WO2019148865A1 - 细胞培养监测设备及培养监测方法 - Google Patents

细胞培养监测设备及培养监测方法 Download PDF

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Publication number
WO2019148865A1
WO2019148865A1 PCT/CN2018/107841 CN2018107841W WO2019148865A1 WO 2019148865 A1 WO2019148865 A1 WO 2019148865A1 CN 2018107841 W CN2018107841 W CN 2018107841W WO 2019148865 A1 WO2019148865 A1 WO 2019148865A1
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Prior art keywords
culture
cell
image
cell sheet
image capturing
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PCT/CN2018/107841
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English (en)
French (fr)
Inventor
周适
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京东方科技集团股份有限公司
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Priority to US16/479,704 priority Critical patent/US20200385667A1/en
Publication of WO2019148865A1 publication Critical patent/WO2019148865A1/zh

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
    • C12M41/12—Means for regulation, monitoring, measurement or control, e.g. flow regulation of temperature
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M23/00—Constructional details, e.g. recesses, hinges
    • C12M23/02—Form or structure of the vessel
    • C12M23/10—Petri dish
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M23/00—Constructional details, e.g. recesses, hinges
    • C12M23/22—Transparent or translucent parts
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M23/00—Constructional details, e.g. recesses, hinges
    • C12M23/48—Holding appliances; Racks; Supports
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M25/00—Means for supporting, enclosing or fixing the microorganisms, e.g. immunocoatings
    • C12M25/02—Membranes; Filters
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M25/00—Means for supporting, enclosing or fixing the microorganisms, e.g. immunocoatings
    • C12M25/14—Scaffolds; Matrices
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M27/00—Means for mixing, agitating or circulating fluids in the vessel
    • C12M27/16—Vibrating; Shaking; Tilting
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
    • C12M41/12—Means for regulation, monitoring, measurement or control, e.g. flow regulation of temperature
    • C12M41/14—Incubators; Climatic chambers
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
    • C12M41/30—Means for regulation, monitoring, measurement or control, e.g. flow regulation of concentration
    • C12M41/36—Means for regulation, monitoring, measurement or control, e.g. flow regulation of concentration of biomass, e.g. colony counters or by turbidity measurements
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
    • C12M41/46—Means for regulation, monitoring, measurement or control, e.g. flow regulation of cellular or enzymatic activity or functionality, e.g. cell viability
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/06—Fluid handling related problems
    • B01L2200/0647—Handling flowable solids, e.g. microscopic beads, cells, particles
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/02—Identification, exchange or storage of information
    • B01L2300/023—Sending and receiving of information, e.g. using bluetooth
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
    • B01L3/54—Labware with identification means
    • B01L3/545—Labware with identification means for laboratory containers

Definitions

  • the present disclosure relates to the field of tissue engineering, and in particular to a cell culture monitoring device and a culture monitoring method.
  • CST Cell Sheet Technology mainly uses temperature-reacting culture dishes to harvest cells. CST avoids the use of proteases for cell processing, thus preserving the extracellular matrix and related proteins and factors that are autocrine during culture, allowing cells to be collected in a complete membranous structure.
  • Cell sheet is a research hotspot in the field of tissue engineering in recent years, and has a wide range of applications in the treatment of skin, cornea, heart and periodontal diseases.
  • a cell culture monitoring apparatus comprising:
  • a culture vessel configured to contain a culture medium of the cell sheet
  • An image capture device configured to acquire an image within the culture vessel to monitor a growth state of the cell sheet.
  • the bottom of the culture vessel comprises:
  • a temperature sensitive layer configured to carry the culture substrate
  • a luminescent layer located on a side of the temperature sensitive layer remote from the cell sheet, is configured to provide backlighting for image acquisition of the cell sheet by luminescence.
  • a light transmissive insulating layer is further disposed between the temperature sensitive layer and the luminescent layer, configured to at least partially reduce heat transfer between the temperature sensitive layer and the luminescent layer.
  • the cold light source is located at an adjacent location inside the luminescent layer or outside the luminescent layer.
  • the bottom of the culture vessel further comprises:
  • the method further includes:
  • a temperature regulating element configured to adjust an internal temperature of the culture vessel.
  • a light blocking structure is provided on the outside of the culture vessel.
  • the temperature regulating element comprises a plurality of semiconductor temperature control elements arranged along an outer wall of the culture vessel, the plurality of semiconductor temperature control elements being arranged to form a light blocking structure.
  • the culture vessel is located within an incubator.
  • one or more spatial layers are disposed in the incubator, and the space layer is provided with a bracket configured to support a plurality of the culture containers, and the plurality of the culture containers are fixed or Mounted on the bracket.
  • an integral or detachable positioning structure is provided on the exterior of the culture vessel, the positioning structure mating with the bracket and movable relative to the bracket.
  • the method further includes:
  • a driving mechanism configured to drive the image capturing device to move on the guiding mechanism to adjust an image capturing position between the plurality of the culture containers.
  • the guiding mechanism comprises: a parallel pair of first rails and a second rail disposed between the pair of first rails; the driving mechanism comprises:
  • a first driving mechanism disposed between the pair of first rails and the second rail, configured to drive the second rail along the extending direction of the pair of first rails relative to the paired first Guide rail movement;
  • a second driving mechanism is disposed between the second rail and the image capturing device and configured to drive the image capturing device to move relative to the second rail along an extending direction of the second rail.
  • At least one of the following devices is also included:
  • control device configured to control an external environmental parameter of the culture vessel
  • An image processing apparatus configured to process an image of the cell sheet acquired by the image acquisition device to obtain a growth state of the cell sheet.
  • control device is coupled to the image processing device and configured to adjust an external environmental parameter of the culture vessel based on a growth state of the cell sheet.
  • a culture monitoring method based on the foregoing cell culture monitoring device comprising:
  • the culture substrate of the cell sheet is placed in a culture vessel to grow the cell sheet layer in the culture substrate;
  • an image in the culture container is acquired by the image acquisition device to monitor the growth state of the cell sheet.
  • the cell culture monitoring device includes a guiding mechanism and a driving mechanism disposed above the culture container; the culture monitoring method further includes:
  • the image capturing device is controlled to perform image acquisition, and after the acquisition is completed, the driving mechanism is controlled to drive the image capturing device to move to an image capturing position corresponding to another culture container to continue image capturing.
  • FIG. 1 is a schematic structural view of some embodiments of a cell culture monitoring device according to the present disclosure
  • FIG. 2 is a schematic illustration of a culture vessel in a top view angle in accordance with some embodiments of the cell culture monitoring device of the present disclosure
  • FIG. 3 is a schematic structural view of a culture container in some embodiments of a cell culture monitoring device according to the present disclosure
  • FIG. 4 is a schematic diagram of the bottom structure of a culture vessel in some embodiments of a cell culture monitoring device according to the present disclosure
  • Figure 5 is a schematic illustration of the bottom structure of a culture vessel in accordance with further embodiments of the cell culture monitoring device of the present disclosure
  • Figure 6 is a schematic illustration of the external structure of a culture vessel in some embodiments of a cell culture monitoring device according to the present disclosure
  • FIG. 7 is a schematic structural view of still another embodiment of a cell culture monitoring device according to the present disclosure.
  • FIG. 8 is a schematic view showing the structure of a positioning structure and a bracket in accordance with still another embodiment of the cell culture monitoring device of the present disclosure
  • FIG. 9 is a schematic view showing the structure of a guiding mechanism and a driving mechanism in accordance with some embodiments of the cell culture monitoring device of the present disclosure.
  • FIG. 10 is a schematic flow diagram of some embodiments of a culture monitoring method in accordance with the present disclosure.
  • FIG. 11 is a schematic flow diagram of still other embodiments of a culture monitoring method in accordance with the present disclosure.
  • a particular device when it is described that a particular device is located between the first device and the second device, there may be intervening devices between the particular device and the first device or the second device, or there may be no intervening devices.
  • that particular device can be directly connected to the other device without intervening devices, or without intervening devices directly connected to the other devices.
  • the culture state of the cells is indirectly detected by sampling the culture solution in the cell culture apparatus and by detecting the change in the index of the culture solution.
  • the inventors have found through research that such related technologies do not take into account the specificity of the cell sheet when it is applied to cell culture monitoring such as cell sheets, and it is difficult to measure the area, thickness, uniformity and level of the cell sheet. Achieve effective detection.
  • the present disclosure provides an implementation structure and principle of some embodiments of the cell culture monitoring device.
  • the so-called cell culture includes the culture of a single or multi-layered Cell Sheet.
  • the cultured cells may include, for example, various animal cells and human cells isolated from human, mouse, rat, guinea pig, hamster, chicken, rabbit, pig, sheep, cow, horse, dog, cat, monkey, and the like. .
  • the types of cells may include, for example, keratinocytes, splenocytes, nerve cells, glial cells, pancreatic beta cells, mesangial cells, epidermal cells, epithelial cells (corneal epithelial cells, oral mucosal epithelial cells, amniotic epithelial cells, etc.) Endothelial cells (vascular endothelial cells, corneal endothelial cells, etc.), fibroblasts, parenchymal cells (hepatocytes, corneal parenchymal cells, etc.), muscle cells including smooth muscle cells such as vascular smooth muscle cells, fat cells, synovial cells, cartilage Cells, bone cells, osteoblasts, osteoclasts, breast cells, hepatocytes, periosteal-derived cells or mesenchymal cells, or precursor cells of cells of the aforementioned type.
  • the cells may also include stem cells or cancer cells such as embryonic stem cells (ESCs) and mesenchymal stem cells (MSCs).
  • the biological source of the cells may be homologous, heterologous or the same individual as long as it can be cultured at the cellular level. That is to say, these cells can be either allogenic cells or different Xenogenic cells. These cells may also be of the same type (Same Type) and different types (Different Type), for example, the same type of cells of the same animal, different kinds of cells of the same animal, the same kind of cells of the heterologous animal, Different types of cells of a heterologous animal, cells of the same type of cells of the same individual, cells of different types of the same individual, and the like.
  • FIG. 1 is a schematic structural view of some embodiments of a cell culture monitoring device according to the present disclosure, in conjunction with the schematic view of the culture vessel of FIG. 2 in a plan view.
  • the cell culture monitoring device of this embodiment comprises:
  • a culture vessel 100 configured to hold a culture substrate 500 of the cell sheet 600;
  • the image capture device 200 is configured to acquire an image within the culture container 100 to monitor the growth state of the cell sheet 600.
  • the culture container 100 has a housing space capable of accommodating the culture substrate 500 and the cell sheet 600, and provides a growth space for the cell sheet 600.
  • the culture substrate 500 is capable of providing nutrient material to the cell sheet 600 to enable it to grow from a single cell or cell colony into a cell sheet 600 having a certain area and thickness.
  • composition and culture conditions of the culture medium can be adjusted or changed according to the difference of the cells to be cultured.
  • appropriate culture media and culture conditions can be selected and designed according to the guidelines of the CSH Protocols.
  • the image capture device 200 can employ various types of imaging elements that can perform imaging imaging, such as cameras based on CCD or CMOS imaging, imagers based on infrared imaging or infrared thermal imaging, and the like.
  • the image capture device 200 is capable of acquiring images in the culture container 100 in accordance with a predetermined time interval, in accordance with an instruction or in real time.
  • the image acquired by image acquisition device 200 can reflect the growth of cell sheet 600 at different stages of growth.
  • the important index data of the cell sheet 600 such as the area, the thickness, the degree of uniformity, and the like, can be further obtained by analyzing and processing the image, thereby satisfying the condition monitoring requirement in the cell sheet culture process.
  • the cell sheet 600 can be collected in time by the collection tool.
  • the coverage area of the cell sheet 600 in the image is measured.
  • the thickness of the cell sheet 600 is determined by measuring the alignment of the transmittance of the cell sheet 600 in the image with a preset standard value.
  • the degree of uniformity and/or flatness of the cell sheet 600 is determined by measuring the heat profile and/or infrared map of the cell sheet 600 in the image.
  • the culture container 100 includes a culture dish body 110 and a dish cover 120 which can be covered on the top of the culture dish body 110 to effect closure of the internal space of the culture dish body 110.
  • the inner wall and the bottom of the culture dish body 110 enclose an accommodation space of the culture substrate 500 and the cell sheet 600.
  • An integral or detachable positioning structure 130 may be disposed outside of the culture vessel 100.
  • the positioning structure 130 can be fixedly disposed outside the outer wall of the culture dish body 110 to facilitate the position setting and adjustment of the culture dish 110.
  • a recessed structure 131 that mates with the guide can be disposed on the positioning structure 130.
  • a temperature sensitive material is commonly used in the related art as a substrate layer (ie, a temperature sensitive layer) to carry a cell sheet, such as poly N-isopropyl acrylamide (PIPAAm), A complex of PIPAAm with methacrylic acid, a short peptide of lysine A6K, and the like.
  • PIPAAm poly N-isopropyl acrylamide
  • the temperature sensitive material is very sensitive to temperature. When the temperature sensed by the temperature sensitive material reaches a certain condition, such as heating/cooling to a specific temperature, the cell sheet changes from a state close to the temperature sensitive material to an easily detachable state, thereby facilitating Harvesting and collection of cell sheets.
  • some cell culture monitoring schemes use the light receiving portion of the measuring unit to receive light from the illuminating portion through the culture solution in the cell culture device, and determine the culture condition of the cells based on the received light.
  • the heat of the light source used in the light-emitting portion may cause local overheating to damage the cell sheet and may cause temperature-sensitive materials attached to the cell sheet. Adverse effects make monitoring of cell sheets very difficult.
  • the bottom of the culture container 100 includes the temperature sensitive layer 140 and the luminescent light.
  • the layer 170, the luminescent layer 170 is located on a side of the temperature sensitive layer 140 away from the cell sheet 600.
  • the temperature sensitive layer 140 is configured to carry the culture substrate 500.
  • the luminescent layer 170 is configured to provide backlighting for image acquisition of the cell sheet 600 by luminescence (eg, based on luminescence from the cold light source 160) to ensure quality of image acquisition.
  • the luminescence process of the luminescent light does not generate significant heat, such as fluorescence, phosphorescence, bacterior light, etc., so the amount of heat generation is small, and the temperature of the temperature-sensitive layer 140 is less affected. It is also not easy to cause local overheating.
  • the luminescent layer 170 can be provided with light dispersing particles, which can scatter the cold light emitted by the cold light source 160 into the entire luminescent layer 170, thereby providing a more uniform backlight effect, thereby improving the quality of the captured image.
  • the cold light source 160 can generate luminescence based on the principles of photoluminescence, the principle of cathodoluminescence, or the principle of high energy particle illumination.
  • the cold light source 160 may employ a luminescent light emitting diode, a cold light, a fluorescent plate or a luminescent sheet, or the like.
  • the cold light source 160 can be disposed at an abutting location outside of the luminescent layer 170.
  • the cold light source 160 may be disposed on the positioning structure 130 at a position adjacent to the outside of the luminescent layer 170.
  • the cold light source 160 is disposed outside the cold light layer 170, and the influence of the heat generation amount of the cold light source 160 on the temperature sensitive layer 140 can be further reduced.
  • FIG. 5 is a schematic illustration of the bottom structure of a culture vessel in accordance with further embodiments of the cell culture monitoring device of the present disclosure.
  • a cold light source 160 may be disposed inside the luminescent layer 170 to make the structure of the culture vessel 100 more compact.
  • a single cold light source 160 can be provided inside the luminescent layer 170.
  • a plurality of granular or elongated cold light sources 160 may be disposed inside the luminescent layer 170 and arranged randomly or in a predetermined array.
  • a light transmissive thermal insulation layer 150 is further disposed between the temperature sensitive layer 140 and the luminescent layer 170 .
  • the light transmissive insulating layer 150 can be configured to at least partially reduce heat transfer between the temperature sensitive layer 140 and the luminescent layer 170 to further reduce the adverse effects of heat of the luminescent layer 170 on the temperature sensitive layer 140.
  • the light-transmissive heat-insulating layer 150 may be made of a heat-insulating material that is at least partially transmitted through the luminescent layer 170, such as glass or plastic that can transmit light.
  • the bottom of the culture vessel 100 further includes a wiring layer 180.
  • the wiring layer 180 is located on a side of the luminescent layer 170 away from the cell sheet layer 600.
  • FIG. 6 is a schematic illustration of the external structure of a culture vessel in some embodiments of a cell culture monitoring device in accordance with the present disclosure.
  • a plurality of semiconductor temperature control elements 190 are closely arranged on the outer wall of the culture vessel 100.
  • These semiconductor temperature control elements 190 can be used as temperature regulating elements to adjust the internal temperature of the culture vessel 100 to achieve finer temperature control during cell sheet 600 culture, thereby ensuring stable growth of the cell sheet 600.
  • the temperature regulating element can also stabilize the temperature sensitive layer 140 during growth of the cell sheet 600.
  • the temperature adjusting element can also adjust the internal temperature of the culture container 100 to a temperature range in which the temperature sensitive layer 140 is easily separated from the cell sheet 600, thereby reducing the difficulty of collecting the cell sheet 600. .
  • the temperature adjustment element is not limited to including the semiconductor temperature control element 190 shown in FIG.
  • the temperature regulating element may also include an electronic thermostat or a vapor thermostat or the like.
  • the temperature adjustment member in some embodiments may be disposed inside the culture container 100 or on the outer side of the culture container 100, but not in close contact with the outer wall of the culture container 100 or the like.
  • the outer side of the culture container 100 is provided with a light shielding structure.
  • FIG. 6 shows a plurality of semiconductor temperature control elements 190 arranged along the outer wall of the culture vessel 100, and these semiconductor temperature control elements 190 can be arranged to form a light shielding structure. Since the semiconductor material is non-transparent with respect to visible light, it is possible to form a light-shielding structure, thereby achieving a more rapid temperature adjustment effect and limiting the leakage of internal cold light or the penetration of external light.
  • FIG. 7 is a schematic illustration of the structure of other embodiments of cell culture monitoring devices in accordance with the present disclosure.
  • the culture vessel 100 is disposed within the incubator 300.
  • the incubator 300 is capable of providing an external environment suitable for the growth of the cell sheet 600, increasing the controllability and reliability of the external environment in which the cell sheet 600 is grown.
  • the incubator 300 is very convenient in many aspects such as arrangement, setting, and use, and is advantageous for use by an operator.
  • one or more spatial layers are provided within the incubator 300.
  • a bracket 310 configured to support a plurality of the culture containers 100 is disposed in the space layer, and the plurality of the culture containers 100 are fixedly or detachably mounted on the bracket 310, thereby satisfying batches of cells
  • the culture and monitoring requirements of the sheet are beneficial to achieve large-scale and industrial cultivation of the cell sheet and reduce the production cost of the cell sheet.
  • the culture vessel may be included A control device that controls the external environmental parameters of 100.
  • input of external environmental parameters is received through the operation panel 330 on the incubator 300 of FIG. 7, and display of related parameters and monitoring data is performed through the display screen 320.
  • image processing devices may also be included in other embodiments.
  • the image processing device may be configured to process an image of the cell sheet 600 acquired by the image capture device 200 to obtain a growth state of the cell sheet 600.
  • the control device is coupled to the image processing device and configured to adjust an external environmental parameter of the culture vessel based on a growth state of the cell sheet.
  • the computer 400 is connected to the image capturing device 200 in the incubator 300, and processes the image of the cell sheet 600 acquired by the image capturing device 200 to obtain various important indexes of the cell sheet 600. Data, such as area, thickness, uniformity and level of peace.
  • control device may have data processing through a central processing unit (CPU) or a field programmable logic array (FPGA) or a single chip microcomputer (MCU) or a digital signal processor (DSP) or an application specific integrated circuit (ASIC).
  • CPU central processing unit
  • FPGA field programmable logic array
  • MCU single chip microcomputer
  • DSP digital signal processor
  • ASIC application specific integrated circuit
  • control device and the image processing device may be integrated in the same processor or separately implemented by different processors.
  • control device can be connected to the image processing device.
  • connection can be implemented by including a wireless network, a wired network, and/or any combination of a wireless network and a wired network, and the like.
  • the network may include a local area network, the Internet, a telecommunications network, an internet of things based on the Internet and/or telecommunications network, and/or any combination of the above networks, and the like.
  • the wired network can communicate by, for example, twisted pair, coaxial cable or optical fiber transmission.
  • the wireless network can be, for example, a mobile communication network, Bluetooth, Zigbee or Wi-Fi.
  • FIG. 8 is a schematic illustration of the configuration of the positioning structure in cooperation with the stent in accordance with further embodiments of the cell culture monitoring device of the present disclosure.
  • the exterior of the culture vessel 100 is provided with an integral or detachable positioning structure 130 that mates with the bracket 310 and relative to the bracket 310 it can move.
  • the stent 310 in the form of an elongated rod shown in FIG. 8 can be in a guiding fit with the concave structure 131 of the positioning structure 130 in addition to the supporting action on the culture container 100, so that the culture container 100 is on the holder 310.
  • the layout and position adjustment are more convenient.
  • Figures 7 and 8 illustrate the arrangement of multiple and multiple sets of culture vessels 100 within the incubator 300, respectively, in some embodiments.
  • One or more image capture devices 200 may be provided in each layer of the incubator 300.
  • the image capture device 200 can be moved to an image capture location corresponding to different culture containers 100 for image capture operations.
  • the image capture device 200 may also be disposed outside the incubator 300 to simplify the configuration of the incubator 300 and reduce the footprint of the incubator 300.
  • the image capture location of image capture device 200 can be manually adjusted by an operator. In other embodiments, automated adjustment means can also be employed to save manpower and improve efficiency.
  • Figure 9 is a schematic illustration of the construction of a steering mechanism and a drive mechanism in accordance with some embodiments of the cell culture monitoring device of the present disclosure.
  • the cell culture monitoring device further includes a guiding mechanism 700 and a driving mechanism 800.
  • the guiding mechanism 700 is disposed above the culture container 100
  • the driving mechanism 800 is configured to drive the image capturing device 200 to move on the guiding mechanism 700, thereby conveniently implementing the image collecting device 200 in a plurality of the training
  • the image collection position is adjusted between the containers 100.
  • these examples allow for image acquisition at different times in a position-adjusted manner using a smaller number of image capture devices, thereby reducing the use of image capture devices while reducing monitoring requirements.
  • Product cost and energy consumption are examples of energy consumption.
  • Fig. 9 shows an example of a fitting structure of a guide mechanism and a drive mechanism which are simple and easy to implement.
  • the guiding mechanism 700 includes a parallel pair of first rails 710 and a second rail 720 disposed between the pair of first rails 710.
  • the drive mechanism 800 includes a first drive mechanism 810 and a second drive mechanism 820.
  • the first driving mechanism 810 is disposed between the pair of first rails 710 and the second rails 720, and is configured to drive the second rails 720 along the extending direction of the pair of first rails 710 with respect to The pair of first guide rails 710 move.
  • a second driving mechanism 820 is disposed between the second rail 720 and the image capturing device 200 and configured to drive the image capturing device 200 along the extending direction of the second rail 720 relative to the second rail 720 motion.
  • the first driving mechanism 810 and the second driving mechanism 820 can adopt various driving power components (such as a motor, a gas pump or a hydraulic pump, etc.), and select a transmission structure as needed (for example, a rack and pinion transmission structure, a gear train transmission structure) Or pulley block transmission mechanism, etc.).
  • the control driving mechanism 800 drives the image capturing device 200 to move on the guiding mechanism 700 to move the image capturing device 200 to the corresponding corresponding to the culture container 100.
  • Image capture location For example, the first driving mechanism 810 can first drive the second rail 720 to move along the pair of first rails 710 to the column where the culture container 100 is located, and the second driving mechanism 820 drives the image capturing device 200 to move to the culture container. Above the 100, the location is the image acquisition location corresponding to the culture vessel 100. In other examples, the driving steps of the first drive mechanism 810 and the second drive mechanism 820 may be reversed or performed simultaneously.
  • the image capturing device 200 can be controlled to perform image acquisition, and after the acquisition is completed, the driving mechanism 800 is controlled to drive the image capturing device 200 to move to another image capturing position corresponding to the culture container 100 to continue image collection. .
  • the present disclosure also provides corresponding embodiments of culture detection methods.
  • Figure 10 a schematic flow diagram of some embodiments of a method of monitoring a culture according to the present disclosure.
  • the culture monitoring method may include:
  • Step S1 the culture substrate 500 of the cell sheet 600 is placed in the culture container 100, in order to grow the cell sheet 600 in the culture substrate 500;
  • Step S2 During the growth of the cell sheet 600, an image in the culture container 100 is acquired by the image capture device 200 to monitor the growth state of the cell sheet 600.
  • the culture substrate 500 and the single cell or cell colony which is the basis for the growth of the cell sheet 600 can be placed into the culture container 100 by an operator manually or by a tool, and then during the growth of the cell sheet 600.
  • the image in the culture container 100 is acquired by an operator manually or by an automatic control image acquisition device 200 by a control device.
  • a guiding mechanism 700 and a driving mechanism 800 disposed above the culture container 100 may also be included.
  • Corresponding culture monitoring methods may also include:
  • Step S3 controlling the driving mechanism 800 to drive the image capturing device 200 to move on the guiding mechanism 700 to adjust the image capturing device 200 to move to an image capturing position corresponding to the culture container 100;
  • Step S4 controlling the image acquisition device 200 to perform image acquisition, and controlling the driving mechanism 800 to drive the image acquisition device 200 to move to another image collection position corresponding to the culture container 100 to continue image collection after the acquisition is completed.
  • the driving operation of the driving mechanism 800 and the image capturing operation of the image capturing device 200 can be controlled by a single or different control device, which can be implemented by a general purpose or special purpose computing device running a control program.
  • Step S3 and step S4 may be embodied as step S2 of the foregoing method embodiment, or may be performed independently of the step S2.

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Abstract

提供一种细胞培养监测设备及培养监测方法，细胞培养监测设备包括：培养容器，配置成容纳细胞片层的培养基质；和图像采集装置，配置成获取培养容器内的图像，以监测细胞片层的生长状态。

Description

细胞培养监测设备及培养监测方法
相关申请的交叉引用
本申请是以申请号为201810089834.2，申请日为2018年1月30日的中国专利申请为基础，并主张其优先权，该中国专利申请的公开内容在此作为整体引入本申请中。
技术领域
本公开涉及组织工程领域，尤其涉及一种细胞培养监测设备及培养监测方法。
背景技术
细胞片层技术(Cell Sheet Technology，简称CST)主要应用温度反应培养皿收获细胞。CST避免了对细胞使用蛋白酶进行处理，因此保留了培养过程中细胞自分泌的细胞外基质及相关蛋白和因子，使得细胞能够以一层完整的膜状结构收集。细胞片层是近年来组织工程领域的研究热点，在皮肤、角膜、心脏及牙周等相关疾病的治疗中具有广泛的应用。
发明内容
在本公开的一个方面，提供一种细胞培养监测设备，包括：
培养容器，配置成容纳细胞片层的培养基质；和
图像采集装置，配置成获取所述培养容器内的图像，以监测所述细胞片层的生长状态。
在一些实施例中，所述培养容器的底部包括：
温敏层，配置成承载所述培养基质；和
冷光层，位于所述温敏层远离所述细胞片层的一侧，配置成通过冷光为所述细胞片层的图像采集提供背光。
在一些实施例中，在所述温敏层与所述冷光层之间还设有透光隔热层，配置成至少部分地减少所述温敏层与所述冷光层之间的热量传递。
在一些实施例中，所述冷光源位于所述冷光层的内部或所述冷光层外侧的邻接位置。
在一些实施例中，所述培养容器的底部还包括：
布线层，位于所述冷光层远离所述细胞片层的一侧。
在一些实施例中，还包括：
温度调节元件，配置成调节所述培养容器的内部温度。
在一些实施例中，在所述培养容器的外侧设有遮光结构。
在一些实施例中，所述温度调节元件包括沿所述培养容器的外壁排列的多个半导体温控元件，多个所述半导体温控元件排列形成遮光结构。
在一些实施例中，所述培养容器位于培养箱内。
在一些实施例中，在所述培养箱内设有一或多个的空间层，在所述空间层设有配置成支撑多个所述培养容器的支架，多个所述培养容器固定地或可装拆地安装在所述支架上。
在一些实施例中，在所述培养容器的外部设有一体式或可分离式的定位结构，所述定位结构与所述支架配合，并相对于所述支架可移动。
在一些实施例中，还包括：
导向机构，设置在所述培养容器的上方；和
驱动机构，配置成驱动所述图像采集装置在所述导向机构上移动，以便在多个所述培养容器之间进行图像采集位置的调整。
在一些实施例中，所述导向机构包括：平行的成对第一导轨和设置在所述成对第一导轨之间的第二导轨；所述驱动机构包括：
第一驱动机构，设置在所述成对第一导轨与所述第二导轨之间，配置成驱动所述第二导轨沿所述成对第一导轨的延伸方向相对于所述成对第一导轨运动；和
第二驱动机构，设置在所述第二导轨与所述图像采集装置之间，配置成驱动所述图像采集装置沿所述第二导轨的延伸方向相对于所述第二导轨运动。
在一些实施例中，还包括以下装置中的至少一种：
控制装置，配置成对所述培养容器的外部环境参数进行控制；
图像处理装置，配置成对所述图像采集装置获取的所述细胞片层的图像进行处理，以获得所述细胞片层的生长状态。
在一些实施例中，所述控制装置与所述图像处理装置连接，并配置成根据所述细胞片层的生长状态对所述培养容器的外部环境参数进行调整。
在本公开的另一个方面，提供一种基于前述的细胞培养监测设备的培养监测方法，包括：
将细胞片层的培养基质放入培养容器，以便在所述培养基质中生长所述细胞片层；
在所述细胞片层的成长过程中，通过所述图像采集装置获取所述培养容器内的图像，以监测所述细胞片层的生长状态。
在一些实施例中，所述细胞培养监测设备包括设置在所述培养容器的上方的导向机构和驱动机构；所述培养监测方法还包括：
控制所述驱动机构驱动所述图像采集装置在所述导向机构上移动，以调整所述图像采集装置移动到与所述培养容器对应的图像采集位置；
控制所述图像采集装置执行图像采集，并在采集完毕后控制所述驱动机构驱动所述图像采集装置移动到另一个培养容器对应的图像采集位置继续图像采集。
附图说明
构成说明书的一部分的附图描述了本公开的实施例，并且连同说明书一起用于解释本公开的原理。
参照附图，根据下面的详细描述，可以更加清楚地理解本公开，其中：
图1是根据本公开细胞培养监测设备的一些实施例的结构示意图；
图2是根据本公开细胞培养监测设备的一些实施例中培养容器在俯视角度的示意图；
图3是根据本公开细胞培养监测设备的一些实施例中培养容器的结构示意图；
图4是根据本公开细胞培养监测设备的一些实施例中培养容器的底部结构示意图；
图5是根据本公开细胞培养监测设备的另一些实施例中培养容器的底部结构示意图；
图6是根据本公开细胞培养监测设备的一些实施例中培养容器的外部结构示意图；
图7是根据本公开细胞培养监测设备的另一些实施例的结构示意图；
图8是根据本公开细胞培养监测设备的另一些实施例中定位结构与支架配合的结构示意图；
图9是根据本公开细胞培养监测设备的一些实施例中导向机构和驱动机构配合的结构示意图；
图10是根据本公开培养监测方法的一些实施例的流程示意图；
图11是根据本公开培养监测方法的另一些实施例的流程示意图。
应当明白，附图中所示出的各个部分的尺寸并不是按照实际的比例关系绘制的。此外，相同或类似的参考标号表示相同或类似的构件。
具体实施方式
现在将参照附图来详细描述本公开的各种示例性实施例。对示例性实施例的描述仅仅是说明性的，决不作为对本公开及其应用或使用的任何限制。本公开可以以许多不同的形式实现，不限于这里所述的实施例。提供这些实施例是为了使本公开透彻且完整，并且向本领域技术人员充分表达本公开的范围。应注意到：除非另外具体说明，否则在这些实施例中阐述的部件和步骤的相对布置应被解释为仅仅是示例性的，而不是作为限制。
本公开中使用的“第一”、“第二”以及类似的词语并不表示任何顺序、数量或者重要性，而只是用来区分不同的部分。“包括”或者“包含”等类似的词语意指在该词前的要素涵盖在该词后列举的要素，并不排除也涵盖其他要素的可能。“上”、“下”、“左”、“右”等仅用于表示相对位置关系，当被描述对象的绝对位置改变后，则该相对位置关系也可能相应地改变。
在本公开中，当描述到特定器件位于第一器件和第二器件之间时，在该特定器件与第一器件或第二器件之间可以存在居间器件，也可以不存在居间器件。当描述到特定器件连接其它器件时，该特定器件可以与所述其它器件直接连接而不具有居间器件，也可以不与所述其它器件直接连接而具有居间器件。
本公开使用的所有术语(包括技术术语或者科学术语)与本公开所属领域的普通技术人员理解的含义相同，除非另外特别定义。还应当理解，在诸如通用字典中定义的术语应当被解释为具有与它们在相关技术的上下文中的含义相一致的含义，而不应用理想化或极度形式化的意义来解释，除非这里明确地这样定义。
对于相关领域普通技术人员已知的技术、方法和设备可能不作详细讨论，但在适当情况下，所述技术、方法和设备应当被视为说明书的一部分。
在发明人所知的技术中，为了测量细胞培养装置中细胞的培养情况，通过在细胞培养装置中进行培养液的取样，并通过检测培养液的指标变化来间接检测细胞的培养状况。发明人经研究发现，此类相关技术在应用到细胞片层等细胞培养的监测时，没 有考虑到细胞片层的特殊性，难以对细胞片层的面积、厚度、均一程度和平整程度等指标实现有效的检测。
考虑到细胞片层等细胞培养的需求，难以将上述相关细胞培养装置的监测方案用于面积、厚度、均一程度和平整程度等中的一项或多项的监测。有鉴于此，为了满足包括细胞片层等细胞在培养过程中的状态监测要求，本公开提供了细胞培养监测设备的一些实施例的实现结构及原理。
在下述中，所称的细胞培养，包括单层或多层的细胞片层(Cell Sheet)的培养。所培养的细胞，例如可以包括从人、小鼠、大鼠、豚鼠、仓鼠、鸡、兔子、猪、绵羊、牛、马、狗、猫、猴子等中分离出的各种动物细胞和人体细胞。细胞的类型例如可以包括角化细胞、脾细胞、神经细胞、胶质细胞、胰腺β细胞、肾小球膜细胞、表皮细胞、上皮细胞(角膜上皮细胞、口腔粘膜上皮细胞、羊膜上皮细胞等)、内皮细胞(血管内皮细胞、角膜内皮细胞等)、成纤维细胞、实质细胞(肝实质细胞、角膜实质细胞等)、包括血管平滑肌细胞等平滑肌细胞的肌细胞、脂肪细胞、滑膜细胞、软骨细胞、骨细胞、造骨细胞、破骨细胞、乳腺细胞、肝细胞、骨膜来源的细胞或间质细胞，或者前述类型的细胞的前体细胞。细胞还可以包括胚胎干细胞(ESC)和间充质细胞干细胞(MSC)等干细胞或癌细胞。
在下述中，细胞的生物来源可以是同种、异种或同一个体，只要其可以在细胞水平进行培养即可。也就是说，这些细胞既可以是同种异型(Allogenic)的细胞，也可以是不同种类(Xenogenic)的细胞。这些细胞还可以是相同类型(Same Type)和不同类型(Different Type)的细胞，例如可以包括同种动物的相同种类的细胞、同种动物的不同种类的细胞、异种动物的相同种类的细胞、异种动物的不同种类的细胞、同一个体的相同种类的细胞、同一个体的不同种类的细胞等。
图1是根据本公开细胞培养监测设备一些实施例的结构示意图，结合图2中培养容器在俯视角度的示意图。本实施例的细胞培养监测设备包括：
培养容器100，配置成容纳细胞片层600的培养基质500；和
图像采集装置200，配置成获取所述培养容器100内的图像，以监测所述细胞片层600的生长状态。
在图1中，培养容器100具有能够容纳培养基质500和细胞片层600的容纳空间，并为细胞片层600提供生长空间。培养基质500能够为细胞片层600提供营养物质，以使其能够从单细胞或细胞集落生长成具有一定面积和厚度的细胞片层600。
容易理解，根据所培养的细胞的差异，可相应的调整或改变培养基质的组成和培养条件。例如，可以根据冷泉港试验设计手册(CSH Protocols)的相关指导选择和设计合适的培养基质和培养条件。
图像采集装置200可采用各类可实现拍摄成像的摄像元件，例如基于CCD或CMOS成像的摄像头、基于红外成像或红外热成像等成像仪等。图像采集装置200能够按照预设的时间间隔、按照指令或实时地对培养容器100内的图像进行获取。图像采集装置200获取到的图像能够反映出细胞片层600在不同生长阶段的生长状况。并且，通过对图像的分析和处理能够进一步获得细胞片层600的重要指标数据，例如面积、厚度、均一程度和平整程度等，从而满足细胞片层培养过程中的状态监测要求。当监测到细胞片层600达到要求的面积等指标时，可以通过收集工具及时对细胞片层600进行收集。
举例来说，在获得图2所示的培养容器100内细胞片层600的图像时，测量图像中细胞片层600的覆盖面积。又例如，通过测量图像中细胞片层600的透光度与预设标准值的比对来确定细胞片层600的厚度。或者，通过测量图像中细胞片层600的热分布图和/或红外图来确定细胞片层600的均一程度和/或平整程度。
图3是根据本公开细胞培养监测设备一些实施例中培养容器的结构示意图。在图3中，培养容器100包括培养皿体110和皿盖120，皿盖120可以覆盖在培养皿体110的顶部，实现培养皿体110内部空间的封闭。培养皿体110的内壁和底部围成了培养基质500和细胞片层600的容纳空间。
在培养容器100的外部可以设置一体式或可分离式的定位结构130。参考图3，在一些实施例中，定位结构130可以固定设置在培养皿体110的外壁外侧，其作用是方便培养皿体110的位置设置和调整。在另一些实施例中，为了方便定位结构130相对于导向装置(例如导轨等)的位置调整，可以在定位结构130上设置与导向装置配合的凹入结构131。
对于细胞片层的收获和采集来说，相关技术中常用温敏材料作为基底层(即温敏层)来承载细胞片层，此类材料例如包括聚N-异丙基丙烯酰胺(PIPAAm)、PIPAAm与甲基丙烯酸的复合物、赖氨酸短肽A6K等。温敏材料对温度非常敏感，当温敏材料所感受到的温度达到特定条件时，例如升温/降温到特定温度时，细胞片层从紧贴温敏材料的状态转变成容易脱离的状态，从而方便细胞片层的收获和采集。
在发明人所知的技术中，有些细胞培养监测方案利用测量单元的受光部来接收发 光部透过细胞培养装置中的培养液后的光线，并根据接收光线来确定细胞的培养情况。当将此类细胞培养监测方案应用到细胞片层的培养监测时，发光部所使用的光源的热量可能造成局部过热而损伤细胞片层，并且可能会对细胞片层所附着的温敏材料造成不利影响，从而使细胞片层的监测变得十分困难。
为了使图像采集装置200进行图像采集时获得良好质量的图像，并且减少对温敏材料的不利影响，参考图4所示的培养容器的底部结构，培养容器100的底部包括温敏层140和冷光层170，冷光层170位于所述温敏层140远离所述细胞片层600的一侧。温敏层140配置成承载所述培养基质500。冷光层170配置成通过冷光(例如基于冷光源160发出的冷光)为所述细胞片层600的图像采集提供背光，来确保图像采集的质量。
由于冷光层170所提供的背光为冷光(Luminescence)，冷光的发光过程不产生显著的热量，例如荧光、磷光、菌光等，因此发热量很小，对温敏层140的温度影响较小，也不容易造成局部过热。
可选地，冷光层170内可设有光色散颗粒，能够将冷光源160发出的冷光散射到整个冷光层170中，从而提供更加均匀的背光效果，进而提升采集图像的质量。
可选地，冷光源160可基于光致发光原理、阴极射线发光原理或者高能粒子发光原理来产生冷光。在本公开的一些实施例中，冷光源160可采用冷光发光二极管、冷光线、荧光板或冷光片等。
参考图4，在一些实施例中，冷光源160可设置在冷光层170外侧的邻接位置。例如，在培养容器100的外部设有一体式或可分离式的定位结构130的实施例中，冷光源160可设置在定位结构130上靠近冷光层170外侧的位置。将冷光源160设置在冷光层170的外侧，能够进一步减少冷光源160的发热量对温敏层140的影响。
冷光源160的设置位置不限于此。图5是根据本公开细胞培养监测设备另一些实施例中培养容器的底部结构示意图。参考图5，冷光源160可以设置在冷光层170的内部，以使培养容器100的结构更加紧凑。在一些实施例中，冷光层170内部可设有单一的冷光源160。在另一些实施例中，冷光层170内部可设有多个颗粒式或长条式冷光源160，并随机布置或按照预设阵列布置。
参考图4和图5，在一些实施例中，温敏层140与所述冷光层170之间还设有透光隔热层150。透光隔热层150可配置成至少部分地减少所述温敏层140与所述冷光层170之间的热量传递，以便进一步降低冷光层170的热量对温敏层140的不利影响。 透光隔热层150可由至少部分透过冷光层170的冷光的隔热材料制成，例如能够透光的玻璃或塑料等。
仍参考图4和图5，在一些实施例中，培养容器100的底部还包括布线层180。该布线层180位于所述冷光层170远离所述细胞片层600的一侧。通过将布线层180设置在培养容器100的底部，能够使培养容器100的结构更加紧凑，也方便了外接线路的布置。
图6是根据本公开细胞培养监测设备一些实施例中培养容器的外部结构示意图。在图6中，培养容器100的外壁紧密排列了多个半导体温控元件190。这些半导体温控元件190可作为温度调节元件对培养容器100的内部温度进行调节，以实现细胞片层600培养过程中更精细化的温度控制，从而确保细胞片层600的稳定生长。另外，温度调节元件还能够使温敏层140在细胞片层600的生长过程中保持稳定。当需要进行细胞片层600的采集时，温度调节元件还可以将培养容器100的内部温度调整到温敏层140易于与细胞片层600相脱离的温度范围，以降低细胞片层600的采集难度。
温度调节元件不仅限于包括图6所示的半导体温控元件190。在另一些实施例中，温度调节元件还可以包括电子式温控器或者蒸汽式温控器等。在设置位置方面，一些实施例中的温度调节元件可以设置在培养容器100的内部，或者设置在培养容器100的外侧，但不与培养容器100的外壁紧密贴合等。
为了防止冷光外泄或者外面的光透入培养容器而影响到采集的图像质量，在一些实施例中，培养容器100的外侧设有遮光结构。例如，图6中示出沿所述培养容器100的外壁排列的多个半导体温控元件190，这些半导体温控元件190能够通过排列形成遮光结构。由于半导体材料相对于可见光是非透明的，因此能够形成遮光结构，从而既实现更加快速的温度调节作用，又能够限制内部冷光的外泄或外部光的透入。
图7是根据本公开细胞培养监测设备另一些实施例的结构示意图。在图7所示的另一些实施例中，培养容器100设置在培养箱300内。培养箱300能够提供适宜细胞片层600生长的外部环境，增加细胞片层600生长的外部环境的可控性和可靠性。培养箱300在布置、设置以及使用等多个方面都非常方便，有利于操作人员的使用。
参考图7，在一些实施例中，在培养箱300内设有一或多个空间层。在所述空间层设有配置成支撑多个所述培养容器100的支架310，多个所述培养容器100固定地或可装拆地安装在所述支架310上，这样就满足了批量的细胞片层的培养和监测要求，从而有利于实现细胞片层的大规模和工业化的培养，降低细胞片层的生产成本。
为了实现培养容器100的外部环境参数(例如培养箱300的工作参数设置，如温度、工作时间、工作参数、开启和关闭等)的控制，在另一些实施例中可以包括可对所述培养容器100的外部环境参数进行控制的控制装置。例如，通过图7中培养箱300上的操作面板330接收外部环境参数的输入，并通过显示屏320进行相关参数和监测数据的显示。
为了实现细胞片层600的图像的处理，在另一些实施例中还可以包括图像处理装置。图像处理装置可配置成对所述图像采集装置200获取的所述细胞片层600的图像进行处理，以获得所述细胞片层600的生长状态。在一些实施例中，控制装置与所述图像处理装置连接，并配置成根据所述细胞片层的生长状态对所述培养容器的外部环境参数进行调整。例如在图7中，计算机400与培养箱300中的图像采集装置200连接，并对图像采集装置200获取的所述细胞片层600的图像进行处理，以获得细胞片层600的各项重要指标数据，例如面积、厚度、均一程度和平整程度等。
例如，控制装置、图像处理装置等可以通过中央处理器(CPU)或者现场可编程逻辑阵列(FPGA)或者单片机(MCU)或者数字信号处理器(DSP)或者专用集成电路(ASIC)等具有数据处理能力和/或程序执行能力的处理器实现。
例如，控制装置、图像处理装置可以集成在同一处理器中，或分别由不同的处理器实现。
例如，控制装置可以连接图像处理装置。
例如，连接可以通过包括无线网络、有线网络、和/或无线网络和有线网络的任意组合等实现。网络可以包括局域网、互联网、电信网、基于互联网和/或电信网的物联网、和/或以上网络的任意组合等。有线网络例如可以采用双绞线、同轴电缆或光纤传输等方式进行通信，无线网络例如可以采用移动通信网络、蓝牙、Zigbee或者Wi-Fi等通信方式。
图8是根据本公开细胞培养监测设备另一些实施例中定位结构与支架配合的结构示意图。参考图3和图8，在一些实施例中，培养容器100的外部设有一体式或可分离式的定位结构130，所述定位结构130与所述支架310配合，并相对于所述支架310可移动。图8中示出的细长杆形式的支架310除了可以实现对培养容器100的支撑作用之外，还能够与定位结构130的凹入结构131进行导向配合，从而使培养容器100在支架310上的布置和位置调整更加便利。
图7和图8分别示出了一些实施例中培养箱300内的多层和多组的培养容器100 的布置形式。在培养箱300的每层都可以设置一个或多个图像采集装置200。图像采集装置200可以运动到不同培养容器100对应的图像采集位置进行图像采集操作。而在另一些实施例中，图像采集装置200也可以设置在培养箱300之外，以简化培养箱300的构成，减小培养箱300的占用空间。
在一些实施例中，图像采集装置200的图像采集位置可以由操作人员手动调整。而在另一些实施例中，也可以采用自动化的调整手段，以节省人力，提高效率。图9是根据本公开细胞培养监测设备一些实施例中导向机构和驱动机构配合的结构示意图。参考图9，在一些实施例中，细胞培养监测设备还包括：导向机构700和驱动机构800。导向机构700设置在所述培养容器100的上方，而驱动机构800配置成驱动所述图像采集装置200在所述导向机构700上移动，从而方便地实现了图像采集装置200在多个所述培养容器100之间进行图像采集位置的调整。对于多个所述培养容器100来说，这些实例允许使用更少数量的图像采集装置以位置调整的方式在不同时段进行图像采集，从而在满足监测要求的同时，减少图像采集装置的使用，降低产品成本和能耗。
图9示出了简便且易于实现的导向机构和驱动机构的配合结构实例。在图9中，导向机构700包括：平行的成对第一导轨710和设置在所述成对第一导轨710之间的第二导轨720。驱动机构800包括：第一驱动机构810和第二驱动机构820。其中，第一驱动机构810设置在所述成对第一导轨710与所述第二导轨720之间，配置成驱动所述第二导轨720沿所述成对第一导轨710的延伸方向相对于所述成对第一导轨710运动。第二驱动机构820设置在所述第二导轨720与所述图像采集装置200之间，配置成驱动所述图像采集装置200沿所述第二导轨720的延伸方向相对于所述第二导轨720运动。第一驱动机构810和第二驱动机构820可采用各种实现驱动作用的动力元件(例如电机、气泵或液压泵等)，并根据需要选用传动结构(例如齿轮齿条传动结构、齿轮组传动结构或滑轮组传动机构等)。
按照图像采集的要求，当需要对某个培养容器100进行图像采集时，可控制驱动机构800驱动图像采集装置200在导向机构700上移动，以使图像采集装置200移动到该培养容器100对应的图像采集位置。举例来说，第一驱动机构810可以先驱动第二导轨720沿成对第一导轨710移动到该培养容器100所在列的上方，第二驱动机构820再驱动图像采集装置200移动到该培养容器100的上方，该位置为该培养容器100对应的图像采集位置。在其他例子中，第一驱动机构810和第二驱动机构820的驱动 步骤可以调换或者同时进行。
当调整到位后，可控制所述图像采集装置200执行图像采集，并在采集完毕后控制所述驱动机构800驱动所述图像采集装置200移动到另一个培养容器100对应的图像采集位置继续图像采集。
前面已对本公开的细胞培养监测设备的各实施例进行了说明。而基于上述任一细胞培养监测设备的实施例，本公开还提供了对应的培养检测方法实施例。如图10所示，为根据本公开培养监测方法的一些实施例的流程示意图。在本实施例中，培养监测方法可包括：
步骤S1、将细胞片层600的培养基质500放入培养容器100，以便在所述培养基质500中生长所述细胞片层600；
步骤S2、在所述细胞片层600的成长过程中，通过所述图像采集装置200获取所述培养容器100内的图像，以监测所述细胞片层600的生长状态。
在上述方法实施例中，可由操作人员手动或通过工具将培养基质500以及作为细胞片层600生长基础的单细胞或细胞集落置入到培养容器100内，然后在细胞片层600的生长过程中，由操作人员手动操作或者由控制装置进行自动控制图像采集装置200来获取所述培养容器100内的图像。
参考图11，在另一些细胞培养监测设备实施例中，还可以包括设置在所述培养容器100的上方的导向机构700和驱动机构800。
对应的培养监测方法还可包括：
步骤S3、控制所述驱动机构800驱动所述图像采集装置200在所述导向机构700上移动，以调整所述图像采集装置200移动到与培养容器100对应的图像采集位置；
步骤S4、控制所述图像采集装置200执行图像采集，并在采集完毕后控制所述驱动机构800驱动所述图像采集装置200移动到另一个培养容器100对应的图像采集位置继续图像采集。
在本实施例中，驱动机构800的驱动操作和图像采集装置200的图像采集操作可由单一或不同的控制装置进行控制，控制装置可以采用运行有控制程序的通用或专用计算设备实现。步骤S3和步骤S4可作为前述方法实施例的步骤S2的具体化，也可独立于该步骤S2执行。
本说明书中多个实施例采用递进的方式描述，各实施例的重点有所不同，而各个实施例之间相同或相似的部分相互参见即可。对于方法实施例而言，由于其整体以及 涉及的步骤与系统实施例中的内容存在对应关系，因此描述的比较简单，相关之处参见系统实施例的部分说明即可。
至此，已经详细描述了本公开的各实施例。为了避免遮蔽本公开的构思，没有描述本领域所公知的一些细节。本领域技术人员根据上面的描述，完全可以明白如何实施这里公开的技术方案。
虽然已经通过示例对本公开的一些特定实施例进行了详细说明，但是本领域的技术人员应该理解，以上示例仅是为了进行说明，而不是为了限制本公开的范围。本领域的技术人员应该理解，可在不脱离本公开的范围和精神的情况下，对以上实施例进行修改或者对部分技术特征进行等同替换。本公开的范围由所附权利要求来限定。

Claims (17)

  1. 一种细胞培养监测设备，包括：
    培养容器，配置成容纳细胞片层的培养基质；和
    图像采集装置，配置成获取所述培养容器内的图像，以监测所述细胞片层的生长状态。
  2. 根据权利要求1所述的细胞培养监测设备，其中，所述培养容器的底部包括：
    温敏层，配置成承载所述培养基质；和
    冷光层，位于所述温敏层远离所述细胞片层的一侧，配置成通过冷光为所述细胞片层的图像采集提供背光。
  3. 根据权利要求2所述的细胞培养监测设备，其中，在所述温敏层与所述冷光层之间还设有透光隔热层，配置成至少部分地减少所述温敏层与所述冷光层之间的热量传递。
  4. 根据权利要求2所述的细胞培养监测设备，其中，所述冷光源位于所述冷光层的内部或所述冷光层外侧的邻接位置。
  5. 根据权利要求2所述的细胞培养监测设备，其中，所述培养容器的底部还包括：
    布线层，位于所述冷光层远离所述细胞片层的一侧。
  6. 根据权利要求1所述的细胞培养监测设备，还包括：
    温度调节元件，用于调节所述培养容器的内部温度。
  7. 根据权利要求2所述的细胞培养监测设备，其中，在所述培养容器的外侧设有遮光结构。
  8. 根据权利要求6所述的细胞培养监测设备，其中，所述温度调节元件包括沿所述培养容器的外壁排列的多个半导体温控元件，多个所述半导体温控元件排列形成遮光结构。
  9. 根据权利要求1所述的细胞培养监测设备，其中，所述培养容器位于培养箱内。
  10. 根据权利要求9所述的细胞培养监测设备，其中，在所述培养箱内设有一或多个的空间层，在所述空间层设有配置成支撑多个所述培养容器的支架，多个所述培养容器固定地或可装拆地安装在所述支架上。
  11. 根据权利要求10所述的细胞培养监测设备，其中，在所述培养容器的外部设有一体式或可分离式的定位结构，所述定位结构与所述支架配合，并相对于所述支架可移动。
  12. 根据权利要求1所述的细胞培养监测设备，还包括：
    导向机构，设置在所述培养容器的上方；和
    驱动机构，配置成驱动所述图像采集装置在所述导向机构上移动，以便在多个所述培养容器之间进行图像采集位置的调整。
  13. 根据权利要求12所述的细胞培养监测设备，其中，所述导向机构包括：平行的成对第一导轨和设置在所述成对第一导轨之间的第二导轨；所述驱动机构包括：
    第一驱动机构，设置在所述成对第一导轨与所述第二导轨之间，配置成驱动所述第二导轨沿所述成对第一导轨的延伸方向相对于所述成对第一导轨运动；和
    第二驱动机构，设置在所述第二导轨与所述图像采集装置之间，配置成驱动所述图像采集装置沿所述第二导轨的延伸方向相对于所述第二导轨运动。
  14. 根据权利要求1所述的细胞培养监测设备，还包括以下装置中的至少一种：
    控制装置，配置成对所述培养容器的外部环境参数进行控制；
    图像处理装置，配置成对所述图像采集装置获取的所述细胞片层的图像进行处理，以获得所述细胞片层的生长状态。
  15. 根据权利要求14所述的细胞培养监测设备，其中，所述控制装置与所述图像处理装置连接，并配置成根据所述细胞片层的生长状态对所述培养容器的外部环境参数进行调整。
  16. 一种基于权利要求1～15任一所述的细胞培养监测设备的培养监测方法，包括：
    将细胞片层的培养基质放入培养容器，以便在所述培养基质中生长所述细胞片层；
    在所述细胞片层的成长过程中，通过所述图像采集装置获取所述培养容器内的图像，以监测所述细胞片层的生长状态。
  17. 根据权利要求16所述的培养监测方法，其中，所述细胞培养监测设备包括设置在所述培养容器的上方的导向机构和驱动机构；所述培养监测方法还包括：
    控制所述驱动机构驱动所述图像采集装置在所述导向机构上移动，以调整所述图像采集装置移动到与所述培养容器对应的图像采集位置；
    控制所述图像采集装置执行图像采集，并在采集完毕后控制所述驱动机构驱动所述图像采集装置移动到另一个培养容器对应的图像采集位置继续图像采集。
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