CN117143725A - An automatic human-like large-scale stem cell culture equipment with low loss rate - Google Patents

Abstract

An automatic culture device for human-like large-scale stem cells with low loss rate belongs to the field of biological equipment, and comprises a plurality of culture modules, a circulation module, a shaking module and a pressure regulating module; the circulation module is connected with the mixing module and the culture module, so that the culture medium circularly flows between the mixing module and the culture module; the shaking module is connected with the culture module and used for driving the culture module to shake; the pressure regulation module may apply a periodically varying pressure field within the cell culture tank. The invention realizes the low loss rate and the expandable large-scale cell culture in the process of uniformly mixing the cell and the nutrient substances, realizes the control of key parameters of the cell culture environment imitating the environment in human body by a multi-physical field decoupling method, has the characteristic of greatly improving the quantity and the quality of the cells cultured in vitro, and provides technical support for the development of the cell treatment industry.

Description

An automatic human-like large-scale stem cell culture equipment with low loss rate

Technical field

The invention belongs to the field of biological equipment, and specifically relates to a human-like large-scale stem cell automatic culture equipment with a low loss rate.

Background technique

As a major international strategic and forward-looking scientific topic, stem cell therapy has obvious therapeutic effects on heart failure, neurodegenerative diseases and many other serious diseases that lack effective treatments. Large-scale, high-quality, safe and stable cell culture equipment is a bottleneck restricting the promotion of stem cell therapy. Three-dimensional cell culture technology and equipment can significantly improve the production efficiency of stem cells, ensure production quality, and improve application effects. Therefore, it is of great significance to develop an equipment that improves the quality of cell products, increases the quantity of stem cell production, and has automated production functions.

In large-scale stem cell culture, cell culture volume has a direct impact on cell growth and accumulation of metabolites. Generally speaking, a larger culture volume provides a greater supply of nutrients and oxygen, thereby promoting cell growth and reproduction. However, too large a culture volume may also cause some problems. For example, a larger culture volume may result in a non-uniform culture environment, resulting in differential growth of cells at different locations. The existing solution is to use a large amount of stirring and mixing after the overall culture volume to improve the environmental uniformity between different locations in the equipment. For example, Ou Guoqiang's patent number CN112852633A "A stem cell large-scale culture bioreactor system" The problem of uneven stirring of the culture liquid in the bioreactor tank is solved by adding a spray device and a stirring device. However, mechanical movements such as stirring, spraying, and aeration will inevitably produce shear forces that damage cells, increase cell loss during cell culture, affect cell survival, and limit the increase in culture density. Therefore, it is necessary to invent a low cell loss method. High-efficiency, scalable large-scale stem cell culture equipment.

In order to improve the quality of stem cell production and ensure various physiological indicators of cells, the culture environment will be precisely controlled when cells are cultured in vitro. Existing cell culture equipment mainly detects and regulates pH, temperature and dissolved oxygen concentration in the cell culture environment. For example, Chen Haijia passed the patent number CN115803428A "A method for efficient preparation of exosomes using a stem cell large-scale culture device" The culture microenvironment control system monitors and regulates pH, temperature and dissolved oxygen in the reactor. However, studies have proven that the pressure of the human body environment has an impact on the cell state, proliferation rate, pluripotency and stability of stem cells. However, in the existing stem cell culture process, it is difficult to accurately couple the pressure with environmental parameters such as pH and dissolved oxygen concentration. Regulation, so there is no controllable pressure cultivation environment. Existing stem cell culture methods still have the problem of being unable to control pressure, dissolved oxygen, pH and other environmental factors at the same time, resulting in differences in cell status, proliferation rate, etc. compared with actual survival in the body, hindering the development of stem cell therapy. Therefore, there is a need to invent a stem cell culture device that simulates the human internal environment including blood pressure.

In addition, compared with traditional manual culture equipment, automated production equipment can significantly improve production efficiency and achieve continuous, high-throughput culture operations, thereby avoiding tedious and time-consuming steps that may occur in manual operations. At the same time, with advanced control systems and sensor technology, the culture environment can be monitored and adjusted in real time to ensure the stability and consistency of culture conditions, thereby improving the efficiency and quality of stem cell culture.

In summary, clinical production urgently needs a low-loss, scalable, fully-automatic large-scale stem cell culture equipment. This equipment can couple the pH, pressure and dissolved oxygen concentration of the culture environment during the cell culture process to simulate human cells in vitro. The purpose of the in vivo cell growth environment is to improve the quantity and quality of cultured stem cells, thereby promoting the development of my country's cell therapy industry.

Contents of the invention

The purpose of the present invention is to propose a low loss rate, scalable, fully automatic large-scale stem cell culture equipment in view of the problems existing in the existing cell culture technology in vitro. This equipment can couple the pH and pressure of the culture environment during the cell culture process. and dissolved oxygen concentration, to achieve the purpose of greatly improving the quality and quantity of cultured cells, and providing technical support for the development of the cell therapy industry.

The present application provides a low loss rate human-like large-scale stem cell automatic culture equipment, which includes a fixed table, a culture module is installed on the fixed table; a sterile cover for covering the culture module is provided on the fixed table, A sterilization device is provided in the sterile cover; a control cabinet is provided on one side of the fixed table; a mixing module and a device cabinet are fixed inside the fixed table, where the device cabinet is installed on the left side of the mixing module and passes through the trachea. , silicone tubes, data lines, communication lines, etc.; it also includes several circulation modules, shaking modules, and pressure adjustment modules; the circulation module connects the mixing module and the culture module, so that the enriched culture medium in the mixing module flows into the The culture module, the oligotrophic culture medium in the culture module flows into the mixing module; the shaking module is connected with the culture module and is used to drive the culture module to shake; the pressure regulation module is connected to the culture module Module connection is used to adjust the air pressure within the culture module.

In some possible implementations of the present application, the circulation module includes a first peristaltic pump, a second peristaltic pump, a first one-way valve, a second one-way valve, a first pinch valve, and a second pinch valve; The circulation die hole has two pairs of ports, the first peristaltic pump, the first one-way valve, and the first pinch valve are connected in series between one pair of ports, and the second peristaltic pump, The second one-way valve and the second pinch valve are connected in series between another pair of ports, one port of each pair of ports is connected to the culture module, and the other port of each pair of ports is connected to the Hybrid module connections.

In some possible implementations of the present application, the pressure regulation module includes a first air pipe, a second air pipe, a second bacterial filter, a third bacterial filter, a pressure sensor, and a pressure regulating proportional valve; the second air pipe , the third bacterial filter, the pressure sensor, and one port of the pressure regulating proportional valve are connected in sequence, and the other port of the first trachea, the second bacterial filter, and the pressure regulating proportional valve is connected in sequence. The ports are connected in sequence, and the first trachea and the second trachea are respectively connected to the culture module.

In some possible embodiments of the present application, the culture module includes a culture platform fixedly installed on a fixed platform and several independently working cell culture tanks detachably installed inside the culture platform; the shaking module It includes several shakers; the shakers are all fixed at the bottom end of the culture platform; a flexible semi-wrapped heater can be detachably installed on the table of the shaker; the culture tank of the culture module can be detachably installed on the flexible semi-wrapped heater. In the heater, both the culture tank of the culture module and the flexible semi-wrapped heater can move following the movement of the shaker table.

In some possible embodiments of the present application, a sensor set and a cell screen are provided inside the cell culture tank; a connecting tube is fixed on the lid of the cell culture tank, and one end of the connecting tube is located on the cell culture tank. Outside the tank, the other end is located in the cell culture tank. The connecting tube includes a long tube A, a short tube A and a sampling tube. One end of the sampling tube is located in the cell culture tank; the cell screen is fixed on The periphery of the long tube A is assembled coaxially with the long tube A, and there is a distance between its bottom and the bottom end of the long tube A; the sensor group is a non-contact measurement device, fixedly installed in the cell culture tank In a separate small room inside.

In some possible implementations of the present application, the mixing module includes a mixing tank, a flexible surrounding heater, a first stirring paddle, a second stirring paddle, a first connecting rod, a second connecting rod, a first stirring motor, The second stirring motor, aeration plate, first bacterial filter, long tube set, short tube set, sampling tube, and liquid discard tube;

The first bacterial filter is connected to the aeration disk;

The control cabinet is equipped with a central controller and a multi-parameter transmitter; the sensor group is connected to the multi-parameter transmitter through signal lines and communication lines, and can transmit information to the multi-parameter transmitter and accept multi-parameter transmission. control of organs;

A gas introduction module, a sample processing module and a waste liquid module are installed in the device cabinet; the gas introduction module includes an air flow regulator, a nitrogen flow regulator, a carbon dioxide flow regulator, an oxygen flow regulator, a four-mix and one-inlet system. air valve;

One ends of the air flow regulator, the nitrogen flow regulator, the carbon dioxide flow regulator, and the oxygen flow regulator are respectively connected to one end of the four-mix-one air intake valve. The other end of the valve is connected to the first bacterial filter;

The sample processing module includes an automatic sampler, a cell analyzer, an automatic sampler, and a reagent rack;

The cell analyzer is connected to the automatic sampler, and the automatic sampler is connected to the sampling tube; the reagent rack is connected to the automatic sampler, and the automatic sampler is connected to the sampling tube. connect;

The waste liquid module includes a waste liquid barrel and a liquid waste pump;

The waste liquid bucket is connected to the liquid discarding pump, and the liquid discarding pump is connected to the liquid discarding pipe.

In some possible implementations of the present application, the flexible surrounding heater is fixed around the periphery of the mixing tank to keep the temperature in the mixing tank constant; a long pipe clamp and a short pipe clamp are fixed on the lid of the mixing tank. Pipe clamp; several long pipes can be detachably installed in the long pipe clamp to form a long pipe group, which is used to connect the circulation module to export the culture medium out of the mixing tank; the short pipe clamp can be detachably installed in the short pipe clamp Several short tubes together form a short tube group, which is used to connect the circulation module and introduce the culture medium into the mixing tank; a first stirring motor is detachably installed on the left side of the lid of the mixing tank, and one end of the first connecting rod It is connected to the power output end of the first stirring motor, and the other end is connected to the first stirring paddle. The first stirring paddle is located in the mixing tank near the left side of the aeration plate; the right side of the lid of the mixing tank The second stirring motor can be detachably installed. One end of the second connecting rod is connected to the power output end of the second stirring motor, and the other end is connected to the second stirring paddle. The second stirring paddle is located in the mixing tank on the right side of the aeration plate. side position; the aeration disk is located in the mixing tank near the bottom of the mixing tank, and is connected to one end of the first bacterial filter through a silicone tube.

In some possible implementations of the present application, the circulation module includes a first circulation unit and a second circulation unit. One end of the first circulation unit is connected to the long tube A in the cell culture tank, and the other end is connected to The short tubes in the short tube group in the mixing tank are connected; one end of the second circulation unit is connected to the long tube in the long tube group in the mixing tank, and the other end is connected to the short tube A in the cell culture tank. Connection; the first circulation unit includes a first peristaltic pump, a first one-way valve, and a first pinch valve. One end of the first peristaltic pump is connected to the long pipe A, and the other end is connected to the first one-way valve, The first one-way valve is connected to the first pinch valve, and one end of the first pinch valve that is not connected to the first one-way valve is connected to the short pipe of the short pipe group; the second circulation unit includes a third Two peristaltic pumps are connected to a second one-way valve and a second pinch valve. One end of the second peristaltic pump is connected to the short pipe A, and the other end is connected to the second one-way valve. The second one-way valve is connected to the second pinch valve. The pinch valve is connected, and one end of the second pinch valve that is not connected to the second one-way valve is connected to the long pipe in the long pipe group.

In some possible implementations of this application, the four-mix-one air inlet valve in the gas introduction module is connected to one end of the first bacterial filter through a silicone tube, and the other end of the four-mix one air inlet valve is connected to the air flow regulator and the nitrogen flow rate. regulator, carbon dioxide flow regulator, and oxygen flow regulator. The other ends of the air flow regulator, nitrogen flow regulator, carbon dioxide flow regulator, and oxygen flow regulator are connected to corresponding gas cylinders through tracheas;

The sample outlet of the automatic sampler in the sample processing module is connected to the detection chamber in the cell analyzer through a sample transport tube, and the sample taken out from the automatic sampler can be directly transported to the cell analyzer for observation and detection; The sampling port of the automatic sampler is connected to the sampling tube on the lid of the cell culture tank through a silicone tube; the reagent rack is installed in the device cabinet, and the reagent rack is used to place reagent bottles to facilitate the automatic feeding The sampler injects samples; the automatic sampler is connected to the sampling tube on the lid of the mixing tank through a silica gel tube, and new reagents are added into the mixing tank;

One end of the waste liquid pump in the waste liquid module is connected to the liquid waste pipe on the lid of the mixing tank through a silicone tube. The other end of the liquid waste pump that is not connected to the liquid waste pipe is connected to a silicone tube and the silicone tube is directly inserted into the waste liquid barrel.

In some possible implementations of this application, the sensor group includes a temperature sensor, a dissolved oxygen concentration sensor, a pH sensor and a carbon dioxide concentration sensor; and/or, the first bacterial filter, the second bacterial filter, The third bacterial filter is used to bidirectionally filter fine impurities and bacteria in the gas; and/or, the cell culture tank of the culture module and the mixing tank in the mixing module are both sealed pressure-resistant containers.

The invention has the following advantages:

(1) The present invention realizes cell culture that can reduce the cell loss rate during the cell-nutrient mixing process. The mixing of substances in existing culture equipment is mainly provided by stirring paddles and aeration disks, which inevitably causes shear stress damage to cells. If the shaking mechanism is simply used for mixing, as the culture system increases, the liquid surface contacts the air for oxygen transmission, and there is an obvious dissolved oxygen concentration gradient in the culture tank, which is not enough to support the dissolved oxygen demand of all cells in the tank. Therefore, the present invention separates the aeration of oxygen and other gases and the mixing of nutrients from cell culture, and sets up a separate mixing module. Traditional stirring and aeration are used to ensure that the nutrients in the culture medium are fully and evenly mixed. The cell culture module The gentle shaking of the shaker ensures the uniform dispersion of the cells, and then the circulation system replaces the mixed nutrient-rich medium in the mixing module with the nutrient-depleted medium in the cell culture module, thereby improving nutrient transmission. On the basis of the characteristics and enhanced mixing stability, the shear damage caused to cells by mixing power sources such as stirring and aeration is eliminated, achieving a low cell loss rate during the cell-nutrient mixing process.

(2) The present invention realizes scalable large-scale cell culture based on low cell loss rate. The existing methods to expand the culture scale of stem cells are mainly to increase the number of culture dishes or increase the culture volume of a single tank. However, in the former case, cells can only grow adherently, and the number is much lower than the number of cells attached to microcarriers and proliferating in the cell culture tank. The latter has the problem of uneven distribution of nutrients in the tank. If mixing, spraying and other mixed power sources are added, the damage of the shear force to the cells/cell-microcarriers will be increased, causing cell death and increasing the cost of the cell culture process. loss rate. The method of the present invention based on the invention point (1) adds multiple cell culture tanks to the culture module. Each culture tank can work independently and can be combined with the mixing module for cell culture. By selecting the number of cell culture tanks, Enable scalable large-scale cell culture.

(3) The present invention realizes the control of key environmental parameters of human internal environment simulation cultivation based on multi-physical field decoupling. Existing cell culture equipment and methods fail to solve the problem of simultaneously regulating pressure, pH, dissolved oxygen concentration and temperature in the culture environment; the present invention is based on high-sensitivity sensor feedback signals and uses a built-in control model to solve multiple physical fields of the cell culture environment. Coupled analysis, comprehensive control adjusts the cell culture temperature through the heater and temperature controller. After the temperature stabilizes, the ratio, rate and time of the four gases of air, oxygen, nitrogen, and carbon dioxide introduced into the cell culture tank are controlled. Pressure, pH and dissolved oxygen concentration in the cell growth environment; solves the problem that existing technology cannot simultaneously control environmental pressure, pH, dissolved oxygen concentration and temperature during in vitro culture, so that cells are always in a culture environment similar to that in the human body. Ensure the production quality of cells to the greatest extent.

(4) The present invention realizes automated cell culture based on the invention points (1), (2) and (3). At present, labor-intensive cell culture methods are used, such as manual addition of culture media, sampling, testing, and expansion of the culture system. Most of the operating steps are likely to cause contamination and produce large errors. Therefore, the equipment is replaced by automated operation without the need for a large number of experimenters. Complete the mass production of cells and improve the reproducibility and stability of the cell culture process to meet production needs.

Description of the drawings

Figure 1 shows a schematic structural diagram of a human-like large-scale stem cell automatic culture equipment with low loss rate provided by an exemplary embodiment of the present application.

Figure 2 shows a schematic structural diagram of the connection of various modules of a human-like large-scale stem cell automatic culture equipment with a low loss rate provided by an exemplary embodiment of the present application.

Figure 3 shows a schematic structural diagram of a culture module provided by an exemplary embodiment of the present application.

Figure 4 shows a schematic structural diagram of the connection between the culture module, the pressure adjustment module, and the sample processing module provided by an exemplary embodiment of the present application.

Figure 5 shows a schematic structural diagram of the connection of the culture module, circulation module, and mixing module provided by an exemplary embodiment of the present application.

Figure 6 shows a schematic structural diagram of the connection of the mixing module, gas introduction module, and waste liquid module provided by an exemplary embodiment of the present application.

Figure 7 illustrates a schematic structural diagram of a long tube and a cell screen provided by an exemplary embodiment of the present application.

Markings in the attached picture:

1. Control cabinet; 2. Culture module; 3. Mixing module; 4. Circulation module; 5. Device cabinet; 6. Sterile cover; 7. Pressure adjustment module; 8. Fixed table; 1.1. Central controller; 1.2. Multi-parameter transmitter; 2.1, cell culture tank; 2.2, culture table; 2.1.1, flexible semi-wrapped heater; 2.1.2, sensor group; 2.1.3, shaker; 2.1.4, cell screen; 2.1.5, long tube A; 2.1.6, short tube A; 2.1.7, sampling tube; 3.1, mixing tank; 3.2, flexible surround heater; 3.3, first stirring paddle; 3.4, first connecting rod; 3.5. The first stirring motor; 3.6. Aeration plate; 3.7. The first bacterial filter; 3.8. Long tube set; 3.8.1. Long tube clamp; 3.9. Short tube set; 3.9.1. Short tube clamp ; 3.10, sampling tube; 3.11, liquid discard tube; 3.12, second stirring paddle; 3.13, second connecting rod; 3.14 second stirring motor; 4.1, first peristaltic pump; 4.2, first one-way valve; 4.3, The first pinch valve; 4.4, the second peristaltic pump; 4.5, the second one-way valve; 4.6, the second pinch valve; 5.1, gas introduction module; 5.2, sample processing module; 5.3, waste liquid module; 5.1.1 , air flow regulator; 5.1.2, nitrogen flow regulator; 5.1.3, carbon dioxide flow regulator; 5.1.4, oxygen flow regulator; 5.1.5, four-mix one intake valve; 5.2.1, automatic sampling 5.2.2, cell analyzer; 5.2.3, automatic sampler; 5.5.4, reagent rack; 5.3.1, waste liquid bucket; 5.3.2, liquid discard pump; 7.1, first trachea; 7.2, Second air pipe; 7.3, second bacterial filter; 7.4, third bacterial filter; 7.5, pressure sensor; 7.6, pressure regulating proportional valve.

Detailed ways

Various exemplary embodiments, features, and aspects of the present application will be described in detail below with reference to the accompanying drawings. The same reference numbers in the drawings identify functionally identical or similar elements. Although various aspects of the embodiments are illustrated in the drawings, the drawings are not necessarily drawn to scale unless otherwise indicated. The word "exemplary" as used herein means "serving as an example, example, or illustrative." Any embodiment described herein as "exemplary" is not necessarily to be construed as superior or superior to other embodiments. In addition, in order to better explain the present application, those skilled in the art should understand that numerous specific details are given in each embodiment below. The present application may be practiced without certain specific details. In some embodiments, methods, means, and components that are well known to those skilled in the art are not described in detail in order to highlight the gist of the present application.

As shown in Figures 1 to 6, the embodiment of the present application is a human-like large-scale stem cell automatic culture equipment with a low loss rate, including a fixed table 8; a culture module 2 is installed on the fixed table 8; There is a sterile cover 6 for covering the culture module 2, and a sterilization device is provided inside the sterile cover 6; a control cabinet 1 is provided on one side of the fixed table 8; Mixing module 3 and device cabinet 5, of which the device cabinet 5 is installed on the left side of the mixing module 3 and is connected through air pipes, silicone tubes, data lines, communication lines, etc.; it also includes several circulation modules 4, shaking modules, and pressure adjustment modules 7; The circulation module 4 connects the mixing module 3 and the culture module 2, so that the rich culture medium in the mixing module 3 flows into the culture module 2, and the poor culture medium in the culture module 2 flows into the mixing module 3 ; The shaking module is connected to the culture module 2 and is used to drive the culture module 2 to shake; the pressure adjustment module 7 is connected to the culture module 2 and is used to adjust the air pressure in the culture module 2.

In some exemplary implementations of this embodiment, the circulation module 4 includes a first peristaltic pump 4.1, a second peristaltic pump 4.4, a first one-way valve 4.2, a second one-way valve 4.5, and a first pinch valve. 4.3. Second pinch valve 4.6; the circulation module 4 shown has two pairs of ports, and the first peristaltic pump 4.1, the first one-way valve 4.2, and the first pinch valve 4.3 are connected in series in one of them. Between pairs of ports, the second peristaltic pump 4.4, the second one-way valve 4.5, and the second pinch valve 4.6 are connected in series between another pair of ports, and one port of each pair of ports is connected to the other port. The culture module 2 is connected, and the other port of each pair of ports is connected to the mixing module 3.

In some exemplary implementations of this embodiment, the pressure regulating module 7 includes a first air pipe 7.1, a second air pipe 7.2, a second bacterial filter 7.3, a third bacterial filter 7.4, a pressure sensor 7.5, a pressure regulating Proportional valve 7.6; one of the ports of the second air pipe 7.2, the third bacterial filter 7.4, the pressure sensor 7.5, and the pressure regulating proportional valve 7.6 are connected in sequence, and the first air pipe 7.1, the third Two bacterial filters 7.3 and the other port of the pressure regulating proportional valve 7.6 are connected in sequence, and the first trachea 7.1 and the second trachea 7.2 are respectively connected to the culture module 2; the pressure regulating proportional valve 7.6 controls The ratio of exhaust and return air is to maintain a stable pressure in the cell culture tank.

In some exemplary implementations of this embodiment, the culture module 2 includes a culture platform 2.2 fixedly installed on the fixed platform 8 and several independently working cell cultures detachably installed inside the culture platform 2.2. Tank 2.1; the shaking module includes several shakers 2.1.3; the shakers 2.1.3 are all fixed at the bottom end of the culture table 2.2; the shaker 2.1.3 table can be detachably installed with flexible semi-wrapped heating 2.1.1; the culture tank of the culture module 2 can be detachably installed in the flexible semi-wrapped heater 2.1.1. Both the culture tank of the culture module 2 and the flexible semi-wrapped heater 2.1.1 can follow the shaking. The bed 2.1.3 moves when the table top moves; the function of the flexible semi-wrapped heater 2.1.1 is to keep the temperature in the cell culture tank constant.

In some exemplary implementations of this embodiment, a sensor group 2.1.2 and a cell screen 2.1.4 are provided inside the cell culture tank 2.1; a connecting tube is fixed on the lid of the cell culture tank 2.1, so One end of the connecting tube is located outside the cell culture tank 2.1, and the other end is located inside the cell culture tank 2.1. The connecting tube includes a long tube A2.1.5, a short tube A2.1.6 and a sampling tube 2.1.7. One end of the sampling tube 2.1.7 is located in the cell culture tank 2.1; the cell screen 2.1.4 is fixed on the periphery of the long tube A2.1.5 and is coaxially assembled with the long tube A2.1.5, with its bottom There is a gap between the bottom end of the long tube A2.1.5, and the cell screen is a barrel-shaped mesh structure, which is used to prevent the cells/cell microcarrier mixture in the cell culture tank from flowing out with the culture medium when the circulation system is turned on. ; The sensor group 2.1.2 is a non-contact measurement device, fixedly installed in an independent chamber in the cell culture tank 2.1.

In some exemplary implementations of this embodiment, the mixing module 3 includes a mixing tank 3.1, a flexible surround heater 3.2, a first stirring paddle 3.3, a second stirring paddle 3.12, a first connecting rod 3.4, a second Connecting rod 3.13, first stirring motor 3.5, second stirring motor 3.14, aeration plate 3.6, first bacterial filter 3.7, long tube set 3.8, short tube set 3.9, sampling tube 3.10, liquid discarding tube 3.11; described The first bacterial filter 3.7 is connected to the aeration disk 3.6;

In some exemplary implementations of this embodiment, the control cabinet 1 is equipped with a central controller 1.1 and a multi-parameter transmitter 1.2; the central controller 1.1 has functions such as signal acquisition, signal processing, and data storage; The sensor group 2.1.2 is connected to the multi-parameter transmitter 1.2 through signal lines and communication lines, can transmit information to the multi-parameter transmitter 1.2, and accept the regulation of the multi-parameter transmitter 1.2;

In some exemplary implementations of this embodiment, a gas introduction module 5.1, a sample processing module 5.2 and a waste liquid module 5.3 are installed in the device cabinet 5; wherein the gas introduction module 5.1 includes an air flow regulator 5.1. 1. Nitrogen flow regulator 5.1.2, carbon dioxide flow regulator 5.1.3, oxygen flow regulator 5.1.4, four-mix one intake valve 5.1.5; the air flow regulator 5.1.1, the nitrogen flow rate One end of the regulator 5.1.2, the carbon dioxide flow regulator 5.1.3, and the oxygen flow regulator 5.1.4 are respectively connected to one end of the four-mix-one intake valve 5.1.5. The other end of the air valve 5.1.5 is connected to the first bacterial filter 3.7; the four gases are pre-mixed through the four-mix-one air inlet valve 5.1.5, and then filtered out impurities through the first bacterial filter 3.7 , and finally pass into the mixing tank 3.1 through the aeration disk 3.6;

In some exemplary implementations of this embodiment, the sample processing module 5.2 includes an automatic sampler 5.2.1, a cell analyzer 5.2.2, an automatic sampler 5.2.3, and a reagent rack 5.2.4; The cell analyzer 5.2.2 is connected to the automatic sampler 5.2.1, and the automatic sampler 5.2.1 is connected to the sampling tube 2.1.7; the reagent rack 5.2.4 is connected to the automatic sampler 5.2 .3 connection, the automatic sampler 5.2.3 is connected to the sampling tube 3.10;

In some exemplary implementations of this embodiment, the waste liquid module 5.3 includes a waste liquid barrel 5.3.1 and a liquid discard pump 5.3.2; the waste liquid barrel 5.3.1 and the liquid discard pump 5.3. 2 is connected, and the liquid discarding pump 5.3.2 is connected with the liquid discarding pipe 3.11.

In some exemplary implementations of this embodiment, the flexible surrounding heater 3.2 is fixed around the periphery of the mixing tank 3.1 to keep the temperature in the mixing tank 3.1 constant; the lid of the mixing tank 3.1 is fixed with Long tube clamp 3.8.1 and short tube clamp 3.9.1; several long tubes can be detachably installed in the long tube clamp 3.8.1 to form a long tube group 3.8, which is used to connect the circulation module 4 to culture The base is led out of the mixing tank 3.1, and several short pipes can be detachably installed in the short pipe clamp 3.9.1 to form a short pipe group 3.9, which is used to connect the circulation module 4 and introduce the culture medium into the mixing tank 3.1; The first stirring motor 3.5 is detachably installed on the left side of the lid of the mixing tank 3.1. One end of the first connecting rod 3.4 is connected to the power output end of the first stirring motor 3.5, and the other end is connected to the first stirring paddle 3.3. The first stirring paddle 3.3 is located in the mixing tank 3.1 close to the left side of the aeration plate 3.6; a second stirring motor 3.14 is detachably installed on the right side of the lid of the mixing tank 3.1, and one end of the second connecting rod 3.13 It is connected to the power output end of the second stirring motor 3.14, and the other end is connected to the second stirring paddle 3.12. The second stirring paddle 3.12 is located in the mixing tank 3.1 near the right side of the aeration plate 3.6; the aeration plate 3.6 Located in the mixing tank 3.1 near the bottom of the mixing tank 3.1, one end is connected to the first bacterial filter 3.7 through a silicone tube; the function of the first stirring paddle 3.3 and the second stirring paddle 3.12 is to mix the mixture through high-speed rotation. Quickly mix everything in the jar.

In some exemplary implementations of this embodiment, the circulation module 4 includes a first circulation unit and a second circulation unit, and one end of the first circulation unit is connected to the long tube A2 in the cell culture tank 2.1. 1.5 is connected, and the other end is connected to the short pipe in the short pipe group 3.9 in the mixing tank 3.1; one end of the second circulation unit is connected to the long pipe in the long pipe group 3.8 in the mixing tank 3.1, and the other end is connected to The short tube A2.1.6 in the cell culture tank 2.1 is connected; the first circulation unit includes a first peristaltic pump 4.1, a first one-way valve 4.2, and a first pinch valve 4.3. The first peristaltic pump 4.1 One end is connected to the long pipe A2.1.5, and the other end is connected to the first one-way valve 4.2. The first one-way valve 4.2 is connected to the first pinch valve 4.3. The first pinch valve 4.3 is not connected to the first one-way valve 4.3. One end connected to the valve 4.2 is connected to the short tube group 3.9; the second circulation unit includes a second peristaltic pump 4.4, a second one-way valve 4.5, and a second pinch valve 4.6. The second peristaltic pump One end of 4.4 is connected to the short pipe A2.1.6, and the other end is connected to the second one-way valve 4.5. The second one-way valve 4.5 is connected to the second pinch valve 4.6, and the second pinch valve 4.6 is not connected to the second pinch valve 4.6. One end of the two one-way valves 4.5 is connected to the long pipe in the long pipe group 3.8.

In some exemplary implementations of this embodiment, the four-mixing-inlet valve 5.1.5 in the gas introduction module 5.1 is connected to one end of the first bacterial filter 3.7 through a silicone tube, and the four-mixing-inlet valve 5.1.5 The other end is connected to the air flow regulator 5.1.1, the nitrogen flow regulator 5.1.2, the carbon dioxide flow regulator 5.1.3, and the oxygen flow regulator 5.1.4. The air flow regulator 5.1.1 and the nitrogen flow regulator 5.1 .2. The other end of the carbon dioxide flow regulator 5.1.3 and the oxygen flow regulator 5.1.4 is connected to the corresponding gas cylinder through the trachea;

In some exemplary implementations of this embodiment, the sample outlet of the automatic sampler 5.2.1 in the sample processing module 5.2 is connected to the detection chamber in the cell analyzer 5.2.2 through a sample delivery tube, and the automatic sampler 5.2.1 can be The sample taken out from the sampler 5.2.1 is directly transported to the cell analyzer 5.2.2 for observation and detection; the sampling port of the automatic sampler 5.2.1 is connected to the sampling port on the cover of the cell culture tank 2.1 through a silicone tube. tube 2.1.7; the reagent rack 5.2.4 is installed in the device cabinet 5, and the reagent rack 5.2.4 is used to place reagent bottles to facilitate the injection of samples by the automatic sampler 5.2.3; the The automatic sampler 5.2.3 is connected to the sampling tube 3.10 on the tank cover of the mixing tank 3.1 through a silicone tube, and new reagents are added into the mixing tank 3.1;

In some exemplary implementations of this embodiment, one end of the waste liquid pump 5.3.2 in the waste liquid module 5.3 is connected to the liquid waste pipe 3.11 on the tank cover of the mixing tank 3.1 through a silicone tube, and the liquid waste pump 5.3.2 is not The other end connected to the waste pipe 3.11 is connected with a silicone tube and the silicone tube is directly inserted into the inside of the waste barrel 5.3.1.

In some exemplary implementations of this embodiment, the sensor group 2.1.2 includes a temperature sensor, a dissolved oxygen concentration sensor, a pH sensor and a carbon dioxide concentration sensor; and/or, the first bacterial filter 3.7, The second bacterial filter 7.3 and the third bacterial filter 7.4 are used to bidirectionally filter fine impurities and bacteria in the gas; and/or, the cell culture tank 2.1 of the culture module 2 and the mixing tank 3.1 in the mixing module 3 are both Sealed pressure-resistant container.

When the above-mentioned human-like large-scale stem cell automatic culture equipment with a low loss rate is used, the culture medium, oxygen and other gases and nutrients are evenly mixed in the mixing module 3. After the mixing is completed, the cells enter the cell culture tank 2.1 through the circulation module 4. /The cell-microcarrier complex is directly in contact with the mixed culture medium in the cell culture tank 2.1. Therefore, there is no need to install modules with functions such as stirring and aeration in the cell culture tank 2.1, which avoids the impact of stirring blades and aeration on the cells. Causes shear stress damage, significantly reduces the cell loss rate in the cell culture process, increases the density of cell culture on the original basis, and increases the number of cultured cells.

In some exemplary implementations of this embodiment, the central controller 1.1 controls the rate, time and proportion of air, carbon dioxide, oxygen and nitrogen introduced into the mixing tank 3.1 through the gas introduction module 5.1; The central controller 1.1 controls the pressure in the cell culture tank 2.1 through the pressure regulating proportional valve 7.6; the central controller 1.1 controls the flexible semi-wrapped heater 2.1 through the feedback information of the multi-parameter transmitter 1.2. 1 and the heating temperature of the flexible surround heater 3.2 to stabilize the internal temperature of the culture module 2 and the mixing module 3; the central controller 1.1 determines the location of the cells based on the information obtained by the cell analyzer 5.2.2 in the sample processing module 5.2 At the culture stage, according to the environmental information required by the cells at this stage in the built-in model, adjust the rotation speed of the two stirring paddles in the mixing tank 3.1, adjust the air flow regulator 5.1.1 and the nitrogen flow regulator in the gas introduction module 5.1 5.1.2, the opening and access time of the carbon dioxide flow regulator 5.1.3, and the oxygen flow regulator 5.1.4; when the central controller 1.1 determines that the cells are in the designated culture stage, it controls the automatic sampler 5.2.3 to transfer the reagents The cytokines in the reagent bottles in rack 5.2.4 are added to the mixing tank 3.1 and added to the cell culture tank 2.1 through the circulation system 4 to meet the needs of cell growth.

In some exemplary implementations of this embodiment, the pH, dissolved oxygen concentration and pressure control of the culture environment mainly rely on the central controller 1.1 to control the pressure adjustment module 7 and the gas introduction module 5.1; first, the target setting is The fixed values are brought into the built-in model to calculate the mass proportion coefficients of carbon dioxide, oxygen, nitrogen and air that need to be introduced into the cell culture tank 2.1. The central controller 1.1 starts the gas introduction module 5.1 and starts to continuously introduce the mixed gas. At the same time, the central controller 1.1 Start the pressure adjustment module 7 to start stably adjusting the pressure in the cell culture tank 2.1; when the sensor group detects a numerical change in pH and/or dissolved oxygen concentration, the central controller 1.1 calculates the four gases that need to be introduced according to the model Proportion, control the flow regulator of the four gases in the gas introduction module 5.1, change the proportion of the four mixed gases, and then continue to pass the mixed gas into the cell culture tank 2.1 until the sensor detection value returns to the set range; in the gas During the continuous air intake process of the introduction module 5.1, the pressure regulating module 7 needs to adjust the opening of the air inlet port, air return port and exhaust port of the pressure regulating proportional valve 7.6 at any time according to the detection value fed back by the pressure sensor 7.5 to make the cell culture tank 2.1 The pressure inside remains stable.

In this embodiment, the application of a low-loss human-like large-scale stem cell automatic culture equipment is as follows:

Step ① Start the equipment, the central controller 1.1 starts the semi-wrapped heater 2.1.1 and the flexible surround heater 3.2, and adjusts the temperature in each cell culture tank 2.1 and mixing tank 3.1 to be constant at 37.2°C;

Step ② The central controller 1.1 controls the automatic sampler 5.2.3 to add the fresh culture medium on the reagent rack 5.2.4 into the mixing tank 3.1;

Step ③ The central controller 1.1 starts the gas introduction module 5.1 and controls the air flow regulator 5.1.1, nitrogen flow regulator 5.1.2, carbon dioxide flow regulator 5.1.3, and oxygen flow regulator 5.1.4 according to the environmental indicators input by the user. opening and access time;

Step ④ Set the rotation speed, and the central controller 1.1 starts the first stirring motor 3.5 and the second stirring motor 3.14 to drive the first stirring paddle 3.3 and the second stirring paddle 3.12 to rotate at high speed;

Step ⑤ The central controller 1.1 starts the pressure adjustment module 77. The central controller 1.1 controls the discharge volume of the mixed gas in the pressure regulating proportional valve 7.6 through the feedback value of the pressure sensor 7.5 to adjust the pressure value and change period in the cell culture tank 2.1;

Step ⑥ Add cells/cell-microcarrier complex in the middle area between the inner wall of the cell culture tank 2.1 and the cell screen 2.1.4. The central controller 1.1 starts the shaker 2.1.3 and inputs the set speed to make the cells/cell-microcarriers in the area. uniformly distributed;

Step ⑦ The central controller 1.1 starts the circulation module 4 to circulate the medium from the mixing module 3 to the cell culture tank 2.1 in the culture module 2. The central controller 1.1 regulates the first peristaltic pump 4.1 and the second peristaltic pump 4.1 in the circulation module 4.1. The rotation speed of the peristaltic pump 4.4 controls the exchange speed of the culture medium in the cell culture tank 2.1 in the mixing module 3 and the culture module 2;

Step ⑧Set the sampling frequency and sampling volume. The central controller 1.1 controls the automatic sampler 5.2.1 to collect the cells and surrounding culture medium in the cell culture tank 2.1, and send the collected samples to cell analysis through the connected sample delivery tube. Instrument 5.2.2, cell analyzer 5.2.2 detects cell morphology, viable cell number/proportion, glucose/urea/lactic acid/inorganic salt content;

Step 9. The central controller 1.1 determines the culture stage of the cells based on the information obtained by the cell analyzer 5.2.2, and adjusts the two stirring paddles in the mixing tank 3.1 based on the environmental information required by the cells at that stage in the preset program. Rotation speed, adjust the opening and access time of the air flow regulator 5.1.1, nitrogen flow regulator 5.1.2, carbon dioxide flow regulator 5.1.3, and oxygen flow regulator 5.1.4 in the gas introduction module 5.1; in the central control When the device 1.1 determines that the cells are in the designated culture stage, the automatic sampler 5.2.3 is controlled to add the cytokines in the reagent bottle in the reagent rack 5.2.4 to the mixing tank 3.1, and then adds it to the cell culture tank 2.1 through the circulation system 4 for cell growth. need;

Step ⑩ During the entire cultivation process, the sensor group 2.1.2 monitors the temperature, dissolved oxygen concentration, pH value and carbon dioxide concentration in the cell culture tank 2.1 in real time, and transmits the signals to the multi-parameter transmitter 1.2. The multi-parameter transmitter 1.2 then collects the The information is transmitted to the central controller 1.1 for viewing, processing and storage; the pressure sensor 7.5 monitors the pressure in the cell culture tank 2.1 in real time during the entire culture process and transmits the information to the central controller 1.1 for viewing, processing and storage; The pH, dissolved oxygen concentration and pressure control of the culture environment rely on the central controller 1.1 to regulate the pressure adjustment module 7 and the gas introduction module 5.1; after the cell culture is completed, the central controller 1.1 starts the waste liquid in the waste liquid module 5.3 Pump 5.3.2, pump the culture medium from the mixing tank 3.1 through the waste pipe 3.11 to the waste barrel 5.3.1, and discard it after treatment.

The embodiments of the present application have been described above. The above description is illustrative, not exhaustive, and is not limited to the disclosed embodiments. Many modifications and variations will be apparent to those skilled in the art without departing from the scope and spirit of the described embodiments. The terminology used herein is chosen to best explain the principles, practical applications, or improvements to the technology in the market, or to enable other persons of ordinary skill in the art to understand the embodiments disclosed herein.

Claims (10)

1. The automatic human-simulated large-scale stem cell culture equipment with low loss rate comprises a fixed table (8), wherein a culture module (2) is arranged on the fixed table (8); the fixed table (8) is provided with a sterile cover (6) for covering the culture module (2), and a sterilizing device is arranged in the sterile cover (6); a control cabinet (1) is arranged on one side of the fixed table (8); the mixing module (3) and the device cabinet (5) are fixedly arranged in the fixed table (8), wherein the device cabinet (5) is arranged at the left side of the mixing module (3) and is connected with the mixing module through an air pipe, a silicone tube, a data line, a communication line and the like; it is characterized in that the method comprises the steps of,

the device also comprises a plurality of circulation modules (4), a shaking module and a pressure regulating module (7);

the circulation module (4) is connected with the mixing module (3) and the culture module (2), so that the culture medium rich in the mixing module (3) flows into the culture module (2), and the culture medium lean in the culture module (2) flows into the mixing module (3);

the shaking module is connected with the culture module (2) and is used for driving the culture module (2) to shake;

the pressure regulating module (7) is connected with the culture module (2) and is used for regulating the air pressure in the culture module (2).

2. The low-loss human-like large-scale stem cell automatic culture apparatus according to claim 1, wherein,

The circulation module (4) comprises a first peristaltic pump (4.1), a first one-way valve (4.2), a first pinch valve (4.3), a second peristaltic pump (4.4), a second one-way valve (4.5) and a second pinch valve (4.6);

the circulation module (4) is provided with two pairs of ports, the first peristaltic pump (4.1), the first one-way valve (4.2) and the first pinch valve (4.3) are sequentially connected in series between one pair of ports, the second peristaltic pump (4.4), the second one-way valve (4.5) and the second pinch valve (4.6) are sequentially connected in series between the other pair of ports, one port of each pair of ports is connected with the culture module (2), and the other port of each pair of ports is connected with the mixing module (3).

3. The low-loss rate human-like large-scale stem cell automatic culture apparatus according to claim 2,

the pressure regulating module (7) comprises a first air pipe (7.1), a second air pipe (7.2), a second bacterial filter (7.3), a third bacterial filter (7.4), a pressure sensor (7.5) and a pressure regulating proportional valve (7.6);

the utility model discloses a pressure regulating device, including first trachea (7.1), second trachea (7.3), pressure regulating proportional valve (7.6), second trachea (7.2) third bacterial filter (7.4) pressure sensor (7.5) one of them port of pressure regulating proportional valve (7.6) connects gradually, first trachea (7.1) second bacterial filter (7.3) another port of pressure regulating proportional valve (7.6) connects gradually, first trachea (7.1) and second trachea (7.2) respectively with cultivate module (2) and connect.

4. The automatic human-like large-scale stem cell culturing apparatus with low loss rate according to claim 3,

the culture module (2) comprises a culture table (2.2) fixedly arranged on a fixed table (8) and a plurality of independently-operable cell culture tanks (2.1) detachably arranged inside the culture table (2.2);

the shaking module comprises a plurality of shaking tables (2.1.3); the shaking tables (2.1.3) are all fixed at the bottom end inside the culture table (2.2); a flexible semi-wrapped heater (2.1.1) is detachably arranged on the table top of the cradle (2.1.3); the culture tank of the culture module (2) is detachably arranged in the flexible semi-wrapped heater (2.1.1), and the culture tank of the culture module (2) and the flexible semi-wrapped heater (2.1.1) can move along with the table-board movement of the shaking table (2.1.3).

5. The low-loss human-like large-scale stem cell automatic culture apparatus according to claim 4, wherein,

a sensor group (2.1.2) and a cell screen (2.1.4) are arranged in the cell culture tank (2.1); a communicating pipe is fixed on the tank cover of the cell culture tank (2.1), one end of the communicating pipe is positioned outside the cell culture tank (2.1), the other end of the communicating pipe is positioned in the cell culture tank (2.1), the communicating pipe comprises a long pipe A (2.1.5), a short pipe A (2.1.6) and a sampling pipe (2.1.7), and one end of the sampling pipe (2.1.7) is positioned in the cell culture tank (2.1); the cell screen (2.1.4) is fixed on the periphery of the long tube A (2.1.5), is coaxially assembled with the long tube A (2.1.5), and the bottom of the cell screen is spaced from the bottom end of the long tube A (2.1.5); the sensor group (2.1.2) is a non-contact measuring device and is fixedly arranged in an independent small chamber in the cell culture tank (2.1).

6. The apparatus for automatically culturing human-like large-scale stem cells with low loss rate according to claim 5,

the mixing module (3) comprises a mixing tank (3.1), a flexible surrounding type heater (3.2), a first stirring paddle (3.3), a second stirring paddle (3.12), a first connecting rod (3.4), a second connecting rod (3.13), a first stirring motor (3.5), a second stirring motor (3.14), an aeration disc (3.6), a first bacterial filter (3.7), a long tube group (3.8), a short tube group (3.9), a sample injection tube (3.10) and a liquid discarding tube (3.11);

the first bacterial filter (3.7) is connected with the aeration disc (3.6);

a central controller (1.1) and a multi-parameter transmitter (1.2) are arranged in the control cabinet (1); the sensor group (2.1.2) is connected with the multi-parameter transmitter (1.2) through a signal line and a communication line, can transmit information to the multi-parameter transmitter (1.2) and receives the regulation and control of the multi-parameter transmitter (1.2);

a gas introduction module (5.1), a sample treatment module (5.2) and a waste liquid module (5.3) are arranged in the device cabinet (5); wherein the gas introducing module (5.1) comprises an air flow regulator (5.1.1), a nitrogen flow regulator (5.1.2), a carbon dioxide flow regulator (5.1.3), an oxygen flow regulator (5.1.4) and a four-mixing air inlet valve (5.1.5);

One end of the air flow regulator (5.1.1), the nitrogen flow regulator (5.1.2), the carbon dioxide flow regulator (5.1.3) and the oxygen flow regulator (5.1.4) are respectively connected with one end of the four-mixing air inlet valve (5.1.5), and the other end of the four-mixing air inlet valve (5.1.5) is connected with the first bacterial filter (3.7);

the sample processing module (5.2) comprises an automatic sampler (5.2.1), a cell analyzer (5.2.2), an automatic sampler (5.2.3) and a reagent rack (5.2.4);

the cell analyzer (5.2.2) is connected with the automatic sampler (5.2.1), and the automatic sampler (5.2.1) is connected with the sampling tube (2.1.7); the reagent rack (5.2.4) is connected with the automatic sampler (5.2.3), and the automatic sampler (5.2.3) is connected with the sampling tube (3.10);

the waste liquid module (5.3) comprises a waste liquid barrel (5.3.1) and a waste liquid pump (5.3.2);

the waste liquid barrel (5.3.1) is connected with the liquid discarding pump (5.3.2), and the liquid discarding pump (5.3.2) is connected with the liquid discarding pipe (3.11).

7. The apparatus for automatically culturing human-like large-scale stem cells with low loss rate according to claim 6,

the flexible surrounding type heater (3.2) is fixed around the periphery of the mixing tank (3.1) so as to keep the temperature in the mixing tank (3.1) constant; a long pipe clamp (3.8.1) and a short pipe clamp (3.9.1) are fixed on the tank cover of the mixing tank (3.1); a plurality of long pipes are detachably arranged in the long pipe clamp (3.8.1) to form a long pipe group (3.8) together, the long pipe clamp is used for connecting the circulation module (4) to guide a culture medium out of the mixing tank (3.1), a plurality of short pipes are detachably arranged in the short pipe clamp (3.9.1) to form a short pipe group (3.9) together, and the short pipe clamp is used for connecting the circulation module (4) to guide the culture medium into the mixing tank (3.1); a first stirring motor (3.5) is detachably arranged on the left side of the top of the mixing tank (3.1), one end of a first connecting rod (3.4) is connected with the power output end of the first stirring motor (3.5), the other end of the first connecting rod is connected with a first stirring paddle (3.3), and the first stirring paddle (3.3) is positioned at a position, close to the left side of the aeration disc (3.6), in the mixing tank (3.1); a second stirring motor (3.14) is detachably arranged on the right side of the top of the mixing tank (3.1), one end of a second connecting rod (3.13) is connected with the power output end of the second stirring motor (3.14), the other end of the second connecting rod is connected with a second stirring paddle (3.12), and the second stirring paddle (3.12) is positioned in the mixing tank (3.1) and is close to the right side of the aeration disc (3.6); the aeration disc (3.6) is positioned in the mixing tank (3.1) and is close to the bottom of the mixing tank (3.1), and one end of the aeration disc is connected with one end of the first bacterial filter (3.7) through a silica gel tube.

8. The apparatus for automatically culturing human-like large-scale stem cells with low loss rate according to claim 7,

the circulation module (4) comprises a first circulation unit and a second circulation unit, one end of the first circulation unit is connected with a long pipe A (2.1.5) in the cell culture tank (2.1), and the other end of the first circulation unit is connected with short pipes in a short pipe group (3.9) in the mixing tank (3.1); one end of the second circulation unit is connected with a long pipe in a long pipe group (3.8) in the mixing tank (3.1), and the other end of the second circulation unit is connected with a short pipe A (2.1.6) in the cell culture tank (2.1); the first circulation unit comprises a first peristaltic pump (4.1) and a first check valve (4.2) and a first pinch valve (4.3), one end of the first peristaltic pump (4.1) is connected with a long pipe A (2.1.5), the other end of the first peristaltic pump is communicated with the first check valve (4.2), the first check valve (4.2) is connected with the first pinch valve (4.3), and one end of the first pinch valve (4.3) which is not connected with the first check valve (4.2) is connected with the short pipe group (3.9); the second circulation unit comprises a second peristaltic pump (4.4), a second check valve (4.5) and a second pinch valve (4.6), one end of the second peristaltic pump (4.4) is connected with a short pipe A (2.1.6), the other end of the second peristaltic pump is communicated with the second check valve (4.5), the second check valve (4.5) is connected with the second pinch valve (4.6), and one end of the second pinch valve (4.6) which is not connected with the second check valve (4.5) is connected with a long pipe in the long pipe group (3.8).

9. The low-loss human-like large-scale stem cell automatic culture apparatus according to claim 8, wherein,

a four-mixing air inlet valve (5.1.5) in the gas introducing module (5.1) is connected with one end of a first bacterial filter (3.7) through a silica gel pipe, the other end of the four-mixing air inlet valve (5.1.5) is connected with an air flow regulator (5.1.1), a nitrogen flow regulator (5.1.2), a carbon dioxide flow regulator (5.1.3) and an oxygen flow regulator (5.1.4), and the other ends of the air flow regulator (5.1.1), the nitrogen flow regulator (5.1.2), the carbon dioxide flow regulator (5.1.3) and the oxygen flow regulator (5.1.4) are connected with corresponding gas cylinders through air pipes;

the sample outlet of the automatic sampler (5.2.1) in the sample processing module (5.2) is connected with a detection cell in the cell analyzer (5.2.2) through a sample conveying pipe, so that the sample taken out of the automatic sampler (5.2.1) can be directly conveyed into the cell analyzer (5.2.2) for observation and detection; the sampling port of the automatic sampler (5.2.1) is connected with a sampling tube (2.1.7) on the tank cover of the cell culture tank (2.1) through a silica gel tube; the reagent rack (5.2.4) is arranged in the device cabinet (5), and the reagent rack (5.2.4) is used for placing reagent bottles so as to facilitate sample injection of the automatic sample injector (5.2.3); the automatic sampler (5.2.3) is connected with a sampling pipe (3.10) on the tank cover of the mixing tank (3.1) through a silica gel pipe, and new reagent is added into the mixing tank (3.1);

One end of a liquid discarding pump (5.3.2) in the waste liquid module (5.3) is connected with a liquid discarding pipe (3.11) on a tank cover of the mixing tank (3.1) through a silica gel pipe, and the other end of the liquid discarding pump (5.3.2) which is not connected with the liquid discarding pipe (3.11) is connected with the silica gel pipe and the silica gel pipe is directly inserted into the waste liquid barrel (5.3.1).

10. The low-loss human-like large-scale stem cell automatic culture apparatus according to claim 9,

the sensor group (2.1.2) comprises a temperature sensor, a dissolved oxygen concentration sensor, a pH sensor and a carbon dioxide concentration sensor; and/or the number of the groups of groups,

the first bacterial filter (3.7), the second bacterial filter (7.3) and the third bacterial filter (7.4) are used for bidirectionally filtering fine impurities and bacteria in the gas; and/or the number of the groups of groups,

the cell culture tank (2.1) of the culture module (2) and the mixing tank (3.1) of the mixing module (3) are both closed pressure-resistant containers.