CN120843269A - A device for cell research and development and application - Google Patents
A device for cell research and development and applicationInfo
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- CN120843269A CN120843269A CN202510703671.2A CN202510703671A CN120843269A CN 120843269 A CN120843269 A CN 120843269A CN 202510703671 A CN202510703671 A CN 202510703671A CN 120843269 A CN120843269 A CN 120843269A
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Abstract
The invention discloses a device for researching and applying cells, which comprises a cell culture module, a cell detection module, a cell sorting module and a cell function regulation and control module which are sequentially connected, wherein the cell culture module is combined with a temperature and humidity sensor, a gas concentration sensor and a regulating device through a microfluidic chip to accurately regulate and control a culture environment, the cell detection module integrates a flow cytometer detection unit and a microscopic imaging detection unit to obtain cell information in multiple dimensions, the cell sorting module utilizes a microfluidic sorting chip to realize accurate sorting according to detection results, the cell function regulation and control module comprises a gene editing unit and a physical stimulation unit to realize diversified cell function regulation and control, and the device is integrally and integrally designed to effectively solve the problems of single function and complex operation of traditional equipment, remarkably improve the efficiency and quality of cell research and application, and has important scientific research and application values.
Description
Technical Field
The invention belongs to the technical field of cell biology, and relates to a device for cell research and development and application.
Background
In the process of cell research and development and application, the prior art has a plurality of defects, the traditional cell culture device is single in function, the accurate regulation and control of various environmental parameters such as temperature, humidity and gas concentration in the process of cell culture are difficult to meet, the function of detecting the cell state on line in real time is lacking, abnormal conditions cannot be timely found and regulated in the process of cell culture, and the quality and efficiency of cell culture are affected.
Disclosure of Invention
The invention aims to provide a device for cell research and development and application, which solves the technical problems.
The technical scheme adopted by the invention is as follows:
the device for cell research and development and application comprises a device main body, wherein a cell culture module, a cell detection module, a cell sorting module and a cell function regulation and control module which are sequentially connected are arranged in the device main body;
The cell culture module comprises a culture chamber, a humidity adjusting device, a gas supply device, a transparent observation window, a temperature sensor, a humidity sensor, a gas concentration sensor, a semiconductor refrigerating and heating sheet, a heating wire, a humidity adjusting device and a gas supply device, wherein the inner wall of the culture chamber is provided with a microfluidic chip, and a plurality of microchannels which are mutually communicated are arranged on the microfluidic chip;
The cell detection module comprises a flow cytometer detection unit and a microscopic imaging detection unit, wherein the flow cytometer detection unit comprises a sheath fluid flow path, a sample flow path and a detection light path, the sheath fluid flow path and the sample flow path are respectively connected with the culture fluid output end of the cell culture module, and the detection light path is provided with a fluorescence detector and a scattering detector;
the cell sorting module comprises a microfluidic sorting chip and a sorting control unit, wherein a plurality of sorting channels and collecting channels are arranged on the microfluidic sorting chip, and the sorting channels are connected with the output end of the cell detection module;
The cell function regulation and control module comprises a gene editing unit and a physical stimulation unit, wherein the gene editing unit comprises a CRISPR-Cas9 delivery system, the CRISPR-Cas9 delivery system is connected with the output end of the cell sorting module, the physical stimulation unit comprises an electric stimulation device, a light stimulation device and a mechanical stimulation device, the electric stimulation device is provided with an electrode array, the light stimulation device is provided with an LED light source array, the mechanical stimulation device is provided with a micro-mechanical vibrator, and the electrode array, the LED light source array and the micro-mechanical vibrator are respectively connected with the output end of the cell sorting module.
The cell culture module comprises a culture chamber, wherein a microfluidic chip is arranged on the inner wall of the culture chamber, a plurality of mutually communicated microchannels are arranged on the microfluidic chip and used for conveying culture fluid and gas, a temperature sensor, a humidity sensor and a gas concentration sensor are arranged in the culture chamber and used for respectively monitoring the temperature, the humidity and the gas concentration in the culture chamber in real time, a semiconductor refrigerating and heating sheet and a heating wire are arranged on the outer wall of the culture chamber and are electrically connected with the temperature sensor, the temperature in the culture chamber is automatically regulated by a signal fed back by the temperature sensor, a humidity regulating device and a gas supply device are further arranged and are electrically connected with the humidity sensor, the gas supply device is electrically connected with the gas concentration sensor and used for regulating the humidity and the gas concentration in the culture chamber according to signals fed back by the humidity sensor and the gas concentration sensor, and a transparent observation window is arranged at the top of the culture chamber and used for observing the cell culture state.
The cell detection module comprises a flow cytometry detection unit and a microscopic imaging detection unit, wherein the flow cytometry detection unit comprises a sheath liquid flow path, a sample flow path and a detection light path, the sheath liquid flow path and the sample flow path are respectively connected with a culture liquid output end of the cell culture module and used for introducing cell culture liquid into the flow cytometry detection unit, a fluorescence detector and a scattering detector are arranged on the detection light path and used for detecting fluorescence signals and scattering light signals of cells so as to acquire information of the size, the shape, the internal structure and cell surface markers of the cells, the microscopic imaging detection unit comprises a microscope, a CCD camera and an image analysis system, an objective table of the microscope is connected with the culture liquid output end of the cell culture module and used for placing cell samples, the CCD camera is connected with the microscope and used for shooting cell images, and the image analysis system is connected with the CCD camera and used for carrying out analysis processing on the shot cell images and acquiring morphological and kinetic information of the cells.
The cell sorting module comprises a microfluidic sorting chip and a sorting control unit, wherein a plurality of sorting channels and collecting channels are arranged on the microfluidic sorting chip, the sorting channels are connected with the output end of the cell detection module, the collecting channels are used for collecting sorted cells, the sorting control unit is electrically connected with the flow cytometer detection unit and the microscopic imaging detection unit, and according to cell information acquired by the cell detection module, the accurate sorting of specific cells is realized by controlling the fluid pressure and the electric field intensity in the microfluidic sorting chip.
The cell function regulation and control module comprises a gene editing unit and a physical stimulation unit, wherein the gene editing unit comprises a CRISPR-Cas9 delivery system, the CRISPR-Cas9 delivery system is connected with the output end of the cell sorting module and used for delivering a gene editing tool to target cells to edit cell genes, the physical stimulation unit comprises an electric stimulation device, a light stimulation device and a mechanical stimulation device, the electric stimulation device is provided with an electrode array, the light stimulation device is provided with an LED light source array, the mechanical stimulation device is provided with a micro-mechanical vibrator, the electrode array, the LED light source array and the micro-mechanical vibrator are respectively connected with the output end of the cell sorting module, and the functions and behaviors of the cells are regulated and controlled by applying different forms of physical stimulation.
Further, the inner wall of the micro-channel of the micro-fluidic chip is coated with a cell affinity coating, and the cell affinity coating is one or a combination of a plurality of fibronectin, laminin or collagen and is used for promoting cell wall-attached growth.
Further, the temperature sensor, the humidity sensor and the gas concentration sensor are miniaturized and high-precision sensors, and the detection precision of the sensors is +/-0.1 ℃ and +/-1% RH and +/-0.1% gas concentration respectively, so that the accurate monitoring of the culture environment is realized.
The humidity adjusting device comprises an ultrasonic humidifier and a dehumidifier, wherein the ultrasonic humidifier is used for increasing the humidity in the culture cavity, and the dehumidifier adopts a condensation dehumidification mode and is used for reducing the humidity in the culture cavity.
Further, the gas supply device comprises a plurality of gas storage tanks for respectively storing carbon dioxide, oxygen and nitrogen, and the flow and the proportion of each gas are precisely controlled by a mass flow controller so as to meet the requirements of different cell cultures on the gas environment.
Further, the temperature regulating system composed of the semiconductor refrigerating and heating plate and the heating wire adopts a PID control algorithm, and the working states of the semiconductor refrigerating and heating plate and the heating wire are automatically regulated according to the difference value between the real-time temperature fed back by the temperature sensor and the set temperature, so that the rapid and stable regulation of the temperature is realized.
Further, the detection light path of the flow cytometer detection unit is also provided with a forward scattering light detector and a side scattering light detector, wherein the forward scattering light detector is used for detecting cell size information, and the side scattering light detector is used for detecting cell internal granularity and complexity information.
Further, dielectrophoresis electrode arrays are arranged in the sorting channels of the microfluidic sorting chip, and the sorting control unit realizes dielectrophoresis sorting of cells by controlling the electric field intensity and frequency of the dielectrophoresis electrode arrays.
Further, the CRISPR-Cas9 delivery system delivers the gene editing tool to the target cell using one or more of liposome transfection, electroporation transfection, or viral vector transfection.
The micro-mechanical vibrator of the mechanical stimulation device has the advantages that the mechanical stimulation device can be used for stimulating the cells, the mechanical stimulation of the cells can be realized, and the mechanical stimulation of the cells can be realized.
In summary, due to the adoption of the technical scheme, the beneficial effects of the invention are as follows:
1. the device for research, development and application of cells is characterized in that a cell culture module realizes accurate transportation of culture solution and gas through a microfluidic chip, and combines various sensors and adjusting devices, so that the temperature, humidity and gas concentration in a culture cavity can be accurately regulated and controlled in real time, a stable and proper growth environment is provided for the cells, and the success rate and quality of cell culture are improved;
2. In the invention, the cell detection module integrates a flow cytometer detection unit and a microscopic imaging detection unit, can detect cells in real time from a plurality of dimensions, acquire various information of the cells, and provide accurate data support for subsequent cell sorting and function regulation;
3. According to the invention, the cell sorting module is based on the information acquired by the cell detection module, and the high-efficiency and accurate sorting of specific cells is realized through the micro-fluid sorting chip and the sorting control unit, so that the sorting process is rapid and accurate, and the risks of cell damage and pollution are reduced;
4. In the invention, the cell function regulation and control module provides gene editing and various physical stimulation modes, can carry out diversified and accurate function regulation and control on cells according to different research demands, and expands the scope and depth of cell research and development and application;
5. in the invention, a plurality of functional modules of cell culture, detection, sorting and functional regulation are integrated, so that the connection and operation links between devices are reduced, the operation complexity and cost are reduced, and the efficiency of cell research and development and application is improved.
Drawings
For a clearer description of the technical solutions of embodiments of the present invention, the drawings that are needed in the embodiments will be briefly described below, it being understood that the following drawings only illustrate some embodiments of the present invention and should not be considered limiting in scope, and other related drawings can be obtained according to these drawings without inventive effort for a person skilled in the art, wherein:
FIG. 1 is a schematic diagram of the structure of the present invention;
the label in the figure comprises a 1-device main body, a 2-cell culture module, a 3-cell detection module, a 4-cell sorting module and a 5-cell function regulating module.
Detailed Description
The present invention will be described in further detail with reference to the drawings and examples, in order to make the objects, technical solutions and advantages of the present invention more apparent. It should be understood that the particular embodiments described herein are illustrative only and are not intended to limit the invention, i.e., the embodiments described are merely some, but not all, of the embodiments of the invention. The components of the embodiments of the present invention generally described and illustrated in the figures herein may be arranged and designed in a wide variety of different configurations.
Thus, the following detailed description of the embodiments of the invention, as presented in the figures, is not intended to limit the scope of the invention, as claimed, but is merely representative of selected embodiments of the invention. All other embodiments, which can be made by a person skilled in the art without making any inventive effort, are intended to be within the scope of the present invention.
It is noted that relational terms such as "first" and "second", and the like, are used solely to distinguish one entity or action from another entity or action without necessarily requiring or implying any actual such relationship or order between such entities or actions. Moreover, the terms "comprises," "comprising," or any other variation thereof, are intended to cover a non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements does not include only those elements but may include other elements not expressly listed or inherent to such process, method, article, or apparatus. Without further limitation, an element defined by the phrase "comprising one does not exclude the presence of other like elements in a process, method, article, or apparatus that comprises an element.
The features and capabilities of the present invention are described in further detail below in connection with examples.
Example 1
The invention relates to a device for cell research and development and application, as shown in figure 1, which comprises a device main body 1, wherein a cell culture module 2, a cell detection module 3, a cell sorting module 4 and a cell function regulation and control module 5 which are sequentially connected are arranged in the device main body 1;
The cell culture module 2 comprises a culture chamber, wherein a micro-fluidic chip is arranged on the inner wall of the culture chamber, a plurality of micro-channels which are mutually communicated are arranged on the micro-fluidic chip, a temperature sensor, a humidity sensor and a gas concentration sensor are arranged in the culture chamber, a semiconductor refrigerating and heating sheet and a heating wire are arranged on the outer wall of the culture chamber and are electrically connected with the temperature sensor, a humidity adjusting device and a gas supply device are also arranged, the humidity adjusting device is electrically connected with the humidity sensor, the gas supply device is electrically connected with the gas concentration sensor, and a transparent observation window is arranged at the top of the culture chamber;
The cell detection module 3 comprises a flow cytometer detection unit and a microscopic imaging detection unit, wherein the flow cytometer detection unit comprises a sheath fluid flow path, a sample flow path and a detection light path, the sheath fluid flow path and the sample flow path are respectively connected with the culture fluid output end of the cell culture module, and the detection light path is provided with a fluorescence detector and a scattering detector;
the cell sorting module 4 comprises a micro-fluid sorting chip and a sorting control unit, wherein the micro-fluid sorting chip is provided with a plurality of sorting channels and collecting channels, and the sorting channels are connected with the output end of the cell detection module;
The cell function regulation and control module 5 comprises a gene editing unit and a physical stimulation unit, wherein the gene editing unit comprises a CRISPR-Cas9 delivery system, the CRISPR-Cas9 delivery system is connected with the output end of the cell sorting module, the physical stimulation unit comprises an electric stimulation device, a light stimulation device and a mechanical stimulation device, the electric stimulation device is provided with an electrode array, the light stimulation device is provided with an LED light source array, the mechanical stimulation device is provided with a micro-mechanical vibrator, and the electrode array, the LED light source array and the micro-mechanical vibrator are respectively connected with the output end of the cell sorting module.
The inner wall of the micro-channel of the micro-fluidic chip is coated with a cell affinity coating, and the cell affinity coating is one or a combination of a plurality of fibronectin, laminin or collagen.
The temperature sensor, the humidity sensor and the gas concentration sensor are miniaturized and high-precision sensors, and the detection precision of the sensors is +/-0.1 ℃ and +/-1% RH and +/-0.1% gas concentration respectively.
The humidity adjusting device comprises an ultrasonic humidifier and a dehumidifier, and the dehumidifier adopts a condensation dehumidification mode.
The gas supply device comprises a plurality of gas storage tanks and respectively stores carbon dioxide, oxygen and nitrogen.
The temperature regulating system composed of the semiconductor refrigerating and heating sheet and the heating wire adopts a PID control algorithm.
The detection light path of the flow cytometer detection unit is also provided with a forward scattering light detector and a side scattering light detector.
A dielectrophoresis electrode array is arranged in a sorting channel of the microfluidic sorting chip.
The CRISPR-Cas9 delivery system employs one or more of liposome transfection, electroporation transfection or viral vector transfection.
The electrode array of the electric stimulation device adopts flexible electrodes, the LED light source array of the optical stimulation device can emit light with different wavelengths, and the vibration frequency and the vibration amplitude of the micro-mechanical vibrator of the mechanical stimulation device can be adjusted.
The cell suspension and the culture solution are injected into a culture cavity through a micro-channel of a micro-fluidic chip, a temperature sensor, a humidity sensor and a gas concentration sensor monitor environmental parameters in the culture cavity in real time and feed data back to a corresponding adjusting device, when the temperature sensor detects that the temperature deviates from a set value, a semiconductor refrigerating and heating plate and a heating wire are automatically started to adjust the temperature in the culture cavity, the humidity adjusting device and a gas supply device adjust the humidity and the gas concentration in the culture cavity according to feedback signals of the humidity sensor and the gas concentration sensor, scientific researchers can observe the cell culture state through a transparent observation window, and a cell detection module can also be utilized to periodically detect cells.
When the cell is required to be detected, the cell culture solution flows into the cell detection module from the cell culture module, the cell culture solution is introduced into the flow cytometry detection unit through the sheath solution flow path and the sample flow path, the cell sequentially passes through the detection light path under the wrapping of the sheath solution, the fluorescence signal and the scattered light signal of the cell are detected by the fluorescence detector and the scattered light detector, the relevant information of the cell is obtained, the cell sample is placed on the objective table of the microscope in the microscopic imaging detection unit, the CCD camera shoots the cell image, and the image is analyzed and processed by the image analysis system, so that the morphological and kinetic information of the cell is obtained.
And then, by controlling the fluid pressure and the electric field intensity in the micro-fluid sorting chip, the target cells enter the corresponding collecting channels, thereby realizing the accurate sorting of the cells.
For the physical stimulation regulation and control, scientific researchers can select an electric stimulation device, a light stimulation device or a mechanical stimulation device according to experimental purposes, and apply corresponding physical stimulation to cells through an electrode array, an LED light source array or a micromechanical vibrator to regulate and control the functions and behaviors of the cells.
Example two
The invention relates to a device for cell research and development and application, which comprises the following specific implementation modes:
culture, detection, sorting and gene editing of human embryonic stem cells
The method comprises the steps of mixing a human embryonic stem cell suspension with a culture solution containing a specific growth factor, injecting the mixture into a culture cavity through a microfluidic chip of a cell culture module, setting the initial temperature in the culture cavity to be 37 ℃, setting the humidity to be 95%, setting the gas concentration to be 5% CO 2、20%O2 and 75% N 2, monitoring environmental parameters in real time by a temperature sensor, a humidity sensor and a gas concentration sensor, quickly adjusting a semiconductor refrigerating and heating plate and a heating wire under a PID control algorithm when the temperature fluctuates by +/-0.2 ℃, enabling the temperature to be restored to a set value, automatically starting an ultrasonic humidifier or a dehumidifier to adjust if the humidity deviates from +/-2% RH, accurately maintaining the gas proportion through feedback of the gas concentration sensor by a gas supply device, and observing the cell state by a scientific research person through a transparent observation window in the culture process.
Cell detection, namely, after culturing for 48 hours, introducing part of cell culture solution into a cell detection module, wherein in a flow cytometry detection unit, sheath solution enters at a flow rate of 0.5mL/min and sample solution enters at a flow rate of 0.1mL/min, cells sequentially pass through a detection light path under the wrapping of the sheath solution, a forward scattering light detector, a side scattering light detector and a fluorescence detector detect the information of cell size, internal granularity and surface markers, and in a microscopic imaging detection unit, the cell morphological structure is observed by using a phase difference objective lens of an inverted microscope, so that the differentiation signs of part of cells are found.
Cell sorting, namely determining the undifferentiated human embryonic stem cells as target cells by a sorting control unit according to the cell surface marker information acquired by the cell detection module. The accurate separation is realized by controlling the fluid pressure in the micro-fluid separation chip to be 50kPa and simultaneously applying an electric field with the frequency of 1MHz and the intensity of 10V/cm on the dielectrophoresis electrode array, so that the undifferentiated human embryonic stem cells enter a specific collecting channel.
And (3) cell function regulation, namely conveying the sorted undifferentiated human embryonic stem cells to a gene editing unit of a cell function regulation module, conveying a CRISPR-Cas9 complex carrying a specific gene editing sequence into the cells by adopting a liposome transfection mode, and editing specific genes of the cells so as to study the influence of the genes on cell differentiation.
Example III
The invention relates to a device for cell research and development and application, which comprises the following specific implementation modes:
Tumor cell culture, detection, sorting and physical stimulation regulation and control
Cell culture, namely injecting tumor cell suspension and conventional culture solution into a culture cavity through a microfluidic chip, wherein the culture temperature is set to 36.5 ℃, the humidity is 90%, and the gas concentration is 5% CO 2 and 95% N 2. In the culture process, each sensor monitors environmental parameters in real time, and the culture environment is ensured to be stable.
Cell detection cells were detected after 72 hours of incubation. Detecting fluorescence signals and scattered light signals of cells in a flow cytometry detection unit at a sheath fluid flow rate of 0.6mL/min and a sample fluid flow rate of 0.15mL/min to obtain information of cell proliferation activity, and observing the protein expression condition of fluorescent markers in the cells in a microscopic imaging detection unit by using a fluorescent objective lens.
Cell sorting, namely determining tumor cells with high proliferation activity and high expression of specific proteins as target cells by a sorting control unit according to the cell proliferation activity and protein expression information obtained by a cell detection module. And sorting the target tumor cells to a designated collection channel by adjusting the fluid pressure in the microfluidic sorting chip and the electric field parameters of the dielectrophoresis electrode array.
And (3) regulating the cell function, namely sending the sorted tumor cells into a physical stimulation unit of a cell function regulating module, applying electric stimulation with the frequency of 10Hz and the voltage of 1V by using a flexible electrode of an electric stimulation device, continuously stimulating for 2 hours, simultaneously starting the optical stimulation device, emitting light with the wavelength of 635nm, irradiating the cells for 30 minutes, and researching the influence of different physical stimulations on the migration and invasion capacities of the tumor cells.
The above description is only of the preferred embodiments of the present invention and is not intended to limit the scope of the present invention, and any modifications, equivalents, improvements and modifications within the spirit and principles of the invention will become apparent to those skilled in the art.
Claims (10)
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| CN202510703671.2A CN120843269A (en) | 2025-05-29 | 2025-05-29 | A device for cell research and development and application |
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| CN202510703671.2A CN120843269A (en) | 2025-05-29 | 2025-05-29 | A device for cell research and development and application |
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