JP2013039113A - Method for controlling cell quality and method for producing cell - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a more practical method for controlling cell quality and a method for producing cells.SOLUTION: The method for controlling cell quality includes a step in which in the control of cell quality, a plurality of pieces of cell movement information in a division in different periods based on cell displacement information related to one or more cell displacements in a prescribed period of at least every two divisions of a culture region virtually divided into a plurality of parts of one cell group are acquired and accumulated cell movement information in a division concerning the every divisions is acquired. Then, quality of the cell group is evaluated based on the accumulated cell movement information in a division.

Description

  The present invention relates to a cell quality control method and a cell production method.

  In recent regenerative medicine, cellular medicine, especially stem cell therapy, has attracted attention. In cell therapy, its effectiveness largely depends on the quality of cells such as the degree of differentiation and activity. Currently, many of the various cell culture processes, including stem cells, rely on skilled workers for their management. That is, the simple and clear standard of cell quality control has not yet been provided, and is usually performed by a combination of a plurality of basic techniques. It was also difficult to evaluate cells non-destructively. Therefore, it is difficult to efficiently and stably provide cells with appropriate quality in a timely manner.

  Thus, for example, a method for predicting cell quality using non-destructive cell shape information obtained from cell images has been proposed (Patent Document 1).

JP 2009-44974 A

  In order to accurately acquire cell morphology information, it is usually necessary to perform complicated operations such as detecting the contour of the cell, and there is still difficulty in practical use. In addition, cell shape information is difficult to recognize unless the cell density is low to some extent. Therefore, in the method using the cell shape information, the culture state is limited, for example, it is necessary to be less than sub-conflent (confluence (cell culture density of about 80% of the state where the cells are grown on the entire culture surface)) or less. .

  On the other hand, cell migration is thought to be a highly controlled cellular activity. Therefore, there is a possibility that the amount of movement of cells and the movement speed can be used as an index. However, since cells exhibit completely different behavior depending on their observation position and time, it is difficult to capture dynamic behavior changes such as the amount and speed of cell movement, and it is also difficult to quantify them.

  As described above, in order to promote the practical application of regenerative medicine, development of engineering technology that supports work related to cell culture and the like is strongly demanded. Accordingly, an object of the present disclosure is to provide a more practical cell quality control method and cell production method. More specifically, an object is to provide a cell quality control method and the like that can efficiently and quantitatively evaluate quality.

  The present inventors pay attention to the movement information such as the movement distance of the cell obtained from the image of the cell, and by acquiring the distribution of the movement information accumulated under a certain condition, the quality such as the degree of differentiation of the cell is obtained. We obtained the knowledge that it can be obtained quantitatively and early regardless of the cell density. The disclosure herein is provided based on these findings.

(1) A cell quality control method,
Different cell movement information in the compartment based on the cell displacement information relating to the displacement of one or more cells in a predetermined period of at least two of the sections of the culture region virtually divided into a plurality of one or two or more cell populations Obtaining a plurality of per-period, and obtaining the intra-compartment cumulative cell migration information for each of the compartments,
A quality control method for evaluating the quality of the one or more cell populations based on the intra-compartment cumulative cell migration information.
(2) The quality control method according to (1), wherein the step of acquiring the intra-compartment cumulative cell migration information is performed using image information acquired for the culture region of the one or more cell populations.
(3) The cell displacement information includes start point position information of the one or more cells existing in the section at the start time of the predetermined period, and end point position information of the cells at the end time of the predetermined period. The quality control method according to (1) or (2).
(4) The following (a) to (d):
(A) obtaining two pieces of image information relating to a cell culture state at a time t _n (n is an integer of 1 or more and N or less) and a time t _{n + 1 for} one or more cell populations;
(B) Based on the position information of one or more cells at time t _n of each section in the two image information and the position information of these cells at time t _{n + 1} , the one or more cells Obtaining the cell displacement information about the cell, and obtaining the intra-compartment cell migration information of each of the compartments at t _n to t _{n + 1} based on the cell displacement information,
(C) Repeat (a) and (b) until t = N,
(D) The intra-compartment cumulative cell movement information for each of the sections is acquired based on the N-1 pieces of intra-partition cell movement information acquired for each of the sections, according to any one of (1) to (3) Quality control method.
(5) The quality control method according to any one of (1) to (4), wherein the displacement of the cell is acquired based on a displacement of the center of gravity of the cell.
(6) The quality control method according to any one of (1) to (5), wherein the intra-compartment cumulative cell migration information is evaluated by making a two-dimensional distribution diagram assigned to the corresponding compartment on the culture region.
(7) The quality control method according to any one of (1) to (6), wherein the intra-compartment cumulative cell migration information is evaluated in the form of a histogram.
(8) The quality control method according to any one of (1) to (7), wherein the quality of the one or more cell populations is evaluated for each of the sections.
(9) The quality control method according to any one of (1) to (8), wherein the quality is a cell differentiation degree.
(10) A method for producing a cell population,
A cell population production step of culturing cells originating from the cell population to obtain the cell population;
With
In the production process, intra-compartment cell movement information based on cell displacement information related to displacement of one or more cells in a predetermined period in at least two of the culture regions of the cell population virtually divided into a plurality of sections. A production method for obtaining a plurality of intra-compartment cumulative cell migration information for each of the compartments, and performing quality evaluation of the cell population based on the intra-compartment cumulative cell migration information.
(11) The production method according to (10), wherein in the cell population production step, cells are selectively cultured based on the intra-compartment cumulative cell migration information.
(12) The production method according to (10) or (11), wherein the cell population production step is performed under conditions for differentiating the origin cells into a desired differentiation state.
(13) A cell inspection method,
Different intra-compartment cell movement information based on cell displacement information regarding the displacement of one or more cells in a predetermined period of at least two of each of the culture regions of the cell population derived from the cells virtually divided into a plurality of cells Obtaining a plurality of per-period, and obtaining the intra-compartment cumulative cell migration information for each of the compartments,
A test method for testing the quality of the cell based on the intra-compartment cumulative cell migration information.

It is a figure which shows the outline | summary of the method of quality control of this indication. It is a figure which shows the integration effect of speed. It is a figure which shows a heat-resistant candidate mutant and its heat-resistant temperature. It is a figure which shows the cumulative effect of "the displacement of the cell which exists in a division for every one period." It is a figure which shows the effect of utilizing "the cell displacement obtained in the virtual division." It is a figure which shows an example of the flow for implementing this quality control method. It is a figure which shows the relationship between integration time and the coefficient of variation (CV) of the integration speed in each integration time. It is a figure which shows the relationship between the presence or absence of the integration | accumulation of the speed | rate in each culture time, and skeletal differentiation conditions. It is a figure which shows the relationship (for every cell line) between the integration effect of speed | velocity | rate, and bone differentiation conditions. It is a figure which shows the relationship (with the difference in a passage number) between the integration | accumulation effect of a speed | rate, and bone differentiation conditions. The relationship between the speed integration effect (average value of the integrated speed at 40 h) and ALP activity is shown. It is a figure which shows the image of the analysis object visual field of RPE cell derived from human iPS. It is a figure which shows the relationship (histogram) of the maturity which used the melanin accumulation | storage of a human iPS cell origin RPE cell as one parameter | index, and an integration map. It is a figure which shows the relationship (scatter diagram) of the maturity which made the melanin accumulation | storage of a human iPS cell origin RPE cell one parameter | index, and an integration map. It is a figure which shows the relationship (histogram) of the maturity (luminance histogram) which used the melanin accumulation | storage of a human iPS cell origin RPE cell as one parameter | index, and a shape / velocity. It is a figure which shows the relationship (median speed) of the maturity (pixel) and shape / speed which made melanin accumulation | storage of a human iPS cell origin RPE cell as one parameter | index.

  The disclosure of the present specification relates to a cell quality control method, a cell population production method, and the like. According to the quality control method according to the present disclosure, as shown in FIG. 1, in a plurality of sections assumed in the culture region of the cell population, a plurality of intra-compartment cell movement information for a plurality of periods is obtained, Accumulated cell movement information within the compartment is obtained by, for example, integration. Using this intra-compartment cumulative cell migration information, the quality of each cell population, each region, or even the entire cell population can be evaluated efficiently and quantitatively.

  One advantage of this quality control method is based on the “accumulation” (accumulation) of cell displacement over a plurality of periods in the cell population culturing process, so that instability of the evaluation method can be suppressed or avoided. In other words, since it is not a single point of time or a single time in the culturing process, a decrease in measurement accuracy due to the fact that the short measurement time reflects variations in the dynamic behavior of cells is suppressed or avoided. . Further, since it is not continuous for a long time, a decrease in measurement accuracy due to an error or the like is suppressed or avoided. Furthermore, as a result, as shown in FIG. 2, the quality of the cell population can be evaluated at an early stage.

  Another advantage is that this quality control method uses the accumulation of “displacement of cells existing in the compartment for each period” as shown in the upper part of FIG. As shown, it is not limited to tracking a specific cell over a plurality of periods. If one period is set to be sufficiently shorter than the cell division time and not more than the time to pass through the compartment with respect to the cell mobility, cells due to cell division, fusion, or cell migration It is possible to solve all the difficult points of the cell tracking technique caused by crossing, disappearance from the screen, etc., and to ensure good tracking accuracy, and as a result, the accuracy of the intra-compartment cumulative cell movement information is improved. In addition, the simplicity and efficiency of operation can be improved. Note that the use of cell displacement by tracking specific cells is not excluded.

  Another advantage is that it uses the “cell displacement obtained in the virtual section” of the culture area of the cell population, so that it is possible to control the quality of individual sections and the quality of the entire cell population. Management is also possible. Moreover, as shown in FIG. 4, by making the information for each section into a two-dimensional distribution map (map), it is possible to clarify the local difference in quality in the culture region, and at the same time, it becomes easy to grasp the properties of the entire cell population. Moreover, by making the information for each section into a histogram, quantitative comparison can be facilitated, and evaluation of the entire cell population can be facilitated from the form of the histogram.

  Furthermore, another advantage is that since the displacement of the cells in a plurality of periods is extracted and accumulated independently for each section and each period, a decrease in accuracy due to the obtained information measurement method is suppressed or reduced. It is avoided, the cumulative cell migration information in the compartment can be quantified, and its quantitative property is good. As a result, comparison for each section and comparison as a cell population are facilitated. Quantitative effects are extremely effective in the above-described two-dimensional distribution chart and histogram.

  In addition, another advantage is that the quality control method can avoid the detection of cell morphology in order to use the displacement of cells, thereby controlling the quality of cells regardless of the cell culture state. It can be done.

  Advantages corresponding to the above-described quality control method can also be obtained in the cell population production method and program according to another embodiment of the present disclosure.

  In the present disclosure, the “cell” is not particularly limited. For example, various cells of mammals (human, monkey, cow, horse, rabbit, mouse, rat, guinea pig, hamster, etc.), such as cardiomyocytes, smooth muscle cells, adipocytes, fibroblasts, bone cells, chondrocytes, Bone cells, parenchymal cells, epidermal keratinocytes, keratinocytes, epithelial cells (skin epidermal cells, corneal epithelial cells, conjunctival epithelial cells, oral mucosal epithelium, hair follicle epithelial cells, oral mucosal epithelial cells, airway mucosal epithelial cells, intestinal mucosa Epithelial cells, etc.), endothelial cells (corneal endothelial cells, vascular endothelial cells, etc.), neurons, glial cells, spleen cells, pancreatic β cells, mesangial cells, Langerhans cells, hepatocytes, or progenitor cells thereof, or mesenchymal system Stem cells (MSC), embryonic stem cells (ES cells), embryonic germ cells (EG cells), adult stem cells or induced pluripotent stem cells (iPS cells) can be used . In addition to normal cells, cancer cells such as cancer cells, or established cells such as HeLa cells, CHO cells, Vero cells, HEK293 cells, HepG2 cells, Cos-7 cells, NIH3T3 cells, Sf9 cells, etc. Etc. can be adopted. Adhesive cells such as epithelial cells, mesenchymal cells, and iPS cells are useful for the present disclosure in that their two-dimensional image analysis is easy.

  In the present disclosure, “cell quality” or “cell quality” refers to the growth rate, the remaining division potential, the remaining number of possible divisions (the number of remaining doublings), the degree of differentiation, the activity, and the ability to produce specific factors and proteins. , Abnormality degree, tumor degree, cancer degree and the like. Cell movement is largely related to these “cell quality” or “cell quality” cell states, and the cell displacement that characterizes cell movement can evaluate or manage these various cell states. . For example, regarding the degree of differentiation and activity of the cells, the present inventors have confirmed that when the displacement of the cells is small, the degree of differentiation is advanced and the degree of activity is high.

  In the present disclosure, “cell differentiation degree” refers to the differentiation of cells that have been scheduled to differentiate during culture (eg, differentiation from progenitor cells to mature cells, differentiation from multipotent stem cells to specific cell lineages, etc.). It is an index indicating whether or not it is in a stage. In order to determine the degree of differentiation, cell morphology, presence or absence or expression level of a cell surface marker (protein), presence or absence or production amount of a specific factor are generally used.

  In the present disclosure, the culture region of one or two or more cell populations is virtually divided into a plurality. “Virtually divided into multiple areas” does not mean that the culture area is physically partitioned by grooves or barriers, but is divided only to classify a plurality of cells in the culture area according to their positions. Means that. Therefore, the “virtual section” may be a section formed so as to be visible below the culture area by an identifiable grid or the like applied to the transparent culture substrate. Moreover, the division which permeate | transmits and visually recognizes a culture | cultivation area | region with the transparent sheet-like body in which the grid etc. which were piled up on the culture | cultivation area | region was formed may be sufficient. Furthermore, it may be a section formed by an image such as a grid attached to the image information acquired for the culture region.

  In the present disclosure, the size of the “compartment” is preferably such that about 1 to 2 cells are fractionated. This is because when a larger number of cells are fractionated, it becomes difficult to detect the displacement of the cells in a predetermined period, and if the cells are not fractionated, a section without information is generated. The size of the compartment depends on the cell type, but it is usually preferable that the actual size is 10 μm or more and 100 μm or less. This is because if it is less than 10 μm, the number of compartments in which cells are not fractionated increases, and if it exceeds 100 μm, three or more cells are fractionated in the compartment. Since the compartments need to be optimized according to the progress and movement speed of the target cells, in order to accurately evaluate the target cells, based on the observation results of the culture region of the cell population to be evaluated in advance It is preferable to set. A square shape, a rectangular shape, or a circular shape having a radius having such a reference dimension as one piece can be used, but a square shape is preferable.

  In the present disclosure, the number of “compartments” is not particularly limited, but it is desirable that the number of compartments is about 200 or more in the culture image when all the compartments are totaled.

  In the present specification, the “image information” is not particularly limited to the recording medium and the recording form. Preferably, it is in the form of a known image file that can be recorded on a storage medium applicable to a computer.

  In the present disclosure, “displacement of a cell” means a movement amount obtained as a result of a cell moving within a predetermined period. That is, “cell displacement” means the amount of movement obtained from the start position of the cell at the start time within the predetermined period and the end position of the cell at the end time of the predetermined period, and the movement of the cell over time within the predetermined period. And the amount of movement based on the distance. Note that cell displacement can be determined in various two-dimensional manners, such as the center of gravity of the cell, the outline of the cell, the appropriate number of points on the outline of the cell, internal organs such as the nucleus, and other detectable features (such as brightness and fluorescence) Can be acquired based on the displacement of various feature points.

  In addition to the above-described cell movement amount, the “cell displacement information” regarding the cell displacement may be a speed obtained by dividing the cell movement amount by the time when the movement amount is generated. By adopting the speed as the cell displacement information, it is possible to obtain data that does not depend on the period for obtaining the cell displacement.

  In the present disclosure, “intra-compartment cell movement information” may be information based on cell displacement information. For example, the corresponding cell displacement information is recalculated as an average or a statistical quantity (median, standard deviation, etc.). It may be what you did.

  In the present disclosure, the “intra-compartment cumulative cell migration information” may be based on a plurality of intra-compartment cell migration information. For example, the plurality of intra-compartment cell migration information may be averaged or statistical quantities (median or standard). The deviation may be recalculated.

  Hereinafter, embodiments of the present disclosure will be described in detail.

(Cell quality control method)
The quality control method is preferably applied in the cell population culturing step. In the cultivation process of a cell population, a single or plural cells are used at the start of the cultivation process, and a part of a tissue or organ collected from a living body may be used as a cell. As a process of starting culture and finally obtaining a cell population based on these. Note that the cell population culturing step may involve cell proliferation.

  Hereinafter, an acquisition process of intra-compartment cumulative cell movement information in this quality control method will be described, and an embodiment for performing quality evaluation of a cell population based on the obtained intra-compartment cumulative cell movement information will be described. FIG. 5 shows an example of a flow for carrying out this quality control method.

  In an embodiment described below, a cell culture device, an imaging device for obtaining image information of a culture region, a CPU such as a computer connected to the imaging device, a control device including a storage medium, and a display are provided. Cell quality control system. In the following steps, variously acquired information is appropriately stored in various types of storage media such as a temporary recording medium and a hard disk provided in a computer or the like, even if not specifically mentioned.

(Intra-compartment cumulative cell migration information acquisition process)
In this step, the cells prepared in advance are carried out with or after the culturing step. The culturing step has various forms depending on the purpose of culturing and is not particularly limited. For example, culture conditions according to the purpose such as quality evaluation of collected cells, quality evaluation of cultured cells, quality evaluation of cells used for regenerative medicine, inspection of collected or cultured cells, and the like are set. Those skilled in the art can appropriately set such culture conditions according to the purpose. Further, from the viewpoint of ease of detection of cell displacement, the culture region is preferably formed on a flat culture substrate.

  In the present embodiment, when time t1 is set to 0:00, six pieces of image information are acquired intermittently every 8 hours over a total of 40 hours until time t6 (N = 6), and a total of five sections are obtained. Acquire internal cell movement information. That is, the image information t1 (0 o'clock), t2 (8 o'clock), t3 (16:00), t4 (24 o'clock), and t5 (32 o'clock) are acquired.

(Step S10)
In this step, first, image information t1 is obtained for the cell culture region at time t1 (step S10). It is preferable that the culture area as a target for acquiring the image information t1 is fixed to a predetermined area in advance.

  The culture region to be imaged may be the entire cell population or a part thereof. Further, the size of the target region is not particularly limited, but it is preferable that the target region is wider. Further, the area of the culture region to be imaged can be selected in consideration of the size of each cell, cell density, and cell migration speed, rather than being defined in units of length.

  In step S10, image information t1 is acquired so as to be viewed from above the culture region so that the displacement of the cells moving along the flat culture region can be detected. The method for acquiring image information is not particularly limited as long as it can recognize the position of the cell at a certain time and simultaneously recognize the position of the movement destination of the cell after a certain period.

  The acquisition of image information is typically preferably performed by an imaging device through a microscope, and more preferably through a phase contrast microscope. The time t1 is not particularly limited, but is preferably a time that can be regarded as reaching a culture state suitable for evaluating the quality of cells in culture.

  The acquired image information t1 is stored in a computer storage medium as an image file in a predetermined form.

(Step S20)
Thereafter, the CPU confirms the elapsed time from the time t1 at regular intervals, and starts step S30 after 8 hours.

(Step S30)
When 8 hours have elapsed from time t1 and time t2 has been reached, image information t2 regarding the cell culture region at time t2 is acquired in the same manner as in step S10, and stored in the storage medium of the computer (step S20). The image information t2 is obtained by imaging the same culture area as the image information t1.

  The interval between the time t1 and the time t2 is 8 hours in this embodiment. Although the said time interval changes also with the kind of cell and culture conditions, from a viewpoint of the further improvement of accuracy, such as quality evaluation, for example, 1 to 4 hours are preferable. This is because, within this range, any adherent cell migration rate is likely to be captured in the same compartment, and cell division occurs very little on the screen. On the other hand, from the viewpoint of more efficiently and cost-effectively evaluating the quality, for example, it is preferable to set the time interval to be longer than 6 hours. This is because by increasing the time interval, the amount of image information can be limited and the processing for analysis can be made more efficient.

(Step S40)
Next, by comparing the image information t1 and the image information t2, the amount of movement of the cell that can be visually recognized by the image information t1 from the time t1 to the time t2 is acquired (step S40). Specifically, the identification information is given to all the cells visible in the image information t1, and the center of gravity of each cell is acquired as the position information t1 of each cell. The center of gravity of the cell can be detected using an image processing program such as Image J, for example.

  Then, by comparing the image information t1 and the image information t2, the same cells as the individual cells visually recognized by the image information t1 are extracted in the image information t2 and associated. For the cell associated in the image information t2, the center of gravity of the cell is acquired as the position information t2.

  With respect to the cells that are visually recognized in the image information t1 and extracted as the same cells in the image information t2, the position information t1 is subtracted from the position information t2, and the movement amount of each cell is acquired. Such an amount of movement is acquired in units of pixels in normal image processing such as Image J. Further, the moving speed is calculated by dividing this moving amount by the time interval of time t2−time t1, and cell displacement information is obtained for each individual cell.

  It should be noted that even a cell that can be visually recognized in the image information t1 is not a displacement amount acquisition target if it is not associated with the same cell in the image information t2. In addition, cells that are not associated in the image information t1 in the image information t2 are also not targeted for movement amount acquisition.

(Step S50)
Next, intra-compartment cell movement information is acquired (step S50). First, in each of the image information t1 and the image information t2, a grid image (50 pixels × 50 pixels, about 200 cell coverage number or more = approximately) for dividing the culture region to be imaged into a plurality of image information. 200 sheets). Identification information such as a number is assigned to each of the plurality of cells (sections) formed by the grid image. The details visually recognized in all the sections of the image information t1 are specified, it is determined whether the cell displacement information 1 is acquired among these cells, and the cell displacement information of the cell from which the cell displacement information is acquired Is obtained as intra-compartment cell movement information. The intra-compartment cell movement information t1 is acquired for all the compartments and stored in the storage medium. In addition, when there is no cell in the partition in the image information t1, the intra-compartment cell movement information for the partition is null.

(Steps S60 to S20 to S50)
After step S50 is completed, the CPU determines whether or not a predetermined number (in this embodiment, 5 times) of intra-compartment cell movement information has been acquired (step S60), and the predetermined number has not been completed. Further, step S20 to step S50 are repeatedly performed to acquire a predetermined number of intra-compartment cell movement information. It should be noted that step S30 is newly performed to obtain image information t3 at time t3, and then, in step S40, the image information t2 and the image information t3 are again compared, and the time t2 of the cell that can be visually recognized with the image information t2 To obtain a displacement amount from time t3 to time t3. That is, the cell identification and identification information in the image information t1 is canceled, and cell displacement information is acquired again based on the image information at the time t2 and the time t3. Thereafter, cell displacement information is obtained in the same manner. On the other hand, when a predetermined number of intra-compartment cell movement information has been acquired, step S70 is performed.

(Step S70)
When a predetermined number of intra-compartment cell movement information is acquired, all the intra-compartment cell movement information is added for each compartment based on the identification information of each compartment to obtain intra-compartment cumulative cell movement information (step S70).

(Step S80)
When the intra-compartment cumulative cell movement information is obtained for each compartment, two-dimensional distribution map information for each color is obtained based on these pieces of information (here, the sum of speeds) (step S80). More specifically, the first color information is in a section where the data value is 90% or higher, the second color information is in a section where the data value is 10% or less, and the third color information or more in other sections. Appropriate number of color information is assigned. Furthermore, two-dimensional distribution map information that can display these color information assigned to each section in the section on the grid image used for each color corresponding to the color information is acquired.

  The quality manager displays such two-dimensional distribution map information on the display, and shows the overall or local cell quality (for example, the degree of differentiation) for each section or for each area composed of a plurality of sections by color distribution. It can be evaluated efficiently and quantitatively. At the same time, the quality of the entire cell population can be evaluated efficiently and quantitatively.

  In the above-described embodiment, since the intra-compartment cumulative cell migration information is acquired using the image information acquired for the culture region, quality evaluation can be performed nondestructively. Furthermore, quality evaluation can be performed in a timely manner along with the culture process, and quality evaluation can be performed after the culture process. In addition, ex-post verification is also possible. In addition, in order to acquire the displacement information of a cell using image information, a human work may be sufficient and available programs, such as ImageJ, may be used.

  In the above-described embodiment, the process of acquiring intra-compartment cumulative cell migration information was performed only for one cell population without adopting a control cell population, but the accuracy and reproducibility of a specific cell population were improved. In order to perform a good quality evaluation, it is preferable to prepare a cell population as a control and similarly perform a step of acquiring intra-compartment cumulative cell migration information. For example, when differentiation induction is performed and quality evaluation is performed on the degree of differentiation, a group in which differentiation induction is not performed on a cell population of the same origin is preferably used as a control cell population. By adopting such a control cell population, it becomes possible to detect changes in a specific cell population with high accuracy using an assay or the like.

  In addition, the effect of the treatment can be evaluated by performing a process of acquiring the intra-compartment cumulative cell migration information for different cell populations subjected to different treatments instead of the control cell population. Furthermore, the process of obtaining the intra-compartment cumulative cell migration information of the control cell population may be combined.

  In the above-described embodiment, the cell displacement information includes the start position information such as the center of gravity of one or more cells existing in one section at the start time of the predetermined period, and the end time of the predetermined period of these cells. Obtained based on end point position information such as the center of gravity. That is, cell displacement information was obtained by performing fragmentary tracking instead of continuous cell tracking. For this reason, tracking is simple, and tracking errors can be reduced to ensure high tracking accuracy. For this reason, the accuracy and quantitativeness of the cell displacement information are also improved. It should be noted that the accuracy and quantitativeness of the cell displacement information can be improved by using a program that can perform continuous tracking of the center of gravity of the cells existing in one section in a predetermined period with a certain level of accuracy.

  In the above-described embodiment, 6 pieces of image information are acquired intermittently every 8 hours over a total of 40 hours up to time t6, where time t1 is 0:00, and a total of 5 intra-compartment cell movement information is acquired. Therefore, stable intra-compartment cumulative cell migration information is obtained, but the present invention is not limited to this. Although the image information depends on the type of cells, it is preferably a period of 10 hours or more, more preferably 20 hours or more, still more preferably 30 hours or more, even more preferably 35 hours or more, and even more preferably 40 hours or more. Preferably, it is acquired multiple times over (acquisition time). In particular, when it is 30 hours or more, stable intra-compartment cumulative cell migration information can be obtained for many cells. In addition, if the image acquisition time is preferably 30 hours or longer, more preferably 35 hours or longer, and even more preferably 40 hours or longer, roughly stable intra-compartment cumulative cell migration information can be obtained in any culture time zone. be able to. In the present embodiment, N = 6, but the present invention is not limited to this, and is appropriately set according to the type of cell, the purpose of quality control, and the like. Furthermore, in the present embodiment, image information is acquired at regular time intervals, and intra-compartment cell movement information is acquired using each image information. That is, an interval is not provided in the acquisition period of the cell movement information in each compartment, but the present invention is not limited to this. For example, as disclosed in Example 2, which will be described later, a plurality of periods with predetermined time intervals (in Example 6, the cell movement information in the compartment within 1 hour is obtained at intervals of 4 hours. May be repeated three times to obtain three pieces of intra-compartment cell movement information), and a plurality of intra-compartment cell movement information may be obtained.

  In the above-described embodiment, intra-compartment cell displacement information is acquired for all of a plurality of sections obtained by adding a grid image to image information. For this reason, cell quality can be evaluated precisely for the entire culture region. In addition, you may limit the division which acquires such displacement information within a division beforehand. This is effective when it is necessary to evaluate the quality of a wide culture area while limiting the amount of information to be acquired.

  In the above-described embodiment, the displacement of the cell is acquired based on the center of gravity of the cell. Based on the center of gravity, compared to the case where cell contour information is used, hardware error and cell contour information in the luminance image have the same form in a high-density culture (conferencing image). So it makes little sense. Compared to this, centroid coordinates are one of the basic calculation indices for image analysis that are provided as standard in any simple software, and are not much affected by image quality errors even if hardware is not shared. Can be acquired stably without. In addition, the hardware can be simplified and more efficient than the case based on the luminance information of a part of the cells. Luminance information changes greatly depending on the imaging environment, and in order to create a quantitative environment, it is essential to share hardware such as a dark room imaging environment and the unification of camera models to unify the number of pixel bits Because. It has been found that when acquiring cell displacement based on the center of gravity of the cell, it is substantially stable to an error in the center of gravity of about 25% of the cell size. Further, it is known that almost the same result is obtained by f-test or the like when the error of the center of gravity is in the range of about 12.5% of the cell size. If the particle tracking algorithm in the image such as Particle Tracking Velocimetry (PTV) is used, the center of gravity detection and displacement information of the above visible cells can be automatically calculated.

  In the above-described embodiment, after obtaining two pieces of image information tn and image information t + 1 at different times, and comparing them, the center of gravity of the cells in both pieces of information is detected to obtain position information. It is not limited to. When the image information tn is acquired, prior to the acquisition of the image information t + 1, the position information may be acquired in advance by specifying a cell for the image information tn and adding identification information.

  In the above-described embodiment, after acquiring a predetermined number of intra-compartment cell movement information, the intra-compartment cumulative cell movement information is acquired and evaluated based on these, but the present invention is not limited to this. The acquisition form of the intra-compartment cumulative cell migration information can be appropriately changed as necessary, and the intra-compartment cumulative cell migration information is the information at any point in time from t1 to tN. It may be based. Typically, up to an arbitrary time can be accumulated including t1. For example, once intra-compartment cell migration information is obtained, these may be accumulated and / or evaluated in a timely or sequential manner. Furthermore, the cell displacement information, the intra-compartment cell movement information, and the intra-compartment cumulative movement information may be obtained after obtaining a predetermined number of pieces of image information.

  In the above-described embodiment, the two-dimensional distribution map information is acquired using the color information as the intra-compartment cumulative cell movement information. However, the present invention is not limited to this. The intra-compartment cumulative cell movement information is information based on the displacement of the cell and can be numerical information, and therefore various evaluation methods are possible. Further, the two-dimensional distribution map information can naturally be constituted by other image information such as shading information and patterns without depending on the color information. In addition, the criteria (such as the upper 90%) for giving such color information and shading information can be changed as appropriate. Further, the intra-compartment cumulative cell movement information can be efficiently evaluated without using the two-dimensional distribution map information. For example, height information such as a bar graph corresponding to the information can be given to each section and displayed three-dimensionally. Further, the intra-compartment cumulative cell movement information may be converted into a histogram. That is, the histogram information may be acquired by using the frequency determined based on Charlie's law or the like for the intra-compartment cumulative cell movement information. By acquiring the histogram information, it is possible to perform tests such as F test and T test on the histogram information under the two conditions to be compared. Alternatively, it is possible to perform evaluation by combining a histogram and numerical information obtained by a statistical method such as a center value or an average value.

(Cell population production method)
A cell population production method according to the present disclosure includes a cell population production step of culturing cells originating from a cell population to obtain the cell population, and in the production step, one cell virtually divided into a plurality of cells A plurality of intra-compartment cell migration information based on cell displacement information relating to the displacement of one or more cells in a predetermined period in at least two sections of the culture region of the population are obtained for different periods, and the intra-compartment cumulative cells for each of the sections The movement information can be acquired, and the quality evaluation of the cell population can be performed based on the intra-compartment cumulative cell movement information. According to this production method, a good quality cell population can be efficiently and stably produced. Various aspects of the quality control method described above can be applied to the acquisition of the intra-compartment cumulative cell migration information and the quality evaluation based on the intra-compartment cumulative cell migration information in this production method. Also in this production method, the step of obtaining intra-compartment cumulative cell migration information for the control cell population may be performed, and quality evaluation may be performed using the intra-compartment cumulative cell migration information for the control cell population. .

  In the cell population production step, the cells may be selectively cultured based on the intra-compartment cumulative cell movement information. That is, if the quality of cells in a specific section is low or deterioration can be detected based on the intra-compartment cumulative cell migration information, some cells in the culture region corresponding to the section are extinguished with a laser or the like. can do. Alternatively, only the cells in the culture region other than the compartment can be separately cultured or subcultured. In this way, a cell population with better quality can be obtained.

  The cell population production step is preferably performed under conditions that cause the origin cells to differentiate into a desired differentiation state. According to such a cell population production process, a cell population used for regenerative medicine or the like can be produced efficiently and stably with good quality.

(Cell inspection method)
The cell inspection method according to the present disclosure includes cell displacement information relating to displacement of one or more cells in a predetermined period of at least two sections of a culture region of a cell population derived from the cells virtually divided into a plurality of sections. Obtaining a plurality of intra-compartment cell migration information for different periods, obtaining intra-compartment cumulative cell migration information for each of the compartments, and examining the quality of the cells based on the intra-compartment cumulative cell migration information Can be. According to this inspection method, since the cells are inspected based on the intra-compartment cumulative cell movement information, the quality of the cells can be evaluated efficiently and accurately. In particular, cell quality can be evaluated by avoiding or suppressing individual differences in cells to be examined and variations in dynamic behavior of cells. Various aspects of the quality control method described above can be applied to the acquisition of the intra-compartment cumulative cell migration information and the evaluation of the cell quality based on the intra-compartment cumulative cell migration information in this inspection method. Also in this test method, the quality evaluation may be performed by using the intra-compartment cumulative cell migration information for the control cell population by performing the step of obtaining the intra-compartment cumulative cell migration information for the control cell population. .

(Cell quality control device and system)
The cell quality control device according to the present disclosure is capable of processing image information about the culture region acquired by the cell culture device, an imaging unit capable of imaging a cell culture region in the cell culture device, and the imaging unit. A control device including various control means, an output means for outputting a processing result by the control means,
The control means includes cell displacement information relating to displacement of one or more cells in a predetermined period of at least two of the culture regions virtually divided into a plurality of cell populations of one or two or more cell populations. Obtaining a plurality of intra-compartment cell migration information for different periods and obtaining intra-compartment cumulative cell migration information for each of the compartments, and the one or more cell populations based on the intra-compartment cumulative cell migration information It is possible to provide an apparatus that is a means for executing the step of evaluating the quality of According to this apparatus, it is possible to provide a cell quality control apparatus capable of evaluating quality efficiently and quantitatively. The cell culture device can acquire various forms, and the imaging means may acquire not only a phase contrast microscope but also a bright field image as long as it can perform contour extraction by image processing in a cell image. The output means may be a printer in addition to the display.

  This apparatus can also take the form of a cell quality management control apparatus provided with the above-described control means, and can also take the form of a cell quality management program for causing the control means such as a computer to execute the above steps. You can also. Furthermore, it is possible to take the form of a storage medium storing such a quality control program.

  This apparatus may be configured as a system, for example. That is, only some of the components of the device may constitute the device, and the remaining components may constitute the quality management system as a configuration outside the device. Moreover, all the components of this apparatus may comprise the quality control system as separate apparatuses.

  The cell quality control method, the cell quality control device, the cell quality control program, etc. according to the present disclosure are not limited to managing the cell quality as long as the purpose is to evaluate the cell quality and quality. It is implemented as a method, apparatus, program, etc. for cell evaluation and examination. The present disclosure includes such forms.

  Hereinafter, examples embodying the present disclosure will be described. These examples are intended to illustrate the present disclosure and not to limit it.

(Quality evaluation using bone differentiation induction images of human MSC)
In this example, the following three types of normal human mesenchymal stem cells MSC (KURABO, Japan) were used to obtain the skeletal differentiation-inducing images over time, and the quality evaluation was performed based on the image information. .

(cell)
Cell line1 (strain number 15000-1, unknown race, Male, 19-year-old),
Cell line2 (strain number 17174, Oriental, Male, 20-year-old),
Cell line3 (strain number 11533, Black, Male, 22-year-old

(Culture conditions)
(1) In a CO2 incubator under 37 ° C, 5% CO2, 95% Air (2) Cell culture dish (Greiner Bio-One, Frickenhausen, Germany)
(3) Growth medium For 1 week until the start of differentiation culture, basal medium: Medium 106S (KURABO), low serum growth additive: LSGS (KURABO), antibiotic: gentamicin (GA) (KURABO), and then differentiation The medium was replaced with an induction medium, cultured for 2 weeks, and the medium was changed once every 3 days. The differentiation induction medium is basal medium: Minimum Essential Medium (MEM, Invitrogen, Gaithersburg, MD, USA), Serum: 10% fetal bovine serum (FBS, Invitrogen, Gaithersburg, MD, USA), Antibiotic: Penicillin Streptomycin (PS, Invitrogen, Gaithersburg, MD, USA).
(4) Photographing of culture images During culture, phase difference images of cells were obtained at intervals of 8 hours using a phase contrast microscope, and images of 5 fields per 1 well were obtained. The magnification was 4 times. Among these, images at 0, 8, 16, 24, 32, 40, 48, 56, 64 and 72 hours after the start of guidance were picked up one by one at random (each lot) and used for image analysis.

(1) To obtain the amount of cell movement, free image analysis software ImageJ is used to cut out a visual field area containing about 200 cells from the image, and each cell in the image is marked by hand for 8 hours. Each cell position information (center of gravity coordinates) was calculated.
(2) In the images before and after each 8-hour interval, the movement distance [pixel] between two points of the center of gravity of those recognized as the same cell was calculated, and the velocity was calculated by dividing by the incubation time of 8 hours.
V = {((X-X0) ² + (Y-Y0) ² ) ^0.5 } / 8
(3) A grid capable of forming 50 pixel x 50 pixel sections in each image is given, and the movement distance from the previous image to the next image of all cells contained in these sections is calculated as an average value from the centroid information of each cell. Defined as “cell velocity” within the compartment.
(4) This is calculated for all the sections, and the upper 90% or more is indicated by the maximum value, the lower 10% or less is indicated by the minimum value, and the middle% is indicated by a color by gradation. A “velocity map” was created at each time by drawing.
(5) The speed information of the same section in the “speed map” calculated at each time is integrated (added) over time to obtain the “integrated speed”, and the top 90% in all sections and all sections Is the maximum value, the lower 10% or less is the minimum value, and the intermediate% is redrawn in color by gradation, and this is referred to as an “integrated speed map”.
(6) Further, the value of each section in these integrated speed maps (integrated maps of 32h or more) is counted according to Charlie's law, and the frequency (number of sections) of each gradation is counted, and the frequency distribution table (histogram) is displayed. Produced. Histograms were prepared for each cell line and for each passage number different from each other, and a summary of the cell lines and passage number.
(7) The comparative examination of the frequency distribution table for each prepared condition was evaluated using the comparison between the median and standard deviation, and the same state test (statistical method) as the distribution by f test.
(8) Furthermore, ALP activity, which is a general index of the degree of bone differentiation, was measured after 2 weeks from the start of culture for both cell groups with and without induction.

  FIG. 6 shows the relationship between the integration time and the coefficient of variation (CV) of the integration rate at each integration time, FIG. 7 shows the relationship between the presence / absence of integration of the rate at each culture time and the skeletal differentiation condition, FIG. 9 shows the relationship between the speed integration effect and the bone differentiation condition (for each cell line), FIG. 9 shows the relationship between the speed integration effect and the bone differentiation condition (with the difference in passage number), and FIG. The relationship between the speed integration effect (average value of the integrated speed at 40 h) and ALP activity is shown.

  As shown in FIG. 6, the cell velocity CV value obtained at each culture time interval showed an unstable tendency regardless of the time course of the culture, whereas the cell velocity obtained at each culture time interval. It has been found that the CV value of the integrated speed obtained by integrating is small as a whole, and that the CV value tends to be smaller and more stable as the integration time increases. In particular, it has been found that when the integrated time is 24 hours (32 hours as an image acquisition time) or more, more preferably 32 hours (40 hours as an image acquisition time) or more, the stabilization is further reduced.

  As shown in FIG. 7, according to the speed map obtained at each culture time interval (upper part of FIG. 7), the difference with / without differentiation induction is difficult to understand even with changes over time, whereas the integrated speed According to the map, it was found that the difference with / without differentiation induction could be clearly grasped with the passage of time, that is, the integration time increased, and the difference could be confirmed even in the early stage of culture.

  As shown in FIG. 8, it was found that even with MSCs derived from different patients, the presence / absence of differentiation induction can be clearly distinguished from the cumulative velocity distribution by making the cumulative velocity a histogram. In addition, as shown in FIG. 9, even when the three cell lines were collectively compared by passage number (P3-P6 or P5-P8), the difference with / without differentiation induction could be clearly distinguished. . In other words, this shows the possibility that this method can be applied universally even if the state of the cells is somewhat different. In addition, the cell line group with a small number of passages has a slower distribution of movement, and the one with a large number of passages accumulates damage due to human manipulation, resulting in greater cell variation. This suggests the possibility of expressing even a change in the degree of quality, such as the existence ratio of “a group with fast speed = a group far from differentiation” increasing.

  As shown in FIG. 10, it was found that the ALP activity was high when the integration rate was small due to differentiation induction, and that the ALP activity was low when the integration rate was large without differentiation induction. That is, it was found that there is a negative correlation (inverse correlation) between the integrated rate and the ALP activity with a high correlation (correlation coefficient = −0.81).

(Quality evaluation using cultured images of retinal pigment epithelium (RPE) cells using human iPS cells)
In this example, differentiation induction images for RPE were obtained over time using human iPS cells (201B7 strain, RIKEN Cell Bank, Japan), and quality evaluation was performed based on the image information.

(cell)
Human iPS cells (201B7 strain, RIKEN Cell Bank, Japan)
(Subculture conditions)
Mouse embryo fibroblasts inactivated by mitomycin-C treatment are used as feeder cells, and the Kyoto University iPS cell culture protocol (http://www.cira.kyoto-u.ac.jp/j/research/protocol.html ).
(Differentiation culture to RPE)
The cells were differentiated into human retinal pigment epithelial cells (RPE) over about 90 days according to the culture protocol reported in the following literature.
Jin Z. B, et al. Modeling retinal degeneration using patient-specific induced
pluripotent stem cells. PLoS One 6.e17084 (2011), Jin ZB, et al. Induced pluripotent stem cells for retinal degenerative diseases: a new perspective on the challenges, J Genetics 88. 417-24 (2009)

(Acquisition of image information)
5 days after the seeding of the colored iPS cell-derived RPE cell-derived colonies, the time was 0 time, 10 minutes, 8 fields, 27.5 hours in total,
A: Appearance is low in melanin (low degree of differentiation)
B: Melanin is sparse in appearance (intermediate differentiation)
C: Melanin is sparse in appearance (intermediate degree of differentiation)
D: Appearance of dark melanin (high degree of differentiation)
These four time-lapse images were selected, and these time-lapse images were acquired and analyzed for six time-points of 3.5, 4.5, 7.5, 8.5, 10.5, and 11.5 hours. FIG. 11 shows an image of the visual field to be analyzed of RPE cells derived from human iPS cells. Hereinafter, the visual field A may be referred to as “NEGA”, and the visual field D may be referred to as [POSI].

(1) Free image analysis software ImageJ is used to acquire dynamic information, and 3.5-4.5, 7.5-8.5, and 10.5-11.5 time intervals. A field-of-view region containing about 200 cells was cut out from the image, and each cell in the image was marked by hand, and the position information (center of gravity coordinates) of each cell was calculated every hour.
(2) In the images before and after each one-hour interval, the moving distance [pixel] between the two centers of gravity of those recognized as the same cell was calculated, and the speed was calculated by dividing by the incubation time of 1 hour.
V = V = {((X-X0) ² + (Y-Y0) ² ) ^0.5 } / 1 (= time interval)
(3) A grid capable of forming 50 pixel x 50 pixel sections in each image is given, and the movement distance from the previous image to the next image of all cells contained in these sections is calculated as an average value from the centroid information of each cell. Defined as “cell velocity” within the compartment.
(4) This is calculated for all the sections, and the upper 90% or more is indicated by the maximum value, the lower 10% or less is indicated by the minimum value, and the middle% is indicated by a color by gradation. A “velocity map” was created at each time by drawing.
(5) The speed information of the same section in the “speed map” calculated at each time is integrated (added) over time to obtain the “integrated speed”, and the top 90% in all sections and all sections Is the maximum value, the lower 10% or less is the minimum value, and the intermediate% is redrawn in color by gradation, and this is referred to as an “integrated speed map”.
(6) Further, the frequency values (number of sections) of each gradation in the speed map and the integrated speed map were counted according to Charlie's law, and a frequency distribution table (histogram) was prepared.
(7) The comparative examination of the frequency distribution table for each prepared condition was evaluated using the comparison between the median and standard deviation, and the same state test (statistical method) as the distribution by f test.
(8) Further, for each visual field, the accumulation degree (luminance), cell size, perimeter length, and circularity, which are the standard of RPE differentiation level in the current cell culture, as well as the moving speed, The numerical value in a division was calculated, the integration map in 3.5-11.5 and various numerical maps were created, and the histogram was created.

  FIG. 12 shows the relationship (histogram) between the melanin accumulation maturity of human iPS cell-derived RPE cells and the integration map, and FIG. 13 shows the relationship (scatter) between the melanin accumulation maturity of human iPS cell-derived RPE cells and the integration map. FIG. 14 shows the relationship between melanin accumulation maturity (luminance histogram) and shape / velocity (histogram) of human iPS cell-derived RPE cells, and FIG. 15 shows melanin accumulation of human iPS cell-derived RPE cells. The relationship between maturity (pixel) and shape (median) / speed (median) is shown.

  As shown in FIG. 12, according to the histogram created from various speed maps and integrated speed maps, the speed is widely dispersed and high in NEGA, whereas in POSI, it is concentrated at a low speed. I understood that I can grasp it. Furthermore, as shown in FIG. 13, according to the scatter diagram, in NEGA and POSI, it is possible to clearly grasp the degree of variation, average existence, irregular existence, and change over time, for example, in POSI. It was found that the tendency of speed stabilization could be grasped. In addition, as shown in FIG. 14, a histogram (lower side) of melanin (integrated) and a histogram (upper side) of cell size (integrated), perimeter length (integrated), circularity (integrated), and velocity (integrated) , It was found that the velocity histogram most closely corresponds to the distribution of the melanin histogram. Furthermore, as shown in FIG. 15, the median speed of each speed map and melanin (pixel) showed a high positive correlation. On the other hand, the median of each map of area and circularity and the pixel showed only a low correlation. Further, as shown in FIG. 15, the median speed obtained from each speed map tends to become smaller as the integration time zone becomes slower for both NEGA and POSI, and the accuracy of quality evaluation based on speed is high. I understood it. From the above, it was found that by using the integrated speed in the section, the integrated speed and the degree of differentiation are well correlated in various statistical methods, and the integrated speed is a good indicator of the degree of differentiation. It was also found that quality such as cell differentiation can be evaluated efficiently and with high accuracy by combining various statistical methods with the integrated speed.

Claims (14)

A cell quality control method comprising:
Different cell movement information in the compartment based on the cell displacement information relating to the displacement of one or more cells in a predetermined period of at least two of the sections of the culture region virtually divided into a plurality of one or two or more cell populations Obtaining a plurality of per-period, and obtaining the intra-compartment cumulative cell migration information for each of the compartments,
A quality control method for evaluating the quality of the one or more cell populations based on the intra-compartment cumulative cell migration information.   The quality control method according to claim 1, wherein the step of acquiring the intra-compartment cumulative cell migration information is performed using image information acquired for a culture region of the one or more cell populations.   The cell displacement information is based on start point position information of the one or more cells existing in the section at the start time of the predetermined period and end point position information of the end point of the predetermined period of these cells. The quality control method according to claim 1 or 2. The following (a) to (c):
(A) obtaining two pieces of image information relating to a cell culture state at a time t _n (n is an integer of 1 or more and N or less) and a time t _{n + 1 for} one or more cell populations;
(B) Based on the position information of one or more cells at time t _n of each section in the two image information and the position information of these cells at time t _{n + 1} , the one or more cells Obtaining the cell displacement information about the cell, and obtaining the intra-compartment cell migration information of each of the compartments at t _n to t _{n + 1} based on the cell displacement information,
(C) Repeat (a) and (b) until t = N,
(D) The quality according to any one of claims 1 to 3, wherein the intra-compartment cumulative cell movement information for each of the sections is acquired based on the N-1 intra-partition cell movement information acquired for each of the sections. Management method.   The quality control method according to claim 1, wherein the displacement of the cell is acquired based on a displacement of the center of gravity of the cell.   The quality control method according to any one of claims 1 to 5, wherein the intra-compartment cumulative cell migration information is evaluated by plotting a two-dimensional distribution diagram assigned to the compartment on the corresponding culture region.   The quality control method according to any one of claims 1 to 6, wherein the intra-compartment cumulative cell migration information is evaluated in the form of a histogram.   The quality control method according to claim 1, wherein the quality of the one or two or more cell populations is evaluated for each section.   The quality control method according to claim 1, wherein the quality is a cell differentiation degree. A method for producing a cell population,
A cell population production step of culturing cells originating from the cell population to obtain the cell population;
With
In the production process, the intra-compartment cell movement information based on the cell displacement information on the displacement of one or more cells in a predetermined period in at least two of the culture regions of one cell population virtually divided into a plurality of sections A plurality of different periods, obtaining intra-compartment cumulative cell migration information for each of the compartments, and evaluating the quality of the cell population based on the intra-compartment cumulative cell migration information.   The production method according to claim 10, wherein in the cell population production step, cells are selectively cultured based on the intra-compartment cumulative cell migration information.   The production method according to claim 10 or 11, wherein the cell population production step is performed under conditions that cause the origin cells to differentiate into a desired differentiation state. A method for examining cells,
Different intra-compartment cell movement information based on cell displacement information regarding the displacement of one or more cells in a predetermined period of at least two of each of the culture regions of the cell population derived from the cells virtually divided into a plurality of cells Obtaining a plurality of per-period, and obtaining the intra-compartment cumulative cell migration information for each of the compartments,
A test method for testing the quality of the cell based on the intra-compartment cumulative cell migration information. A cell quality control device,
A cell culture device;
Imaging means capable of imaging a cell culture region in the cell culture device;
A control device comprising control means capable of processing image information about the culture region acquired by the imaging means;
Output means for outputting a processing result by the control means;
With
In the compartment based on the cell displacement information relating to the displacement of one or two or more cells in a predetermined period of at least two of the sections of the culture region virtually divided into a plurality of one or two or more cell populations Obtaining a plurality of cell migration information for different periods to obtain intra-compartment cumulative cell migration information for each of the compartments, and determining the quality of the one or more cell populations based on the intra-compartment cumulative cell migration information And a step of evaluating.