk is level zero. The risk status is determined based on the pressure reduction risk and / or the leakage risk. Specifically, when the pressure reduction risk is level two and / or the leakage risk is level two, the risk status is a leakage status. When both the pressure reduction risk and the leakage risk are level zero, the risk status is a safe status. When the pressure reduction risk is level one and the leakage risk is level zero, or when the pressure reduction risk is level zero and the leakage risk is level one, the risk status is a pending status. It is understood that the monitoring method in this embodiment may include the same steps as in the above embodiments. For example, the method may also include issuing a warning to the user based on the risk status result, or disinfecting the support device and reactor, etc., which will not be elaborated here.

[0116] In some embodiments, a pressure monitoring module for monitoring the internal pressure of the support body is further provided on the second sidewall of the support device. Accordingly, before S202, the method further includes the step of S208 pre-testing the bioreactor and the support device; wherein, S208 includes the steps of: introducing gas into the reaction bag through the first inlet and monitoring the second pressure change value inside the support body through the pressure monitoring module; keeping the remaining inlets and outlets of the reaction bag and the support device closed at the same time; ending the gas supply and keeping the corresponding first inlet closed; collecting the first pressure change value of the reaction bag within a preset first time period through the first pressure monitoring unit and monitoring the third pressure change value of the support body within the first time period through the pressure monitoring module;

[0117] The bioreactor passes the pre-test based on the first pressure change value, and the support device passes the pre-test based on the second and third pressure change values. Specifically, the bioreactor passes the pre-test when the first pressure change value falls within a preset safety threshold (i.e., the pressure change is 0 or very small). The support device passes the pre-test when the second pressure change value matches the amount of gas introduced (understandably, the more gas introduced, the greater the pressure change) and the third pressure change value falls within a preset safety threshold. For example, in some embodiments, if the pressure in the reaction bag does not change significantly after aeration, the reaction bag is considered to have good sealing performance and meets the requirements. Similarly, in some embodiments, if the pressure inside the support body increases significantly during aeration and remains stable during the first period of testing, the support device is considered to have good sealing performance and meets the requirements.

[0118] Furthermore, in some embodiments, the support device can also achieve high-temperature in-situ sterilization. Furthermore, in some embodiments, the base is also provided with casters to facilitate the transportation of the support device. Furthermore, in some embodiments, the top of the support device is also provided with mounting holes for mounting the stirring device 2.

[0119] In some embodiments, different structures of stirring devices can be selected depending on the culture medium. For example, when the culture medium is cells, the stirring impeller of the stirring device can be blade-shaped. As another example, when the culture medium is bacteria, the stirring impeller of the stirrer can be rod-shaped.

[0120] In some embodiments, the various monitoring units equipped in the reaction bag can all be selected from disposable electrode detection sensors.

[0121] The sensors and reaction bags are all sealed to prevent leakage. The entire bioreactor requires only simple installation and is for single use. The disposable bioreactor provided by this invention can be used for the cultivation of various microorganisms and cells; for example, it can be applied to vaccine production.

[0122] It should be noted that the present invention preferably provides a small-scale integrated bioreactor and supporting device for the cultivation of miniaturized, highly pathogenic microorganisms. Furthermore, this small-scale integrated bioreactor can be scaled up according to the user's actual needs (e.g., from small-scale to pilot-scale), for example, to a bioreactor with a chamber capacity of approximately 100L-200L. The present invention can be widely applied to various types of microorganisms or cells.

[0123] The present invention also provides a semi-automatic control method applicable to the above-mentioned bioreactor and support device, so as to reduce the user's intervention (such as ventilation or disinfection) to a certain extent during the culture of highly pathogenic viruses. Figure 11 An exemplary semi-automatic control method of the present invention is shown, comprising: S301 providing a bioreactor system for microbial growth, the bioreactor system comprising: a bioreactor, and a support device for supporting the bioreactor, wherein the bioreactor comprises: a reaction bag, the reaction bag having a first space and a second space for containing gas and liquid respectively, the reaction bag also having a first inlet and a second inlet for introducing gas and liquid respectively, and the reaction bag also having a working status monitoring module for monitoring the working status parameters of the bioreactor; S302 responding to a user or the working... The status monitoring module sends corresponding control signals to introduce gas and / or liquid into the reaction bag through the first inlet and / or the second inlet for biological culture (microbial culture or cell culture); S303 monitors the pressure of the liquid inside the second space to collect hydraulic pressure, and monitors leakage at the bottom of the reaction bag to collect leakage value; S304 determines the risk status of the bioreactor system based on the hydraulic pressure and the leakage value, the risk status including: safe status, and / or pending status, and / or leakage status; wherein, S304 includes: determining the pressure drop risk of the reaction bag based on the change parameters of the hydraulic pressure; wherein,

[0124] The pressure drop risk is classified into three levels based on the magnitude of the changing parameters: Level 0, Level 1, and Level 2; the leakage risk of the reaction bag is determined based on the leakage value; wherein, the leakage risk is classified into three levels based on the magnitude of the leakage value:

[0125] The risk status is categorized into three levels: Level 0, Level 1, and Level 2. The risk status is determined based on the pressure reduction risk and leakage risk. Specifically, when the pressure reduction risk is Level 2 and / or the leakage risk is Level 2, the risk status is a leakage status. When both the pressure reduction risk and the leakage risk are Level 0, the risk status is a safe status. When the pressure reduction risk is Level 1 and the leakage risk is Level 0, or when the pressure reduction risk is Level 0 and the leakage risk is Level 1, the risk status is a pending status.

[0126] In some embodiments, the method further includes: S305 When a risk state is detected as an undetermined state or a leakage state,

[0127] The corresponding warning signals are sent to the user respectively; S306 responds to the feedback signals sent by the user to perform secondary monitoring of the pressure reduction risk and / or leakage risk of the bioreactor system, so as to receive or correct the warning signals.

[0128] Preferably, in some embodiments, the feedback signal can be manually issued by the staff. Alternatively, in other embodiments, if no feedback signal issued by the staff is monitored or received within a set time (e.g., 5 seconds or 15 seconds), a computer (which can communicate with the bioreactor or support device) can issue a feedback signal pre-set by the staff according to the current risk status.

[0129] In some embodiments, a condensate guiding zone is further formed in the top region of the first space. The condensate guiding zone is used to guide condensate to condense within the guiding zone and to flow back to the second space through the guiding path provided by the condensate guiding zone. Accordingly, the feedback signal includes: a first feedback signal for heating the condensate guiding zone and a second feedback signal for receiving the warning signal. S306 includes the steps of: when the pressure drop risk is level one and the leakage risk is level zero, the heating unit disposed in the condensate guiding zone heats the condensate guiding zone in response to the first feedback signal; when it is detected that the pressure drop risk drops to level zero during or after heating, the risk state is corrected to a safe state in response to the first correction signal issued by the user; otherwise, the risk state is corrected to a leakage state in response to the second correction signal issued by the user.

[0130] Preferably, in this embodiment of the invention, the correction of the risk status needs to be manually performed by staff to avoid misjudgment of the bioreactor. Of course, in other embodiments, the risk status can also be corrected and assessed automatically. In some embodiments, S306 further includes the step of: when the pressure drop risk is detected to be level zero or the leakage risk is level one, continuing to monitor the leakage value, and when the leakage value is detected to remain constant within a preset fourth time period, then in response to the first correction signal, correcting the risk status to a safe status; otherwise, in response to the second correction signal, correcting the risk status to a leakage status. For example, in some embodiments, when the leakage value does not change significantly within a certain period of time (e.g., 5s or 10s), the user can rule out the possibility of leakage in the bioreactor.

[0131] In some embodiments, the bioreactor system further includes: a stirring device, the stirring device including a stirring shaft and a stirring paddle disposed at the end of the stirring shaft; a mounting hole for mounting the stirring device is provided in the central region of the condensate guiding zone, and the stirring shaft is fixedly mounted on the reaction bag through the mounting hole; a heating unit is provided on the side of the stirring shaft located at or near the condensate guiding zone to avoid or limit the formation of condensate.

[0132] In some embodiments, the bioreactor system further includes a safety disinfection module for disinfecting the bioreactor and the support device. The safety disinfection module includes a recovery unit connected to the reaction bag via a recovery pipe, and a chamber for storing disinfectant. The chamber is connected to the reaction bag and the support device via disinfection pipes. The method further includes the steps of: S307 When the risk state is detected as a leakage state, a corresponding disinfection signal is sent to the safety disinfection module; S308 In response to the disinfection signal, the safety disinfection module recovers the liquid inside the reaction bag to the recovery unit, and closes the recovery pipe after the recovery is completed; S309 After the recovery pipe is closed, the safety disinfection module, in response to the disinfection signal, introduces the disinfectant into the reaction bag and the support device.

[0133] In some embodiments, before step 301, the method further includes the step S300: performing a pre-test on the stability of the bioreactor system; wherein, S300 includes: introducing gas into the reaction bag through the first inlet and simultaneously collecting a second pressure change value inside the support device; simultaneously keeping the remaining inlets and outlets of the reaction bag and the support device closed; ending the gas supply and keeping the corresponding first inlet closed; collecting a first pressure change value of the reaction bag within a preset first time period and a third pressure change value of the support device within the first time period; determining whether the stability of the bioreactor system is qualified based on the first, second, and third pressure change values; wherein,

[0134] When the first pressure change value is within the corresponding preset safety threshold, and the second and third pressure change values ​​are consistent with the amount of gas introduced, the bioreactor system passes the pre-test.

[0135] For small-scale cultivation of highly pathogenic microorganisms (such as influenza viruses), this invention selects limited monitoring factors (i.e., internal hydraulic pressure and external leakage value) for core monitoring of the bioreactor's operating status during the cultivation process, providing a low-cost, semi-automated control method that reduces manual intervention. Furthermore, this method can be combined with a condensate guide zone to further reduce interference from other factors on risk monitoring, thereby reducing misjudgments and thus reducing R&D costs to some extent (specifically, avoiding the termination of experiments due to misjudgments and the excessive consumption of human and material resources).

[0136] Based on the above semi-automatic control methods, such as Figure 12As shown, the present invention also provides a semi-automated system, including: a bioreactor system 10, comprising a bioreactor and a support device for supporting the bioreactor, wherein the bioreactor includes: a reaction bag, the reaction bag having a first space and a second space for containing gas and liquid respectively, and the reaction bag also having a first inlet and a second inlet for introducing gas and liquid respectively; a working status monitoring module for monitoring the working status parameters of the bioreactor; a sample addition module 20, configured to introduce gas and / or liquid into the reaction bag through the first inlet and / or the second inlet for biological culture in response to a corresponding control signal issued by the user or the working status monitoring module; a leakage monitoring module 30, configured to monitor the pressure of the liquid inside the second space to collect hydraulic pressure, and monitor the leakage at the bottom of the reaction bag to collect leakage value; and a risk analysis module 40, configured to determine the biological reaction based on hydraulic pressure and leakage value. The system's risk status includes: a safe state, and / or a pending state, and / or a leakage state. The risk analysis module includes: a hydraulic analysis unit 40-1, configured to determine the pressure drop risk of the reaction bag based on changes in hydraulic parameters; where the pressure drop risk is categorized into level 0, level 1, and level 2 based on the magnitude of the changing parameters; a leakage analysis unit 40-2, configured to determine the leakage risk of the reaction bag based on the leakage value; where the leakage risk is categorized into level 0, level 1, and level 2 based on the leakage value; and a risk analysis unit 40-3, configured to determine the risk status based on the pressure drop risk and leakage risk; where the risk status is a leakage state when the pressure drop risk is level 2 and / or the leakage risk is level 2; the risk status is a safe state when both the pressure drop risk and leakage risk are level 0; and the risk status is a pending state when the pressure drop risk is level 1 and the leakage risk is level 0, or when the pressure drop risk is level 0 and the leakage risk is level 1.

[0137] In some embodiments, the sample addition module can be customized by the user to add culture nodes according to the culture status, such as turbidity, so as to determine when to stop the culture, when to automatically add liquid, automatically adjust pH, and introduce different gases or oxygen, etc.

[0138] In some embodiments, the system further includes: an early warning sending module 50, configured to send corresponding early warning signals to the user when the detected risk status is either pending or leakage; and an early warning correction module 60, configured to perform secondary monitoring of the pressure reduction risk and / or leakage risk of the bioreactor system in response to feedback signals from the user, so as to receive or correct the early warning signals.

[0139] In some embodiments, a condensate guiding zone is further formed in the top region of the first space. The condensate guiding zone guides condensate to condense within the guiding zone and flows back to the second space through the guiding path provided by the condensate guiding zone. Accordingly, the feedback signal includes: a first feedback signal for heating the condensate guiding zone and a second feedback signal for receiving the warning signal. The warning correction module 60 includes: a secondary monitoring unit 60-1, configured to heat the condensate guiding zone in response to the first feedback signal when the pressure drop risk is level one and the leakage risk is level zero; and a correction unit 60-2, which, when it is detected that the pressure drop risk drops to level zero during or after heating, corrects the risk state to a safe state in response to the first correction signal issued by the user; otherwise, it corrects the risk state to a leakage state in response to the second correction signal issued by the user.

[0140] To simplify vaccine production processes and reduce initial vaccine development costs, this invention provides a method for producing single-use highly pathogenic microorganism or inactivated vaccines suitable for small-scale vaccine trials. Figure 13 As shown, the method includes the following steps: S001, providing a customizable and assembleable biological culture module, and a management module for controlling the operating status of the biological culture module; wherein, the biological culture module includes: a bioreactor for culturing microorganisms, an intermediate device for processing a liquid containing the microorganisms, and a purification device for purifying the liquid; the intermediate device is connected to any one or both of the bioreactor and the purification device via at least one detachable pipe for collecting the liquid from the bioreactor and / or the purification device and processing the liquid, wherein the detachable pipe has a pipe... At least one leakage sensing line is arranged along the length direction; S002 When the biological culture module is turned on, a first set consisting of at least one reading from at least one of the leakage sensing lines is collected in real time; S003 The first risk state of the biological culture module is determined based on the first set, the first risk state including: leakage object, and / or leakage risk level; S004 The management module generates corresponding control signals or instructions according to the risk state (such as the first risk state, the second risk state) and sends them to the biological culture module; S005 The biological culture module responds to the control signal or instruction by selecting to continue production, suspend production, or stop production. Of course, the production method in this embodiment can also be applied to processes such as the cultivation and inactivation of highly pathogenic microorganisms. Highly pathogenic microorganisms refer to viruses or bacteria that may harm the health of organisms (such as humans) or have adverse effects on the environment.

[0141] To address the needs of small-scale trials in the early stages of vaccine development, this invention adopts a design approach diametrically opposed to the traditional fixed production model. Specifically, it provides a set of flexibly detachable and assembleable biological culture modules, facilitating flexible combination and replacement of the entire production line during the early process design and real-time vaccine production. Furthermore, to ensure the stability and safety of the culture modules assembled on-site, in some embodiments, when at least one leakage is detected, step S004 includes the following step:

[0142] The management module determines whether the number of leaking objects exceeds a preset first threshold (e.g., 5 or 8).

[0143] Alternatively, other quantities can be set according to actual production needs.

[0144] If so, the management module sends a first control signal to the biological culture module indicating a halt to production, wherein,

[0145] The first control signal includes: a stop signal indicating the cessation of production, and a disinfection signal indicating the initiation of disinfection;

[0146] If not, the management module sends a second control signal to the biological culture module indicating a production halt. The second control signal includes: a halting object that needs to be halted; wherein the halting object includes: a leaking object, and one or more of the detachable pipes and / or one or more devices connected to the leaking object.

[0147] For example, in some embodiments, when the number of devices at risk of leakage exceeds five, the biological culture module is in a high-risk state and can be directly shut down, and disinfection of the biological culture module can be initiated to prevent further spread of risk. When the number of devices at risk of leakage is five or less, the operation of only the devices at risk of leakage is suspended, and the user is reminded to replace the leaking device in time. After the user has replaced the device, the biological culture module can resume normal operation.

[0148] The technical solution and effects of this invention will be further explained below using the production process of inactivated bacteria as an example:

[0149] like Figure 14As shown, the bioculture module used for inactivated bacterial culture includes: a bioreactor A (specifically a bioreactor and supporting structure), a clarification and filtration device B1, an ultrafiltration device (for removing the inactivating agent and concentrating the liquid) B2, a first liquid exchange and concentration device B3, a purification device C, a second liquid exchange and concentration device B4, a mixing device B5 (for adding adjuvants and mixing), a dilution device B6, and a collection device D. Each device is sequentially connected via detachable pipes, and the bioreactor's inlet pipe and each detachable pipe are communicatively connected to a management module (such as a computer). Specifically, in some embodiments, when leakage is detected in the clarification and filtration device B1, the operation of the clarification and filtration device B1, the bioreactor, and the pipes connecting them can be simultaneously stopped to prevent the bioreactor from continuing to supply liquid to the clarification and filtration device. Alternatively, only the operation of the clarification and filtration device can be suspended and replaced to complete the dynamic maintenance of the production line. It is understood that the purification device and intermediate device in the embodiments of the present invention can be flexibly configured or adjusted according to actual application requirements. For example, in some other embodiments, the purification device includes one or more of the following: an ultrafiltration deactivation device, an ultrafiltration liquid exchange device, a clarification filtration device, and a supernatant concentration device; the intermediate device includes one or more of the following: a chromatography device and a mixing device.

[0150] Preferably, one or more of the above devices are biosafety devices, such as biosafety clarification filtration devices, biosafety ultrafiltration devices, etc.

[0151] In some embodiments, a diaphragm pump or peristaltic pump for conveying liquid is provided at the interface between the device and the detachable pipe. Preferably, a unidirectional diaphragm pump is used for liquid delivery to improve the safety and stability of the internal liquid during normal operation and replacement pauses of the custom module.

[0152] In some embodiments, the reading includes a current value, and S003 includes the step of generating a first leakage signal including the first risk status information when the current value of the detachable pipe is greater than a preset second threshold, and the first leakage signal is associated with one or two devices connected to the detachable pipe (e.g., the first leakage signal includes or marks leakage object information).

[0153] Accordingly, S004 includes the following steps:

[0154] 1) When the management module detects two first leakage signals associated with the device, it identifies the corresponding device and detachable pipeline as the first leakage object and generates a second control signal indicating a production halt. The second control signal includes information about the first leakage object (e.g., device name or number, and risk level). For example, when leakage signals are detected on both sides of the clarifier / filter B1, the operation of the clarifier / filter B1 is halted, and the pipelines on both sides are closed (i.e., liquid inlet and outlet are stopped).

[0155] In some embodiments, S004 further includes the step: 3) when a first leakage signal is detected associated with the device,

[0156] The corresponding device is identified as the second leaking object, and the detachable pipe is identified as the first leaking object, and a first alarm signal is sent to the user; 4) The management module monitors whether the user issues a corresponding instruction within the first alarm time after the first alarm signal is issued; wherein, the instruction includes: a first instruction indicating cancellation of the alarm, and / or, a second instruction indicating receipt of the alarm, and / or, a third instruction indicating correction of the alarm, and the third instruction includes: the corrected leaking object; if yes, the received instruction is sent to the biological culture module; if no, a second control signal indicating suspension of production is sent directly to the biological culture module. For example, in some embodiments, when only the right pipe of the clarification filter device B1 (i.e., the side connected to the ultrafiltration device B2) is detected to have a leaking signal, manual intervention can be alerted. At this time, the user can determine whether it is necessary to suspend or continue production. For example, users can choose to continue production and cancel the alarm based on actual production conditions (such as the production progress of inactivated bacteria, the level of leakage risk, etc.) (i.e., inputting a first instruction to the management module); or receive the suggested handling method automatically generated by the current management module (i.e., inputting a second instruction to the management module); or, users can add other components (such as bioreactors) as leakage targets based on actual production conditions and replace the leakage targets (i.e., inputting a third instruction to the management module). In some embodiments, the leakage risk level of the first leakage target is higher than that of the second leakage target.

[0157] Typically, the production of biological products such as inactivated bacteria takes several days. Unexpected situations during production (such as leaks or gas leaks) can pose a significant danger to workers. Conversely, directly terminating production due to unforeseen circumstances wastes the initial investment of manpower and equipment, increasing the initial R&D costs of the biological products. To mitigate the conflict between risk avoidance and cost control, this invention proposes a "dynamic maintenance" production mode. Specifically, this embodiment first provides a freely disassembled and reassembled biological culture module. Furthermore, it employs multi-segment leak signal monitoring to monitor the risk status of the biological culture module in real time. While maintaining the operation of the biological culture module, the production line is dynamically managed and maintained based on the real-time monitoring results (e.g., workers can replace specific devices based on multiple leak signals).

[0158] Furthermore, in order to reduce the maintenance difficulty and cost of "dynamic maintenance", the present invention preferably adopts a monitoring mode of internal and external dual leakage signals to optimize the maintenance scheme of dynamic maintenance. For example, in some embodiments, the detachable pipe includes: an internal pipe for conveying fluid, and an external pipe for protecting the internal pipe; wherein, at least one first leakage sensing line is provided on the outer side of the internal pipe along its length, and at least one second leakage sensing line is provided on the outer side of the external pipe along its length; and the reading includes: the current value of the leakage sensing line. Accordingly, step S003 includes: (i) determining that the internal pipe is leaking when the current value of the internal pipe is detected to be greater than a preset second threshold; (ii) generating a second leakage signal (i.e., an internal pipe leakage signal) associated with the detachable pipe, the second leakage signal including: pipe information, and / or leakage risk level; and (iii) determining that the external pipe is leaking when the current value of the external pipe is detected to be greater than a preset second threshold; and (iv) generating a third leakage signal (i.e., an internal pipe leakage signal) associated with the pipe.

[0159] The third leakage signal includes: pipeline information, and / or leakage risk level; accordingly, S004 includes the step:

[0160] When the management module detects that the detachable pipe is associated with the second leakage signal and the third leakage signal, it identifies the detachable pipe as a leakage object and generates a first control sub-signal indicating a production halt based on the leakage object, which is then sent to the biological culture module.

[0161] When the management module detects only the second leakage signal associated with the detachable pipe, it sends a notification to the user.

[0162] Issue the second alarm signal;

[0163] When the management module detects that the detachable pipe is only associated with the third leakage signal, it determines the leakage risk level of the pipe (e.g., external pipe) and the device based on the reading, and generates a second control sub-signal indicating a suspension of production based on the leakage risk level and sends it to the biological culture module.

[0164] For example, in some embodiments, when the management module detects leakage signals in both the inner and outer pipes, it recommends that the user replace the pipes directly. When the management module only detects leakage in the outer pipe, if the risk of leakage in the outer pipe is high (e.g., a large leakage value), it recommends replacement; otherwise, it recommends that the production line continue operating and will also send an alarm to the user. When the management module only detects leakage in the inner pipe, it recommends that the production line operate normally and will send an alarm to the user.

[0165] In some embodiments, the pipes and devices are connected via a quick-release structure, such as... Figure 15b , 15c As shown.

[0166] For example, in some embodiments, such as Figure 15b As shown, the end of the detachable pipe 91 is provided with a first pair of interfaces.

[0167] A second pair of interfaces 92 is provided on the side wall of the device (such as the side wall of a bioreactor bag) to mate with it. The first pair of interfaces includes a first external interface 91-1 located at the end of an external pipe, and a first internal interface 91-2 located at the end of an internal pipe. Correspondingly, the second pair of interfaces 92 includes a second external interface 92-1 that is sealed to the side wall, with the inner wall of the second external interface 92-1 protruding and extending to form the second internal interface 92-2. Further, a first protrusion is provided on one side of the first external interface 91-1, and a corresponding second protrusion is provided on the second external interface. When the first pair of interfaces of the detachable pipe mates with the second pair of interfaces under external force (i.e., when the first pair of interfaces is inserted into the second pair of interfaces), the corresponding wall surfaces of the first pair of interfaces (such as the first external interface) and the second pair of interfaces (such as the second external interface) will fit tightly together. At this time, the detachable pipe 91 is fixedly connected to the second pair of interfaces under the squeezing action inside the second pair of interfaces 92 and the limiting action of the second protrusion. It is understandable that the first inner interface and the second inner interface are respectively provided with corresponding third protrusions and fourth protrusions to achieve mutual cooperation between the inner interfaces.

[0168] Preferably, in some embodiments, the detachable pipe can be made of an elastic material, such as plastic, rubber, etc. Specifically, when the elastic detachable pipe (specifically the first pair of interfaces) mates with the second pair of interfaces, since the outer diameter of the first pair of interfaces is slightly larger than the inner diameter of the second pair of interfaces, the first pair of interfaces will deform to a certain extent under the pressure of the second pair of interfaces, thereby achieving a tight fit with the second pair of interfaces, i.e., achieving good sealing at the interface. Preferably, as... Figure 15c As shown, the second pair of interfaces is sealed to the side wall of the device by means of threaded connection. Specifically, the outer side of the second pair of interfaces is provided with a threaded structure 92-3.

[0169] In some embodiments, the clarification and filtration device includes: a first chamber for filtration, and a second chamber disposed on the outer layer of the first chamber. The first chamber contains a filter medium for filtering the liquid, and one end of the first chamber is connected to a bioreactor or a purification device, and the other end of the first chamber is connected to a purification device, an intermediate device, or a collection device. Leakage sensing units are respectively disposed on the outer walls of the first chamber and the second chamber, and the leakage sensing units are leakage sensors or leakage sensing lines. Specifically, in some embodiments, the clarification and filtration device may be made of a double-layer bio-bag.

[0170] In some embodiments, the alarm signal includes one or more of the following forms: voice alarm, and / or SMS alarm.

[0171] And / or traffic light alarms.

[0172] In some embodiments, the step of determining the leakage risk level of the pipeline leakage and the device leakage based on the reading includes:

[0173] The distribution characteristics of the leakage locations of the pipe (such as an internal pipe) are determined based on the readings, wherein the distribution characteristics include: a continuous distribution connected to both sides of the pipe, and / or a regional distribution disconnected from both sides of the pipe;

[0174] When the distribution characteristic is detected to be continuous (e.g., when a leakage signal is continuously detected along the length of the pipe),

[0175] The leakage risk level at the interface of the device on both sides of the pipeline is determined to be Level 1 (equivalent to high risk), and the leakage risk level of the pipeline is determined to be Level 2 (equivalent to medium-low risk).

[0176] When the distribution characteristic is detected as a regional distribution (e.g., when a small amount of leakage signal is detected only in the middle region of the pipeline), the leakage risk level of the pipeline is determined to be Level 1, and the leakage risk level of the devices on both sides of the pipeline is determined to be Level 2; accordingly, S004 further includes the step:

[0177] A third alarm signal is generated based on the leakage risk level information and sent to the user or a second control sub-signal is sent to the biological culture module.

[0178] like Figure 15a As shown, in some embodiments, when a leak signal is detected in the detachable pipeline connecting bioreactor A and purification device B, the current leak object and leak risk level can be further analyzed based on the leak signal. Specifically, when a unilateral leak is detected, such as a leak signal M1 on the left (specifically, a location-based leak sensing line can detect the specific leak area or location), or a leak signal M2 on the right, then the bioreactor A and the pipeline, or the pipeline and purification device B, are replaced accordingly. Alternatively, when a continuously distributed leak signal M3 is detected, it can be suggested that the user replace bioreactor A, purification device B, and the intermediate pipeline simultaneously. Alternatively, when a leak signal M4 disconnected from the device is detected, it is suggested that the user replace only the pipeline. In this embodiment, the leak risk level of the pipeline and device is preferably determined based on real-time monitoring signals, and a preferred "dynamic maintenance" solution is provided to the user based on the risk level.

[0179] In some embodiments, the first risk status further includes: gas leakage information, and the production method further includes the step of:

[0180] S006 When the biological culture module is turned on, the gas around the biological culture module is collected; S007 The gas is detected to obtain a first index value of the gas; S008 Based on the first index value, it is determined whether the biological culture module is leaking gas, and if so, a leak signal is generated; Correspondingly, S004 includes: 6) When the management module detects that the biological reactor is leaking liquid and simultaneously detects the leak signal, it sends a first control signal indicating that production should be stopped to the biological culture module; and 7) When the management module detects that the biological reactor is leaking liquid but does not detect the leak signal, it sends a second control signal to the biological culture module to suspend production; and 8) When the management module detects the leak signal but the biological reactor is not at risk of leakage, it sends a fifth alarm signal to the user to prompt the user to check the leak signal. In this embodiment of the invention, it is preferable to use an automated monitoring and user collaboration method to perform "dynamic maintenance" of the production line.

[0181] In some embodiments, a pre-inspection of the quality of the on-site assembly line is also performed before production begins. For example, prior to S002, the following steps are also included:

[0182] Liquid is introduced into the biological culture module so that the liquid flows sequentially through the biological reaction device, the purification device, and the collection device;

[0183] Collect a second set consisting of at least one reading from at least one leakage sensing line;

[0184] A third risk state of the biological culture module is determined based on the second set, the third risk state including:

[0185] The object of the leak;

[0186] A fourth alarm signal is generated and sent to the user based on the third risk status.

[0187] Users can manually check the assembly quality of the production line and replace parts based on the fourth alarm signal.

[0188] For example, in some embodiments, if no leakage signal is detected in the production line, the assembly of the production line is determined to be correct.

[0189] It can remind users to start production.

[0190] For example, in some embodiments, when the reading of one of the leakage sensing lines is detected to be greater than a preset threshold, the corresponding detachable pipe and the devices connected to both sides of it are identified as leakage objects.

[0191] Preferably, in some embodiments, one or more devices in the biological culture module can be disposable devices, such as disposable reaction bags, disposable filter bags, disposable collection bags, disposable mixing bags, etc.

[0192] In some embodiments, the collection bag can withstand low temperatures to meet the repeated freeze-thaw requirements of biological products.

[0193] For example, in some embodiments, the bioreactor includes: a bioreactor bag for culturing microorganisms, a first pressure monitoring unit for monitoring the hydraulic pressure in the bioreactor bag, and a first leakage monitoring module correspondingly disposed at the bottom of the bioreactor bag. The method includes the steps of: S102 introducing a preset amount of gas or liquid into the bioreactor bag through a first inlet or a second inlet on the bioreactor bag for culturing microorganisms or cells; S104 monitoring the hydraulic pressure of the first pressure monitoring unit and the leakage value of the first leakage monitoring module in real time; S106 determining the risk status (i.e., the second risk status) of the bioreactor based on the hydraulic pressure and the leakage value, the risk status including: a safe status, and / or a pending status, and / or a leakage status; wherein, S106 includes: S61 determining the pressure drop risk of the bioreactor bag based on the change parameters of the hydraulic pressure; wherein, when When the changing parameter falls within a preset first pressure reduction threshold, the pressure reduction risk is level one; when the changing parameter falls within a preset second pressure reduction threshold, the pressure reduction risk is level two; otherwise, the pressure reduction risk is level zero. S62 determines the leakage risk of the reaction bag based on the leakage value, wherein when the leakage value falls within a preset first leakage threshold, the leakage risk is level one; when the leakage value falls within a preset second leakage threshold, the leakage risk is level two; otherwise, the leakage risk is level zero. S63 determines the risk status based on the pressure reduction risk in S61 and / or the leakage risk in S62; wherein when the pressure reduction risk is level two and / or the leakage risk is level two, the risk status is a leakage status; when both the pressure reduction risk and the leakage risk are level zero, the risk status is a safe status; otherwise, the risk status is a pending status.

[0194] Of course, it is understood that one or more devices (e.g., chromatography apparatus) in the embodiments of the present invention may also be commercially available products.

[0195] In some embodiments, a condensate guiding zone is further formed in the top region of the first space within the reaction bag, for guiding condensate to condense within the condensate guiding zone and to flow back to the second space through the guiding path provided by the condensate guiding zone; correspondingly, the method further includes the steps of: S108 issuing corresponding warning signals to the user when the risk state is detected as pending or leakage state; S110 performing secondary monitoring of the pressure reduction risk and / or leakage risk of the bioreactor in response to the feedback signal issued by the user, or receiving or correcting the warning signal.

[0196] It is understood that the bioreactor and support device in the embodiments of the present invention can be any of the bioreactors and support devices described in the above embodiments, and will not be repeated here.

[0197] In some embodiments, each device is equipped with an aerosol filter at its outlet. In some embodiments, the pore size of each ultrafiltration device can be flexibly configured according to the size of the microorganisms and the size of the active substance. In some embodiments, a biosafety sampling device (negative pressure sampler) or a pre-set, pollution-free sampling bag is also provided to ensure the safety of sampling during the culture process. In some embodiments, the entire production line can automate processes such as microbial concentration, supernatant concentration, liquid exchange, ultrafiltration, inactivation, chromatography, and mixing. In some embodiments, the management module can automate the replenishment or configuration of various culture media and buffers.

[0198] This invention effectively ensures the safety and continuous operation of a disposable production line through real-time risk monitoring and dynamic maintenance. This also allows the production line in this invention to be installed in biosafety isolators or biosafety cabinets with relatively low biosafety protection levels.

[0199] Therefore, in practical applications, staff can complete the initial research and development of highly pathogenic microorganisms or vaccines and other biological products in low-risk biological laboratories (such as P1, P2 laboratories, etc.), which will effectively reduce the difficulty and cost of developing biological products.

[0200] Preferably, the disposable production line provided by the present invention can be used for small-scale (e.g., 2-20L) cultivation of highly pathogenic viruses and bacteria, exploration of conditions for inactivated vaccine production, product line development, etc.

[0201] like Figure 16 As shown, corresponding to the production method provided in the above embodiments, the present invention also provides a production system for disposable highly pathogenic microorganisms or inactivated vaccines, comprising:

[0202] Customizable and assembleable biological culture modules, and a management module for controlling the operating status of the biological culture modules;

[0203] The biological culture module includes: a bioreactor for culturing microorganisms, an intermediate device for processing liquid containing microorganisms, a purification device for purifying liquid, and a collection device. The intermediate device is connected to any one or both of the bioreactor and the purification device via at least one detachable pipe for collecting liquid from the bioreactor and / or the purification device and processing the liquid. At least one leakage sensing line is arranged along the length of the detachable pipe. A leakage reading acquisition module 01 is configured to acquire a first set of readings from at least one leakage sensing line in real time when the biological culture module is turned on.

[0204] The leakage reading processing module 02 is configured to determine the first risk state of the biological culture module through a first set, the first risk state including: leakage object, and / or leakage risk level; the management module is configured to generate corresponding control signals or instructions according to the first risk state and send them to the biological culture module; wherein, the biological culture module responds to the control signals or instructions to choose to continue production, suspend production or stop production.

[0205] In some embodiments, the reading includes a current value, and the leakage reading processing module 02 is further configured to generate a first leakage signal including a first risk status information when the current value of the detachable pipe is greater than a preset second threshold, and the first leakage signal is associated with one or two devices connected to the detachable pipe.

[0206] Accordingly, the management module includes:

[0207] The first monitoring unit identifies the corresponding device and detachable pipeline as the first leakage object when the management module detects two first leakage signals associated with the device and generates a second control signal indicating a production halt. The second control signal includes information about the first leakage object.

[0208] In some embodiments, the management module further includes:

[0209] The second monitoring unit is configured to identify the corresponding device as the second leaking object and the detachable pipe as the first leaking object when a first leaking signal is detected, and to send a first alarm signal to the user.

[0210] The third monitoring unit is configured to monitor whether the user issues a corresponding instruction within a first alarm time after the first alarm signal is issued; wherein the instruction includes: a first instruction indicating cancellation of the alarm, and / or, a second instruction indicating receipt of the alarm, and / or, a third instruction indicating correction of the alarm, and the third instruction includes: the corrected leak object;

[0211] If yes, the received instruction is sent to the biological culture module; otherwise, a second control signal indicating a halt to production is sent directly to the biological culture module.

[0212] In some embodiments, a gas leakage detection module 03 is also included, which is configured to collect the gas around the biological culture module when the biological culture module is turned on; detect the gas to obtain a first index value of the gas; determine whether the biological culture module has leaked gas based on the first index value, and generate a gas leakage signal if so. Accordingly, the management module is configured to: (1) when the management module detects that the biological reactor is a leaking object and simultaneously detects a gas leakage signal, send a first control signal indicating that production should be stopped to the biological culture module; (2) when the management module detects that the biological reactor is a leaking object but does not detect a gas leakage signal, send a second control signal suspending production to the biological culture module; and (3) when the management module detects a gas leakage signal but the biological reactor is not at risk of leaking gas, send a fifth alarm signal to the user to prompt the user to check the gas leakage signal.

[0213] In some embodiments, a warning module 07 for issuing various alarm signals to the user is also included.

[0214] Specifically, embodiments of the present invention can realize functions such as automatic leakage early warning for production lines and automatic disinfection and sterilization of production lines.

[0215] This is to avoid or mitigate the pollution problems caused by aerosol leakage during centrifugation and culture processes in traditional fixed culture modes.

[0216] In some embodiments, a user-defined module 05 is also included, configured to receive control commands input by the user.

[0217] For example, in some embodiments, users can also flexibly customize the nodes from the start to the end of the culture through the user-defined module 05 (such as setting a node to automatically enter culture concentration or collection) to quickly explore different growth conditions.

[0218] In some embodiments, the system further includes a disinfection scheme generation module 08, configured to generate a disinfection scheme based on a control signal, wherein the disinfection scheme includes: a disinfection object, etc.

[0219] In some embodiments, the system further includes: a disinfection module 09, configured to, upon receiving a first control signal,

[0220] Initiate the sterilization process for the biological culture module.

[0221] In some embodiments, the management module is further configured to determine whether the number of leaking objects exceeds a preset first threshold; if so, the management module sends a first control signal to the biological culture module indicating a halt to production, wherein the first control signal includes a stop signal indicating a halt to production and a disinfection signal indicating the initiation of disinfection; if not, the management module sends a second control signal to the biological culture module indicating a suspension of production, wherein the second control signal includes: a suspension object that needs to be suspended; wherein the suspension object includes: a leaking object, and one or more detachable pipes and devices connected to the leaking object;

[0222] In some embodiments, the reading includes a current value, and accordingly, the leakage reading processing module is configured to generate a first leakage signal including first risk status information when the current value of the removable pipe is greater than a preset second threshold, and the first leakage signal is associated with one or two devices connected to the removable pipe.

[0223] Accordingly, the management module is also configured to: (4) when the management module detects that the device is associated with two first leakage signals, identify the corresponding device and detachable pipe as the first leakage object and generate a second control signal indicating that production is suspended; wherein the second control signal includes: information on the first leakage object.

[0224] And / or, the management module is also configured to: (5) when a first leak signal is detected associated with the device,

[0225] The corresponding device is identified as the second leak object and the detachable pipe is identified as the first leak object, and a first alarm signal is sent to the user; (6) The management module monitors whether the user issues a corresponding instruction within the first alarm time after the first alarm signal is issued; wherein the instruction includes: a first instruction indicating cancellation of the alarm, and / or, a second instruction indicating receipt of the alarm, and / or, a third instruction indicating correction of the alarm, and the third instruction includes: the corrected leak object; if yes, the received instruction is sent to the biological culture module; if no, a second control signal indicating suspension of production is sent directly to the biological culture module.

[0226] In some embodiments, the detachable pipe includes: an internal pipe for conveying fluid, and an external pipe for protecting the internal pipe; wherein at least one first leakage sensing line is provided on the outer side of the internal pipe along its length, and at least one second leakage sensing line is provided on the outer side of the external pipe along its length; and the reading includes: the current value of the leakage sensing line, and accordingly, the leakage reading processing module is configured to: (i) determine that leakage has occurred in the internal pipe when the current value of the internal pipe is detected to be greater than a preset second threshold; (ii) generate a second leakage signal associated with the detachable pipe, the second... The leakage signal includes: pipeline information and / or leakage risk level; and (iii) when the current value of the external pipeline is detected to be greater than a preset second threshold, it is determined that the external pipeline has leaked; (iv) a third leakage signal associated with the pipeline is generated, the third leakage signal including: pipeline information and / or leakage risk level; accordingly, the management module is also configured to: when the management module detects that the detachable pipeline is associated with the second leakage signal and the third leakage signal, identify the detachable pipeline as a leakage object, and generate a first control sub-signal indicating a suspension of production based on the leakage object to be sent to the bioculture module.

[0227] And / or, the management module is also configured to: issue a second alarm signal to the user when the management module detects only a second leak signal associated with the detachable pipe; or, when the management module detects only a third leak signal associated with the detachable pipe, determine the leak risk level of the external pipe and device based on the reading, and generate a second control sub-signal indicating a suspension of production based on the leak risk level to be sent to the biological culture module.

[0228] And / or, the management module is also configured to: the clarification filtration device includes: a first chamber for filtration, and a second chamber disposed on the outer layer of the first chamber, the first chamber containing filter material for filtering the liquid, and one end of the first chamber being connected to a bioreactor or a purification device, and the other end of the first chamber being connected to an intermediate device, a purification device, or a collection device. Leakage sensing units are respectively disposed on the outer walls of the first chamber and the second chamber, and the leakage sensing units are leakage sensors or leakage sensing lines.

[0229] In some embodiments, the step of determining the leakage risk level of pipeline leakage and device leakage based on readings includes: (1) determining the distribution characteristics of the leakage location of the internal pipeline based on the readings, wherein the distribution characteristics include: continuous distribution connected to both sides of the pipeline, and / or regional distribution disconnected from both sides of the pipeline; (2) when the distribution characteristics are detected as continuous distribution, the leakage risk level at the interface of the device on both sides of the pipeline is determined to be Level 1, and the leakage risk level of the pipeline is determined to be Level 2; (3) when the distribution characteristics are detected as regional distribution, the leakage risk level of the pipeline is determined to be Level 1, and the leakage risk level of the device on both sides of the pipeline is determined to be Level 2; accordingly, the management module is also configured to: generate a third alarm signal based on the leakage risk level information and send it to the user or a second control sub-signal to be sent to the biological culture module.

[0230] In some embodiments, the system further includes a pre-assessment module configured to: (1) introduce liquid into the biological culture module such that the liquid flows sequentially through the bioreactor, the purification device, and the collection device; (2) acquire a second set consisting of at least one reading on at least one leak sensing line; (3) determine a third risk state of the biological culture module based on the second set, the third risk state including a leaking object; and (4) generate and send a fourth alarm signal to the user based on the third risk state.

[0231] In some embodiments, the bioreactor includes: a bioreactor bag for culturing microorganisms, a first pressure monitoring unit for monitoring the hydraulic pressure in the bioreactor bag, and a first leakage monitoring module correspondingly disposed at the bottom of the bioreactor bag. The management module further includes: a sample addition module 20 configured to, in response to a corresponding control signal issued by a user or a working status monitoring module, introduce gas and / or liquid into the bioreactor bag through a first inlet and / or a second inlet for biological culture; a leakage monitoring module 30 configured to, for pressure monitoring of the liquid inside the second space to collect hydraulic pressure, and to, for leakage monitoring of the bottom of the bioreactor bag to collect leakage value; and a risk analysis module 40 configured to, for determining the risk status of the bioreactor system based on the hydraulic pressure and leakage value, the risk status including: a safe status, and / or a pending status, and / or a leakage status; wherein the risk analysis module includes: a hydraulic pressure analysis unit 40-1 configured to, for determining the pressure reduction risk of the bioreactor bag based on the change parameters of the hydraulic pressure; wherein the pressure reduction risk is divided into the following categories according to the magnitude of the change parameters:

[0232] The system includes three risk levels: Level 0, Level 1, and Level 2. A leakage analysis unit 40-2 is configured to determine the leakage risk of the reaction bag based on the leakage value. The leakage risk is categorized into Level 0, Level 1, and Level 2 based on the leakage value. A risk analysis unit 40-3 is configured to determine the risk status based on the pressure reduction risk and leakage risk. When the pressure reduction risk is Level 2 and / or the leakage risk is Level 2, the risk status is a leakage status. When both the pressure reduction risk and leakage risk are Level 0, the risk status is a safe status. When the pressure reduction risk is Level 1 and the leakage risk is Level 0, or when the pressure reduction risk is Level 0 and the leakage risk is Level 1, the risk status is a pending status.

[0233] In some embodiments, a condensate guiding zone is further formed in the top region of the first space. The condensate guiding zone guides condensate to condense within the guiding zone and flows back to the second space through the guiding path provided by the condensate guiding zone. Accordingly, the feedback signal includes: a first feedback signal for heating the condensate guiding zone and a second feedback signal for receiving the warning signal. The management module further includes: a warning correction module 60, which includes: a secondary monitoring unit 60-1, configured to heat the condensate guiding zone in response to the first feedback signal when the pressure drop risk is level one and the leakage risk is level zero; and a correction unit 60-2, which, when it is detected that the pressure drop risk drops to level zero during or after heating, corrects the risk state to a safe state in response to the first correction signal issued by the user; otherwise, it corrects the risk state to a leakage state in response to the second correction signal issued by the user.

[0234] This invention also provides a method for disposable aseptic microbial culture and purification of effective substances, to achieve safe, stable, and low-cost aseptic microbial culture. In some embodiments, the method includes: S001 providing a customizable and assembleable biological culture module, and a management module for controlling the operating status of the biological culture module; wherein the biological culture module includes: a bioreactor for culturing microorganisms, a purification device connected to the bioreactor via at least one detachable pipe, and a collection device connected to the bioreactor and / or the purification device via at least one detachable pipe, wherein at least one leakage sensing line is arranged along the length of the detachable pipe; S002 when the biological culture module is turned on, a first set consisting of at least one reading on at least one of the leakage sensing lines is collected in real time; S003 the first set is used to determine a first risk state of the biological culture module, the first risk state including: leakage object, and / or leakage risk level;

[0235] S004 The management module generates a corresponding control signal or instruction based on the first risk status and sends it to the biological culture module; S005 The biological culture module responds to the control signal or instruction by selecting to continue production, suspend production, or stop production.

[0236] Further, in some embodiments, the first risk state further includes: the microbial culture state, and the method further includes the steps of: S009 when the biological culture module is turned on, collecting the waste gas generated by the biological culture module during operation; S010 detecting the waste gas to obtain a second index value of the gas; S011 judging the microbial culture state according to the second index value, the culture state including: an abnormal state indicating that culture needs to be paused or culture conditions adjusted, a peak state indicating that the microbial content has met a preset index, and a sustainable state; correspondingly, step S004 includes: when the management module detects that the culture state is at its peak, sending a fourth control signal to the biological culture module indicating that the operation of the bioreactor device should be stopped; or,

[0237] When the management module detects that the cultivation status is abnormal, it sends a fourth alarm signal to the user. It is understood that embodiments of the present invention may include steps that are the same as or similar to any of the above embodiments, and will not be repeated here.

[0238] In this embodiment of the invention, the gases (i.e., waste gases) generated during the cultivation process can be collected for monitoring relevant indicators of the gas-producing microorganisms. Gas detection can be achieved using a gas detection device connected to the biological cultivation module.

[0239] In some embodiments, the purification apparatus further includes one or more of the following: an ultrafiltration deactivation device, an ultrafiltration liquid exchange device, a clarification filtration device, and a supernatant concentration device. Embodiments of the present invention can automatically realize processes such as concentration, supernatant concentration, liquid exchange, ultrafiltration, inactivation, chromatography, and mixing of liquids containing microorganisms. For example, it can automatically control the addition of microbial culture medium, the addition of lysis buffer, liquid exchange, automatic pH adjustment, clarification filtration, and ultrafiltration.

[0240] In some embodiments, the management module can also monitor pressure changes within the reactor during the cultivation process and issue real-time warnings in case of leaks. Additionally, for the cultivation of some gas-producing microorganisms, the module can monitor gas production in real time and increase emissions of waste gases.

[0241] In this embodiment, the production line can be used for automated extraction of biological products such as proteins, polysaccharides, and lipids.

[0242] The production line in this embodiment is particularly suitable for small-scale (e.g., 2-20L) aseptic culture of microorganisms.

[0243] In this embodiment, the entire production line can be operated in a clean bench, biosafety cabinet, or isolator to provide dual protection against contamination. Furthermore, the automated monitoring of the entire line can assist users in quickly determining process conditions.

[0244] Similarly, in contrast to the traditional fixed culture mode, this invention proposes a flexible assembly and "dynamic maintenance" disposable production line for aseptic culture and purification of microorganisms. The flexible assembly feature of this production line can meet the user's early exploration needs for microbial process conditions (specifically, the user can assemble the production line on-site in the laboratory in a quick disassembly manner according to the process design).

[0245] Furthermore, during the real-time operation of the production line, "dynamic maintenance" can be achieved through flexible assembly and risk monitoring, ensuring continuous and stable operation and reducing application costs. Specifically, the "dynamic maintenance" model can improve the effective utilization rate of disposable equipment or pipelines (or, in other words, increase the production qualification rate of disposable production lines). Moreover, this "dynamic maintenance" model reduces the need for manual labor and simplifies manual operations (e.g., by providing optimized dynamic maintenance solutions to simplify production line maintenance), thus facilitating the mass production of the production line.

[0246] Corresponding to the above embodiments, the present invention also provides a disposable microbial aseptic culture and effective substance purification system.

[0247] The system includes: a customizable bioculture module and a management module for controlling the operating status of the bioculture module; wherein the bioculture module includes: a bioreactor for culturing microorganisms, a purification device connected to the bioreactor via at least one detachable pipe, and a collection device connected to the bioreactor and / or purification device via at least one detachable pipe, wherein at least one leakage sensing line is arranged along the length of the detachable pipe; a leakage reading acquisition module 01 is configured to acquire a first set of readings from at least one of the at least one leakage sensing line in real time when the bioculture module is running; a leakage reading processing module 02 is configured to determine a first risk state of the bioculture module based on the first set, the first risk state including: leakage object, and / or leakage risk level; and a management module is configured to generate corresponding control signals or instructions based on the first risk state and send them to the bioculture module; wherein the bioculture module responds to the control signals or instructions by selecting to continue production, suspend production, or stop production.

[0248] In some embodiments, a gas generation monitoring module 04 is further included, which is configured to: (1) collect the waste gas generated by the biological culture module during operation when the biological culture module is turned on; (2) detect the waste gas to obtain a second index value of the gas; and (3) determine the culture status of the microorganisms based on the first index value. The culture status includes: an abnormal state indicating that the culture needs to be paused or the culture conditions adjusted, a peak state indicating that the content of the microorganisms has met the preset index, and a sustainable state. Accordingly, the management module is configured to send a fourth control signal indicating that the operation of the bioreactor is stopped to the biological culture module when the management module detects that the culture status is at its peak state; or, when the management module detects that the culture status is abnormal, send a fourth alarm signal to the user. It is understood that the various devices or modules in the embodiments of the present invention can be flexibly configured according to the actual needs of aseptic microbial culture. For example, the system can implement any step or function in the above embodiments. For another example, the system may include the same or similar devices or modules as in the above embodiments, which will not be described again here.

[0249] It should be noted that the production system in this invention can also be scaled up to meet the needs of vaccine production at different scales (such as pilot-scale vaccine production, commercial vaccine production, etc.) or microbial culture at different scales (such as 50L or other scales). Parameters such as thresholds or threshold ranges in this invention can be freely set by the user through the user-defined module 05. The user in this invention can be a computer connected to the semi-automatic control system or a laboratory operator. Besides being applicable to the culture of highly pathogenic microorganisms (such as bacteria and viruses), this invention can also be applied to other microorganisms that may cause varying degrees of pollution or impact on the environment, or other microorganisms that may cause varying degrees of pathogenicity or impact on humans or animals and plants. Of course, this invention can also be applied to the culture of various biological cells, such as animal cells, plant cells, or single-celled organisms. Through the above description of the embodiments, those skilled in the art can clearly understand that the methods of the above embodiments can be implemented by means of software plus necessary general-purpose hardware platforms; of course, they can also be implemented by hardware, but in many cases, the former is a better implementation method. Based on this understanding, the technical solution of this invention, or the part that contributes to the prior art, can be embodied in the form of a software product. This computer software product is stored in a storage medium (such as ROM / RAM, magnetic disk, optical disk) and includes several instructions to cause a computer terminal (which may be a mobile phone, computer, server, or network device, etc.) to execute the methods described in the various embodiments of this invention. The embodiments of this invention have been described above with reference to the accompanying drawings. However, this invention is not limited to the specific embodiments described above. The specific embodiments described above are merely illustrative and not restrictive. Those skilled in the art, under the guidance of this invention, can make many modifications without departing from the spirit and scope of the claims, and all of these modifications are within the protection scope of this invention.

Claims

1. A method for producing a single-use highly pathogenic microorganism or inactivated vaccine, characterized in that, Including the following steps: S001 provides a customizable bioculture module and a management module for controlling the operating status of the bioculture module; wherein, the bioculture module includes: a bioreactor for culturing microorganisms, an intermediate device for processing a liquid containing the microorganisms, and a purification device for purifying the liquid; the intermediate device is connected to any one or both of the bioreactor and the purification device via at least one detachable pipe for collecting the liquid from the bioreactor and / or the purification device and processing the liquid, wherein at least one leakage sensing line is arranged along the length of the detachable pipe; S002 When the biological culture module is turned on, a first set consisting of at least one reading on at least one of the leakage sensing lines is collected in real time; S003 determines the first risk status of the biological culture module through the first set, the first risk status including: leakage object, and / or leakage risk level; S004 The management module generates corresponding control signals or instructions based on the risk status and sends them to the biological culture module; S005 The biological culture module responds to the control signal or instruction by selecting to continue production, suspend production, or stop production; The bioreactor includes: a bioreactor bag for culturing microorganisms, a first pressure monitoring unit for monitoring the hydraulic pressure in the bioreactor bag, and a first leakage monitoring module correspondingly disposed at the bottom of the bioreactor bag; a condensate guiding zone is also formed in the top region of the first space inside the bioreactor bag to guide condensate to condense in the condensate guiding zone and flow back to the second space through the guiding path provided by the condensate guiding zone; the method further includes the following steps: S102 introduces a preset amount of gas or liquid into the reaction bag through the first or second inlet on the reaction bag to culture microorganisms or cells; S104 monitors the hydraulic pressure of the first pressure monitoring unit and the leakage value of the first leakage monitoring module in real time. S106 determines the second risk state of the bioreactor based on the hydraulic pressure and the leakage value. The second risk state includes: a safe state, and / or a pending state, and / or a leakage state. Specifically, S106 includes: when the changing parameter is within a preset first pressure drop threshold, the pressure drop risk is level one; when the changing parameter is within a preset second pressure drop threshold, the pressure drop risk is level two; otherwise, the pressure drop risk is level zero. When the leakage value is within a preset first leakage threshold, the leakage risk is level one; when the leakage value is within a preset second leakage threshold, the leakage risk is level two; otherwise, the leakage risk is level zero. When the pressure drop risk is level two and / or the leakage risk is level two, the second risk state is a leakage state. When both the pressure drop risk and the leakage risk are level zero, the second risk state is a safe state; otherwise, the second risk state is a pending state. When the second risk state is detected as either pending or leaking, a corresponding warning signal is issued to the user. When the pressure drop risk is level one and the leaking risk is level zero, the condensate guide area is heated in response to the first feedback signal issued by the user. When the pressure drop risk is detected to drop to level zero during or after heating, the risk state is corrected to a safe state in response to the first correction signal issued by the user. Otherwise, the risk state is corrected to a leaking state in response to the second correction signal issued by the user.

2. The method for producing a single-use highly pathogenic microorganism or inactivated vaccine according to claim 1, characterized in that, S004 includes the following steps: The management module determines whether the number of leaking objects exceeds a preset first threshold. If so, the management module sends a first control signal to the biological culture module indicating that production has been stopped, wherein the first control signal includes: a stop signal indicating that production has been stopped, and a disinfection signal indicating that disinfection has been started; If not, the management module sends a second control signal to the biological culture module indicating a production halt, the second control signal including: a halting object requiring production halt; wherein, the halting object includes: a leaking object, and one or more of the detachable pipes and / or one or more devices connected to the leaking object; And / or, the purification apparatus includes one or more of the following: an ultrafiltration deactivation device, an ultrafiltration liquid exchange device, a clarification filtration device, and a supernatant concentration device; the intermediate device includes one or more of the following: a chromatography device and a mixing device; And / or, the device is provided with a diaphragm pump or peristaltic pump for conveying liquid at the interface between the device and the detachable pipe.

3. The method for producing a single-use highly pathogenic microorganism or inactivated vaccine according to claim 1, characterized in that, The reading includes: current value, and S003 includes the following steps: When the current value of the detachable pipe is greater than a preset second threshold, a first leakage signal including first risk status information is generated, and the first leakage signal is associated with one or two devices connected to the detachable pipe. Accordingly, S004 includes the following steps: 1) When the management module detects that the device is associated with two first leakage signals, it identifies the corresponding device and the detachable pipe as the first leakage object and generates a second control signal indicating a production halt; wherein, the second control signal includes: information about the first leakage object; And / or, 2) When a first leakage signal is detected associated with the device, the corresponding device is identified as the second leakage object and the detachable pipe is identified as the first leakage object, and a first alarm signal is sent to the user; 3) The management module monitors whether the user issues a corresponding instruction within the first alarm time after the first alarm signal is issued; wherein, the instruction includes: a first instruction indicating cancellation of the alarm, and / or, a second instruction indicating receipt of the alarm, and / or, a third instruction indicating correction of the alarm, and the third instruction includes: the corrected leakage object; if yes, the received instruction is sent to the biological culture module; if no, a second control signal indicating suspension of production is sent directly to the biological culture module.

4. The method for producing a single-use highly pathogenic microorganism or inactivated vaccine according to claim 1 or 2, characterized in that, The detachable pipe includes: an internal pipe for conveying fluid, and an external pipe for protecting the internal pipe; wherein, at least one first leakage sensing line is provided on the outer side of the internal pipe along its length, and at least one second leakage sensing line is provided on the outer side of the external pipe along its length; and the reading includes: the current value of the leakage sensing line, and correspondingly, step S003 includes: (i) When the current value of the internal pipe is detected to be greater than the preset second threshold, it is determined that the internal pipe is leaking; (ii) Generate a second leakage signal associated with the removable pipe, the second leakage signal including: pipe information, and / or leakage risk level; And (iii) when the current value of the external pipe is detected to be greater than the preset second threshold, it is determined that the external pipe is leaking; (iv) Generate a third leakage signal associated with the pipeline, the third leakage signal including: pipeline information, and / or leakage risk level; Accordingly, S004 includes: When the management module detects that the detachable pipe is associated with both the second and third leakage signals, it identifies the detachable pipe as a leakage target and generates a first control sub-signal indicating a production halt based on the leakage target, which is then sent to the bio-culture module; or, When the management module detects only the second leak signal associated with the detachable pipe, it issues a second alarm signal to the user; or, When the management module detects that the detachable pipe is only associated with the third leakage signal, it determines the leakage risk level of the pipe and the device based on the reading, and generates a second control sub-signal indicating a production halt based on the leakage risk level and sends it to the biological culture module. And / or, the purification device includes: a concentration filtration device, and the concentration filtration device includes: a first chamber for filtration, and a second chamber disposed outside the first chamber, the first chamber being provided with a filter material for filtering the liquid, and one end of the first chamber being connected to a bioreactor, and the other end of the first chamber being connected to a corresponding intermediate device or other purification device. The first chamber and the second chamber are respectively provided with a leakage sensing unit on their outer walls, and the leakage sensing unit is a leakage sensor or a leakage sensing line.

5. The method for producing a single-use highly pathogenic microorganism or inactivated vaccine according to claim 4, characterized in that, The steps for determining the leakage risk level of the pipeline and the device based on the readings include: The distribution characteristics of the leakage location of the pipeline are determined based on the readings, wherein the distribution characteristics include: a continuous distribution connected to both sides of the pipeline, and / or a regional distribution disconnected from both sides of the pipeline; When the distribution feature is detected to be continuous, the leakage risk level at the interface of the device on both sides of the pipeline is determined to be Level 1, and the leakage risk level of the pipeline is determined to be Level 2. When the distribution feature is detected to be a regional distribution, the leakage risk level of the pipeline is determined to be Level 1, and the leakage risk level of the devices on both sides of the pipeline is determined to be Level 2. Accordingly, S004 further includes the step of: A third alarm signal is generated based on the leakage risk level information and sent to the user, or a second control sub-signal is generated and sent to the biological culture module. And / or, alarm signals may include one or more of the following forms: voice alarm, and / or SMS alarm, and / or traffic light alarm.

6. The method for producing a single-use highly pathogenic microorganism or inactivated vaccine according to claim 1, characterized in that, The first risk status also includes: gas leakage information, and the production method further includes the following steps: S006 When the biological culture module is turned on, the gas around the biological culture module is collected; S007 Detects the gas to obtain a first index value for the gas; S008 determines whether the biological culture module is leaking based on the value of the first indicator; if so, a leak signal is generated. Accordingly, S004 includes: When the management module detects that the bioreactor is leaking liquid and simultaneously detects the gas leak signal, it sends a first control signal to the bioculture module indicating that production should be stopped. And when the management module detects that the bioreactor is leaking liquid but does not detect the gas leak signal, it sends a second control signal to the bioculture module indicating that production should be suspended. And when the management module detects the gas leak signal but the bioreactor has no immediate risk of liquid leakage, it sends a fifth alarm signal to the user to prompt the user to check the gas leak signal; and / or, before S002, the following steps are also included: Liquid is introduced into the biological culture module so that the liquid flows sequentially through the biological reaction device, the purification device, and the collection device; Collect a second set consisting of at least one reading from at least one leakage sensing line; The third risk state of the biological culture module is determined based on the second set, and the third risk state includes: leakage object; A fourth alarm signal is generated and sent to the user based on the third risk status.

7. A production system for single-use highly pathogenic microorganisms or inactivated vaccines, characterized in that, include: A customizable bioculture module and a management module for controlling the operating status of the bioculture module; wherein the bioculture module includes: a bioreactor for culturing microorganisms, an intermediate device for processing liquid containing the microorganisms, and a purification device for purifying the liquid; the intermediate device is connected to any one or both of the bioreactor and the purification device via at least one detachable pipe for collecting the liquid from the bioreactor and / or the purification device and processing the liquid, wherein at least one leakage sensing line is arranged along the length of the detachable pipe; The leakage reading acquisition module (01) is configured to acquire, in real time, a first set consisting of at least one reading on at least one of the leakage sensing lines when the biological culture module is turned on; The leakage reading processing module (02) is configured to determine a first risk status of the biological culture module through the first set, the first risk status including: leakage object, and / or leakage risk level; The management module is configured to generate corresponding control signals or instructions based on the risk status and send them to the biological culture module; The biological culture module responds to the control signal or instruction to choose to continue production, pause production, or stop production; The bioreactor includes: a bioreactor bag for culturing microorganisms, a first pressure monitoring unit for monitoring the hydraulic pressure in the bioreactor bag, and a first leakage monitoring module correspondingly disposed at the bottom of the bioreactor bag; a condensate guiding zone is also formed in the top region of the first space inside the bioreactor bag to guide condensate to condense in the condensate guiding zone and to flow back to the second space through the guiding path provided by the condensate guiding zone; the system is also used for: A predetermined amount of gas or liquid is introduced into the reaction bag through the first or second inlet to culture microorganisms or cells. Real-time monitoring of the hydraulic pressure of the first pressure monitoring unit and the leakage value of the first leakage monitoring module; The second risk state of the bioreactor is determined based on the hydraulic pressure and the leakage value. The second risk state includes: a safe state, and / or a pending state, and / or a leakage state. Specifically, it includes: when the changing parameter is within a preset first pressure drop threshold, the pressure drop risk is level one; when the changing parameter is within a preset second pressure drop threshold, the pressure drop risk is level two; otherwise, the pressure drop risk is level zero. Similarly, when the leakage value is within a preset first leakage threshold, the leakage risk is level one; when the leakage value is within a preset second leakage threshold, the leakage risk is level two; otherwise, the leakage risk is level zero. When the pressure drop risk is level two and / or the leakage risk is level two, the second risk state is a leakage state. When both the pressure drop risk and the leakage risk are level zero, the second risk state is a safe state; otherwise, the second risk state is a pending state. When the second risk state is detected as either pending or leaking, a corresponding warning signal is issued to the user. When the pressure drop risk is level one and the leaking risk is level zero, the condensate guide area is heated in response to the first feedback signal issued by the user. When the pressure drop risk is detected to drop to level zero during or after heating, the risk state is corrected to a safe state in response to the first correction signal issued by the user. Otherwise, the risk state is corrected to a leaking state in response to the second correction signal issued by the user.

8. The production system for a single-use highly pathogenic microorganism or inactivated vaccine according to claim 7, characterized in that, The readings include current values, and the leakage reading processing module (02) is further configured to generate a first leakage signal including first risk status information when the current value of the detachable pipe is greater than a preset second threshold, and the first leakage signal is associated with one or two devices connected to the detachable pipe. Accordingly, the management module includes: The first monitoring unit identifies the corresponding device and the detachable pipe as the first leakage object when the management module detects that the device is associated with two first leakage signals and generates a second control signal indicating a production halt; wherein the second control signal includes: information about the first leakage object; And / or, The second monitoring unit is configured to identify the corresponding device as the second leaking object and the detachable pipe as the first leaking object when a first leaking signal is detected associated with the device, and to send a first alarm signal to the user. The third monitoring unit is configured to monitor whether the user issues a corresponding instruction within a first alarm time after the first alarm signal is issued; wherein the instruction includes: a first instruction indicating cancellation of the alarm, and / or, a second instruction indicating receipt of the alarm, and / or, a third instruction indicating correction of the alarm, and the third instruction includes: the corrected leak object; If yes, the received instruction is sent to the biological culture module; otherwise, a second control signal indicating a halt to production is sent directly to the biological culture module.

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