CN116286350A - Multi-gas incubator and control method thereof - Google Patents

Multi-gas incubator and control method thereof Download PDF

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CN116286350A
CN116286350A CN202310236610.0A CN202310236610A CN116286350A CN 116286350 A CN116286350 A CN 116286350A CN 202310236610 A CN202310236610 A CN 202310236610A CN 116286350 A CN116286350 A CN 116286350A
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concentration
temperature
box body
gas
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毒岛弘树
赵光宇
姜森
李璇
徐桂杰
刘锦阳
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Bingshan Songyang Biotechnology Dalian Co ltd
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Abstract

The invention provides a multi-gas incubator and a control method thereof. The structure of incubator includes a box body, a structure with accommodation space for holding cell culture container, a front door at the front end, and CO on the side wall of the box body 2 Concentration sensor and O 2 A concentration sensor and a fan; at least one group of supporting plates are arranged in the box body and used for placing the cell culture container; a humidifying tray arranged at the bottom of the box body and heated by a heater; a multi-gas channel communicated with the box body for selectively introducing CO 2 、O 2 、N 2 And air, enter the culture environment in the box bodyAnd a HEPA filter is arranged at one end part of the multi-air channel connected with the box body. The invention also discloses a control method of the incubator, which comprises the steps of temperature and CO 2 Concentration, O 2 And the sensor feeds back in real time, and the parameters in the control box reach the culture conditions to finish the culture. Effectively solves the problems of mixed bacterial pollution caused by dew condensation and the like when the temperature is reduced, the culture environment is regulated and controlled.

Description

Multi-gas incubator and control method thereof
Technical Field
The invention relates to the field of cell foundation research and cell treatment, in particular to the technical field of foundation and clinical medicine of IPS research, and particularly relates to a multi-gas incubator and a control method thereof.
Background
In the prior art, the cell culture device is easier to realize CO 2 Concentration, O 2 The concentration is controlled to realize the program setting change of the temperature in the culture device along with the time. However, depending on the characteristics of the cell culture apparatus, particularly the cultured IPS cells, different control conditions, and different target cells for cell differentiation, environmental control is particularly important.
The current cell culture device mostly adopts a cell culture bag to culture by using a liquid environment, and the factors of the environment are not much considered, but when the cell culture device is used for culturing cells by adopting an incubator, the influence of each condition on the cells needs to be balanced, nearly saturated humidity conditions (about 90-98% RH) are needed in the cell culture device, when the temperature in the device is reduced to control the culture conditions during the cell culture, the temperature in a container is lower than the dew point temperature, a large amount of dew condensation is generated on the wall surface in the incubator, the mixed bacteria pollution is generated, the final commodity qualification is difficult to realize, the culture differentiation efficiency is reduced, and the reject ratio is higher.
Therefore, it is necessary to provide a culture apparatus for solving the above-mentioned problems.
Disclosure of Invention
According to the above proposalIn the cell differentiation process, when the culture environment is required to be cooled and regulated, the existing incubator can have the technical problems of dew condensation, mixed bacterial pollution and the like, and the multi-gas incubator and the control method thereof are provided. The invention mainly utilizes the communication of a plurality of air pipelines on the incubator, and feeds back the culture environment in the incubator in real time through the sensor arranged in the incubator to CO in the incubator 2 、O 2 、N 2 Concentration real-time control, especially when the incubator is cooled down and regulated, through gaseous circulation control, effectively avoid the production of box inner wall dewfall to realize the effect of high-efficient regulation and control.
The invention adopts the following technical means:
a multi-gas incubator comprising:
the box body is used for accommodating a structure of the cell culture container, the front end of the box body is provided with a front door, and the side wall of the box body is also provided with CO 2 Concentration sensor and O 2 A concentration sensor and a fan;
at least one group of supporting plates, which are arranged in the box body and used for placing the cell culture container;
a humidifying tray arranged at the bottom of the box body and heated by a heater arranged in the wall surface of the inner box at the bottom of the box body;
a multi-gas channel communicated with the box body for selectively introducing CO 2 、O 2 、N 2 And air is used for adjusting the culture environment in the box body, and a HEPA filter is arranged at one side end part of the multi-air channel connected with the box body.
Further, the multi-gas channels are respectively connected with CO 2 Channel, O 2 Channel, N 2 The channels are connected with the air channels, the flow speed of each channel is controlled by various electromagnetic valves, and one or more channels are selectively selected and simultaneously enter the box body through the multi-air channels.
The invention also discloses a control method of the multi-gas incubator, which is characterized in that initial parameter conditions, first target parameter conditions and second target parameter conditions in the multi-gas incubator are respectively set through a temperature sensor and CO 2 Concentration sensor, O 2 Sensor entityAnd (3) feeding back the time, controlling the initial parameter condition to reach the first target parameter condition, and then feeding back the first target parameter condition to reach the second target parameter condition through the sensor to complete the culture.
Further, the initial parameter conditions and the first and second target parameter conditions include, but are not limited to, temperature, CO 2 Concentration, O 2 Concentration, humidity and incubation time.
Further, controlling the initial parameter condition to reach the first target parameter condition includes the steps of:
s11, a first target temperature set in the first target parameters is larger than the initial temperature in the box, and the output of the heater is controlled through PID regulation, so that the temperature is stabilized at the set first target temperature;
s12, setting a first target CO in the first target parameters 2 Defining a first target CO 2 The concentration set point is CO 2 SV1, initial CO 2 Concentration of CO 2 PV1, when CO 2 SV1>CO 2 At PV1, CO 2 Continuously switching on by electromagnetic valve and charging CO 2 Gas until the concentration reaches CO 2 SV1；
S13, setting a first target O in the first target parameters 2 Concentration, defining a first target O 2 The concentration set value is O 2 SV1, initial O 2 Concentration of O 2 PV1, when O 2 PV1>O 2 SV1，N 2 Continuously switching on by electromagnetic valve, and charging N 2 Gas until the concentration reaches O 2 SV1。
Further, controlling the first target parameter condition to reach the second target parameter condition includes the steps of:
s21, stopping the output of the heater when the second target temperature set in the second target parameter is smaller than the first target temperature in the box, simultaneously switching on the air pump, introducing air, reducing the temperature and humidity in the box, stopping the air pump until the actual temperature in the box is equal to the second target temperature, then controlling the heater to work through PID, and maintaining the temperature in the box stable at the second target temperature;
s22, second target parameterA second target CO set in (2) 2 Concentration is greater than actual CO in the tank 2 Concentration, definition of the second target CO 2 The concentration set point is CO 2 SV2, actual CO 2 Concentration of CO 2 PV2, when CO 2 SV2>CO 2 At the time of PV2, CO 2 Continuously switching on by electromagnetic valve and charging CO 2 A gas; until the concentration reaches CO 2 SV2；
S23, setting a second target O in the second target parameters 2 Concentration, definition of the second target O 2 Setting value O 2 SV2, actual O 2 Concentration of O 2 PV2, when O 2 SV2>O 2 At PV2, O 2 Continuously switching on by electromagnetic valve and charging O 2 Gas, finally reach O 2 SV2; when O is 2 SV2<O 2 At PV2, N 2 Continuously switching on by electromagnetic valve, and charging N 2 Gas, finally reach O 2 SV2；
And S24, when the incubation time set by the second target parameter is reached, the incubator gives an alarm to prompt the user to take the incubated sample.
Further, in the step S21, when the second target temperature is stable, the air pump stops working again, and the following gas control is performed, so that excessive gas waste is avoided for regulation.
Compared with the prior art, the multi-gas incubator provided by the invention is a cell culture device, and mainly aims to change the temperature and the concentration of each gas along with the time change through a program control function when the IPS cells are differentiated into target cells.
With the temperature change in the incubator, particularly when the temperature is reduced, a large amount of dew condensation is generated in the container in the prior art, so that the mixed bacteria pollution is caused. Under the condition of temperature reduction, the air pump is operated, and the humidity in the incubator is forcedly reduced by introducing air into the incubator. Then pass through N 2 Using electromagnetic valve to make N 2 Introducing into an incubator, and introducing O into the incubator 2 The concentration is adjusted to the set concentration. At the same time by turning on CO 2 Using electromagnetic valve to make CO 2 Introducing gas into incubator to introduce CO into incubator 2 The concentration is adjusted to the set concentration.
The invention changes the temperature and CO in the cell culture of cell culture research and clinical medicine 2 Concentration, O 2 The program functions of concentration and humidity can change the cell culture environment. Meanwhile, the mixed bacteria pollution caused by dew condensation can be prevented, a stable and controllable culture environment is provided for culturing IPS cells, an optimal cell culture environment is realized, and the incubator has the advantages of reasonable design, cost reduction, easiness in operation and the like.
For the above reasons, the invention can be widely popularized in the field of cell culture.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings that are required in the embodiments or the description of the prior art will be briefly described, and it is obvious that the drawings in the following description are some embodiments of the present invention, and other drawings may be obtained according to the drawings without inventive effort to a person skilled in the art.
FIG. 1 is a schematic diagram of a multi-gas incubator according to the present invention.
In the figure: 1. an inner box wall surface; 2. a front door; 3. a cell culture vessel; 4. a heater; 5. a humidifying tray; 6. humidifying water; 7. a supporting plate; 8. a fan; 9. CO 2 A concentration sensor; 10. o (O) 2 A concentration sensor; 11. CO 2 Using an electromagnetic valve; 12. o (O) 2 Using an electromagnetic valve; 13. n (N) 2 Using an electromagnetic valve; 14. an air pump; 15. HEPA filter.
Detailed Description
It should be noted that, without conflict, the embodiments of the present invention and features of the embodiments may be combined with each other. The invention will be described in detail below with reference to the drawings in connection with embodiments.
For the purpose of making the objects, technical solutions and advantages of the embodiments of the present invention more apparent, the technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings in the embodiments of the present invention, and it is apparent that the described embodiments are only some embodiments of the present invention, not all embodiments. The following description of at least one exemplary embodiment is merely exemplary in nature and is in no way intended to limit the invention, its application, or uses. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
It is noted that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of exemplary embodiments according to the present invention. As used herein, the singular is also intended to include the plural unless the context clearly indicates otherwise, and furthermore, it is to be understood that the terms "comprises" and/or "comprising" when used in this specification are taken to specify the presence of stated features, steps, operations, devices, components, and/or combinations thereof.
The relative arrangement of the components and steps, numerical expressions and numerical values set forth in these embodiments do not limit the scope of the present invention unless it is specifically stated otherwise. Meanwhile, it should be clear that the dimensions of the respective parts shown in the drawings are not drawn in actual scale for convenience of description. Techniques, methods, and apparatus known to those of ordinary skill in the relevant art may not be discussed in detail, but are intended to be part of the specification where appropriate. In all examples shown and discussed herein, any specific values should be construed as merely illustrative, and not a limitation. Thus, other examples of the exemplary embodiments may have different values. It should be noted that: like reference numerals and letters denote like items in the following figures, and thus once an item is defined in one figure, no further discussion thereof is necessary in subsequent figures.
In the description of the present invention, it should be understood that the azimuth or positional relationships indicated by the azimuth terms such as "front, rear, upper, lower, left, right", "lateral, vertical, horizontal", and "top, bottom", etc., are generally based on the azimuth or positional relationships shown in the drawings, merely to facilitate description of the present invention and simplify the description, and these azimuth terms do not indicate and imply that the apparatus or elements referred to must have a specific azimuth or be constructed and operated in a specific azimuth, and thus should not be construed as limiting the scope of protection of the present invention: the orientation word "inner and outer" refers to inner and outer relative to the contour of the respective component itself.
Spatially relative terms, such as "above … …," "above … …," "upper surface at … …," "above," and the like, may be used herein for ease of description to describe one device or feature's spatial location relative to another device or feature as illustrated in the figures. It will be understood that the spatially relative terms are intended to encompass different orientations in use or operation in addition to the orientation depicted in the figures. For example, if the device in the figures is turned over, elements described as "above" or "over" other devices or structures would then be oriented "below" or "beneath" the other devices or structures. Thus, the exemplary term "above … …" may include both orientations of "above … …" and "below … …". The device may also be positioned in other different ways (rotated 90 degrees or at other orientations) and the spatially relative descriptors used herein interpreted accordingly.
In addition, the terms "first", "second", etc. are used to define the components, and are only for convenience of distinguishing the corresponding components, and the terms have no special meaning unless otherwise stated, and therefore should not be construed as limiting the scope of the present invention.
As shown in fig. 1, the present invention provides a multi-gas incubator comprising: the box, the conventional incubator that adopts in this embodiment, sets up two sets of layer boards 7 in the box for hold cell culture container 3 (can be culture dish or blake bottle), its front end is equipped with qianmen 2, still be equipped with CO on the lateral wall of box 2 Concentration sensor 9 and O 2 A concentration sensor 10 and a fan 8 for moderating the environment inside the tank.
The bottom of the box body is provided with a humidifying tray 5, humidifying water 6 is arranged in the humidifying tray, and the humidifying tray is heated by a heater 4 arranged in an inner box wall surface 1 at the bottom of the box body to provide temperature conditions and humidity conditions.
The multi-gas channel is communicated with the inside of the box body, is arranged at the rear part of the box body in the figure, and is selectively introduced with CO according to actual control requirements 2 、O 2 、N 2 And air, adjust the culture environment in the box, the end of one side that many air channels link to each other with the box is equipped with HEPA filter 15. The multi-gas channel is respectively connected with CO 2 Channel, O 2 Channel, N 2 The channels are connected to air channels, each of which controls flow rate through various solenoid valves. Correspondingly, each electromagnetic valve is CO 2 By means of solenoid valves 11, O 2 By means of solenoid valves 12, N 2 With a solenoid valve 13 and an air pump 14. According to the practical situation, single gas or multiple groups of gases can be selected to be introduced simultaneously, and the internal environment is regulated.
Generally, the cell culture of the human body is set to a temperature of 37℃and CO 2 The concentration is 5% and the humidity is 90-95%. In addition, sensitive cells such as fertilized eggs, stem cells and IPS cells are usually at 37℃and CO 2 Concentration 5%, O 2 The concentration is 5% and the humidity is 90-95%.
Example 1
Program 1: at 37 ℃, CO 2 Concentration 5%, O 2 The concentration is 5%, the humidity is 90-95%, and the culture time is 72 hours;
program 2: at a temperature of 32 ℃, CO 2 Concentration 5%, O 2 The concentration is 2%, the humidity is 90-95%, and the culture time is 96 hours.
1. After the multi-gas incubator is electrified, the initial temperature is room temperature, and CO 2 Concentration is 0%, O 2 The concentration is about 21% of the oxygen content of the air, and the temperature sensor (arranged in the box, preferably near the air duct in the central area of the box, for accurately measuring the temperature in the box, not shown in the figure) and the CO are used according to the parameters set in the procedure 1 2 Concentration sensor 9, O 2 The actual values fed back by the concentration sensor 10 are compared, and the specific control method is as follows:
1. the set temperature is larger than the actual temperature in the box, the controller controls the output of the heater 4 through PID adjustment (the conventional control mode can meet the requirement and can refer to the PID control method of the carbon dioxide culture temperature), and finally the temperature is stabilized at the set temperature of 37 ℃.
2. When setting the first target CO 2 Concentration is greater than actual CO in the tank 2 Concentration, defining a first target CO 2 The concentration set point is CO 2 SV1，CO 2 Actual value is CO 2 PV1, when CO 2 SV1>CO 2 At PV1, CO 2 Continuously switching on the electromagnetic valve 11 and charging CO 2 Gas until the concentration reaches CO 2 SV1, stop charging CO 2 The gas, i.e. CO, reaching procedure 1 2 And (5) setting concentration.
3. When setting the first target O 2 Concentration is greater than initial O in the box 2 Concentration, defining first target concentration set value as O 2 SV1, initial O 2 Concentration O 2 PV1, when O 2 PV1>O 2 SV1，N 2 Continuously switching on the electromagnetic valve 13 and charging N 2 Gas until the concentration reaches O 2 SV1, stop charging N 2 The gas, i.e. reaching O in procedure 1 2 And (5) setting concentration.
2. When the incubation time satisfies the time set in procedure 1, the temperature and CO set in procedure 2 are used 2 Concentration, O 2 Concentration parameter and temperature sensor, CO 2 By means of sensors 11, O 2 The actual values fed back by the sensor 12 are compared, and the specific control method is as follows:
1. the second target temperature 32 ℃ is smaller than the first target temperature 37 ℃ in the box, the output of the heater 4 is stopped, the air pump 14 is connected, air is introduced, the temperature and the humidity in the box are reduced, and the air pump 14 is stopped after the second target temperature 32 ℃ is set to be equal to the actual temperature in the box. The heater is then operated by PID control to maintain the temperature in the tank at a set temperature.
2. In the cooling process, a large amount of air is filled, and CO in the tank 2 Concentration and O 2 The concentration will change, corresponding to CO 2 Concentration and meeting are reduced, O 2 Concentration is far lower than O in air in procedure 1 2 Concentration of O at this time 2 The concentration will be higher than the O set in procedure 1 2 Concentration.
3. When the second target CO 2 Concentration is greater than actual CO in the tank 2 Concentration, definition of CO 2 The set value is CO 2 SV2, actual CO 2 Concentration of CO 2 PV2, when CO 2 SV2>CO 2 At the time of PV2, CO 2 Continuously switching on the electromagnetic valve 11 and charging CO 2 Gas until the concentration reaches CO 2 SV2, stop charging CO 2 The gas, i.e. CO in procedure 2 2 And (5) setting concentration.
4. After cooling and ventilation, the actual O in the box at the moment 2 The concentration is greater than the first target O set in procedure 1 2 Concentration (O) 2 Concentration 5%), define a second target O 2 Setting value O 2 SV2, actual O 2 Concentration of O 2 PV2, thus, a second target O 2 Set value O 2 SV2(O 2 Concentration 2%) is smaller than actual O 2 Concentration O 2 PV2, i.e. when O 2 SV2<O 2 At PV2, N 2 Continuously switching on the electromagnetic valve 13 and charging N 2 Gas, finally reach O 2 SV2, stop charging N 2 The gas, i.e. reaching O in procedure 2 2 And (5) setting concentration.
5. When the time set in procedure 2 is satisfied, the incubator sounds an alarm, prompting the user to take the incubated sample.
Finally, it should be noted that: the above embodiments are only for illustrating the technical solution of the present invention, and not for limiting the same; although the invention has been described in detail with reference to the foregoing embodiments, it will be understood by those of ordinary skill in the art that: the technical scheme described in the foregoing embodiments can be modified or some or all of the technical features thereof can be replaced by equivalents; such modifications and substitutions do not depart from the spirit of the invention.

Claims (7)

1. A multi-gas incubator, comprising:
the box body is used for accommodating a structure of the cell culture container, the front end of the box body is provided with a front door, and the side wall of the box body is also provided with CO 2 Concentration sensor and O 2 A concentration sensor and a fan;
at least one group of supporting plates, which are arranged in the box body and used for placing the cell culture container;
a humidifying tray arranged at the bottom of the box body and heated by a heater arranged in the wall surface of the inner box at the bottom of the box body;
a multi-gas channel communicated with the box body for selectively introducing CO 2 、O 2 、N 2 And air is used for adjusting the culture environment in the box body, and a HEPA filter is arranged at one side end part of the multi-air channel connected with the box body.
2. The multi-gas incubator of claim 1, wherein the multi-gas channels are each CO-associated with 2 Channel, O 2 Channel, N 2 The channels are connected with the air channels, the flow speed of each channel is controlled by various electromagnetic valves, and one or more channels are selectively selected and simultaneously enter the box body through the multi-air channels.
3. The method for controlling a multi-gas incubator according to claim 1 or 2, wherein the initial parameter conditions and the first target parameter conditions and the second target parameter conditions in the multi-gas incubator are set by a temperature sensor, CO 2 Concentration sensor, O 2 And the sensor feeds back in real time, and after the initial parameter condition is controlled to reach the first target parameter condition, the sensor feeds back to control the first target parameter condition to reach the second target parameter condition, so that the cultivation is completed.
4. A method of controlling a multi-gas incubator according to claim 3, wherein the initial parameter conditions and the first and second target parameter conditions include, but are not limited to, temperature,CO 2 Concentration, O 2 Concentration, humidity and incubation time.
5. The method of controlling a multi-gas incubator according to claim 4, wherein controlling the initial parameter condition to reach the first target parameter condition comprises the steps of:
s11, a first target temperature set in the first target parameters is larger than the initial temperature in the box, and the output of the heater is controlled through PID regulation, so that the temperature is stabilized at the set first target temperature;
s12, setting a first target CO in the first target parameters 2 Defining a first target CO 2 The concentration set point is CO 2 SV1, initial CO 2 Concentration of CO 2 PV1, when CO 2 SV1>CO 2 At PV1, CO 2 Continuously switching on by electromagnetic valve and charging CO 2 Gas until the concentration reaches CO 2 SV1；
S13, setting a first target O in the first target parameters 2 Concentration, defining a first target O 2 The concentration set value is O 2 SV1, initial O 2 Concentration of O 2 PV1, when O 2 PV1>O 2 SV1，N 2 Continuously switching on by electromagnetic valve, and charging N 2 Gas until the concentration reaches O 2 SV1。
6. The method of controlling a multi-gas incubator according to claim 4, wherein controlling the first target parameter condition to reach the second target parameter condition comprises the steps of:
s21, stopping the output of the heater when the second target temperature set in the second target parameter is smaller than the first target temperature in the box, simultaneously switching on the air pump, introducing air, reducing the temperature and humidity in the box, stopping the air pump until the actual temperature in the box is equal to the second target temperature, then controlling the heater to work through PID, and maintaining the temperature in the box stable at the second target temperature;
s22, setting a second target CO in the second target parameters 2 Concentration is greater than actual CO in the tank 2 Concentration, definition of the second target CO 2 The concentration set point is CO 2 SV2, actual CO 2 Concentration of CO 2 PV2, when CO 2 SV2>CO 2 At the time of PV2, CO 2 Continuously switching on by electromagnetic valve and charging CO 2 A gas; until the concentration reaches CO 2 SV2；
S23, setting a second target O in the second target parameters 2 Concentration, definition of the second target O 2 Setting value O 2 SV2, actual O 2 Concentration of O 2 PV2, when O 2 SV2>O 2 At PV2, O 2 Continuously switching on by electromagnetic valve and charging O 2 Gas, finally reach O 2 SV2; when O is 2 SV2<O 2 At PV2, N 2 Continuously switching on by electromagnetic valve, and charging N 2 Gas, finally reach O 2 SV2；
And S24, when the incubation time set by the second target parameter is reached, the incubator gives an alarm to prompt the user to take the incubated sample.
7. The method according to claim 6, wherein in the step S21, when the second target temperature is stabilized, the air pump is stopped, and the following air control is performed.
CN202310236610.0A 2023-03-13 2023-03-13 Multi-gas incubator and control method thereof Pending CN116286350A (en)

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