JP2010152829A - Cell culture management system - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a cell culture management system, capable of determining an optimum operation scheduled date and time. <P>SOLUTION: A schedule storage part 33 stores information for facility and doctor. A schedule formation part 32 predicts an arrival day when a cell grows by culture into a necessary cellular quantity needed for an operation using the cell, and determines an operation scheduled date and time for the operation using the cell based on the information for facility and doctor in that arrival day. An observation mechanism 22 observes the culture condition of the cell at a predetermined time after start of the cell culture, and the schedule formation part 32 re-predicts the arrival day according to the observation result, and corrects the operation scheduled date and time according to the re-predicted arrival day. The present invention can be applied to, for example, a cell culture management system for managing a culture schedule. <P>COPYRIGHT: (C)2010,JPO&INPIT

Description

  The present invention relates to a cell culture management system, and more particularly, to a cell culture management system capable of determining an optimal scheduled operation date and time.

  Regenerative medicine, which has been actively researched in recent years, restores functions by, for example, culturing cells in a culture container, regenerating lost human tissue, and transplanting the tissue to a patient. It is medical.

  For example, Patent Document 1 discloses a management system that manages a culture plan of collected cells in accordance with a hospital or doctor's treatment system. In this management system, the culture rate of cells in culture is controlled in accordance with the patient's medical condition, the schedule of a hospital or doctor, and the shipment schedule of stem cells or tissues is scheduled.

JP 2004-290147 A

  By the way, in general, since the culture period of the cell greatly depends on the culture state of the cell, as disclosed in Patent Document 1, the culture rate was controlled in accordance with the treatment system of the hospital or doctor. In some cases, abnormalities may occur in the cells. In addition, if the culture does not proceed according to the schedule, a significant schedule change may be performed immediately before the scheduled operation date, and it may be difficult to ensure a sufficient medical system.

  Therefore, there has been a demand for a system that determines the scheduled operation date and time in which the combination of cells is optimal in consideration of both the cell culture state and the treatment system of the hospital or doctor.

  The present invention has been made in view of such a situation, and makes it possible to determine an optimal scheduled operation date and time.

  The cell culture management system of the present invention is a cell culture management system for culturing cells and managing a culture schedule, and storing means for storing information relating to a facility and a doctor; Prediction means for predicting the arrival date to reach the required cell amount required for surgery using a surgery, and surgery using the cells based on information on the facility and doctor on the arrival date predicted by the prediction means Determining means for determining the scheduled operation date and time.

  In the cell culture management system of the present invention, the arrival date when the cells are cultured and proliferated and reach the necessary cell amount necessary for the operation using the cells is predicted, and based on the information on the facility and the doctor on the arrival date The scheduled operation date and time for performing an operation using cells is determined.

  According to the cell culture management system of the present invention, it is possible to determine the optimal scheduled operation date and time.

  Hereinafter, specific embodiments to which the present invention is applied will be described in detail with reference to the drawings.

  FIG. 1 is a block diagram showing a configuration example of an embodiment of a cell culture management system to which the present invention is applied.

  In FIG. 1, a cell culture management system 11 includes a cell culture device 12 having a culture chamber maintained in a culture environment suitable for cell culture, a management device 13 for managing cell culture by the cell culture device 12, and management. A display device 14 for displaying a schedule created by the device 13 and an input device 15 for inputting an operation to the management device 13 are provided.

  In the cell culture management system 11, cells are cultured in the cell culture device 12 in accordance with management by the management device 13. When the cells cultured in the cell culture device 12 (hereinafter referred to as “cultured cells” as appropriate) reach a necessary cell amount necessary for transplantation into the patient, an operation for transplanting the cultured cells into the patient is performed. .

  The cell culture device 12 includes a culture mechanism 21 for maintaining a culture chamber under conditions suitable for cell culture (for example, a temperature of 37 degrees, a humidity of 90% or more, and a carbon dioxide concentration of 5%), the behavior and state of cultured cells, and the like. An observation mechanism 22 for observing cells, a medium exchange mechanism 23 for exchanging a medium required in the cell culture process, a passage mechanism 24 for seeding the proliferated cells, and cell differentiation induction and proliferation. It is configured with a solution addition mechanism 25 for promoting.

  The management device 13 includes a control unit 31 that controls each block included in the cell culture device 12, a schedule creation unit 32 that creates a schedule such as an operation date for transplanting the cultured cells to a patient, and data and schedules input to the management device 13 A schedule storage unit 33 for storing a schedule created by the creation unit 32, a culture data storage unit 34 for storing a database in which information on cell culture is registered, and a database stored in the culture data storage unit 34 are searched. A search unit 35 and an image processing unit 36 for obtaining an amount of cultured cells by processing an image obtained by observation by the observation mechanism 22 are provided.

  The culture data storage unit 34 stores culture history of cells cultured in the past in the cell culture management system 11 (for example, data related to cell growth such as cell type, culture conditions, and proliferation rate), and other information ( For example, a database in which information on the patient of the cell and information on containers and reagents used for the culture is registered is stored. And the schedule preparation part 32 refers to the database of the culture data memory | storage part 34 via the search part 35 based on the information input into the management apparatus 13 when starting the culture | cultivation of a cell with the cell culture apparatus 12, The predicted completion date (date of arrival) at which the cultured cells are predicted to reach the required cell amount is determined.

  Moreover, the schedule preparation part 32 determines the candidate date of an operation date based on the information regarding a patient, the information regarding the facility where surgery is performed, the information regarding the doctor who performs an operation, etc. The schedule creation unit 32 displays an input screen for allowing the user of the cell culture management system 11 to input such information on the display device 14 such as a CRT (Cathode Ray Tube) or an LCD (Liquid Crystal Display). The user operates the input device 15 including a keyboard and a mouse to input necessary information.

  For example, when starting the culture of a cell, the schedule creation unit 32 inputs a patient ID (Identification) for identifying the patient of the cell, and associates the patient ID with information about the patient in the schedule storage unit 33. And create a schedule for the patient. Information about the patient includes detailed conditions such as the date and time that the patient can visit, the date and time that the patient can be hospitalized, the date and time that the patient can operate, other examination information about the patient, and the operation time.

  For example, FIG. 2 shows an operation screen 41 that is operated when information related to a patient is input or displayed.

  The user of the cell culture management system 11 operates the pull-down menu 42 to specify a desired patient ID, or operates the search button 43 to display a search screen for searching for the patient ID, and the search result The desired patient ID can be designated from As a result, the patient ID designated by the user (patient ID “125856” in the example of FIG. 2) is displayed in the pull-down menu 42. When the display buttons 44a to 44e corresponding to various types of information related to the patient are operated, a display screen for displaying each information related to the patient specified by the designated patient ID is displayed. In addition, when the input buttons 45a to 45e corresponding to various types of information related to the patient are operated, an input screen for inputting each information related to the patient specified by the designated patient ID is displayed. Various information about the patient is input using the input screen.

  Next, FIG. 3 shows an operation screen 51 that is operated when inputting or displaying information about a facility, and FIG. 4 is an operation screen operated when inputting or displaying information about a doctor. 61 is shown.

  In the operation screen 51 of FIG. 3, the operating room ID for specifying the desired operating room is specified by operating the pull-down menu 52, or the search screen for searching the operating room ID is displayed by operating the search button 53. The desired operating room ID can be designated from the search result, and the designated operating room ID (“OR0101” in the example of FIG. 3) is displayed in the pull-down menu 52. Then, the display button 54 is operated to display a display screen displaying the reservation status of the operating room specified by the specified operating room ID, or the input button 55 is operated to specify the specified operating room ID. An input screen for inputting a reservation for the operating room to be displayed can be displayed to input (change) the reservation for the operating room.

  In the operation screen 61 of FIG. 4, a pull-down menu 62 is operated to specify a doctor ID for specifying a desired doctor, or a search screen for searching a doctor ID by operating the search button 63 is displayed. The desired doctor ID can be designated from the menu, and the designated doctor ID (“A456” in the example of FIG. 4) is displayed in the pull-down menu 62. Then, the display button 64a or 64b corresponding to the doctor's work schedule (whether or not normal work is possible or not) is operated to display a display screen for displaying the doctor's work schedule specified by the designated doctor ID. Enter or change the attendance schedule by displaying the input screen for entering the attendance schedule of the doctor identified by the designated doctor ID by operating the input button 65a or 65b corresponding to the attendance schedule of the doctor. )can do.

  Information regarding the facility and the doctor is appropriately input to the management device 13 as necessary, and is referred to when determining a candidate for the scheduled surgery date, as will be described later with reference to FIG.

  Further, when starting the culture of a cell, the schedule creation unit 32 stores information on the cultured cell in the schedule storage unit 33 in association with the patient ID of the patient of the cell. Information on cultured cells includes the cell ID that identifies the cultured cells, the input date when the cultured cells were started, the input amount of the cultured cells, the required amount of cultured cells necessary for transplantation to the patient, and the required cell amount. There are critical points that specify the allowable amount, the predicted completion date when the cultured cells are expected to reach the required amount, and the timing for determining whether to modify the schedule.

  Here, in the cell culture management system 11, from the start of cell culture until the scheduled operation date so that appropriate medical cells can be provided while sufficiently constructing the medical system of facilities and doctors. In the meantime, one or more critical points are set. By observing the cultured cells at the critical point, the schedule can be appropriately corrected in accordance with the increase / decrease in the growth rate during the culturing process.

  FIG. 5 shows a display screen 71 that displays information related to cultured cells. Note that the operation screens 41 to 61 in FIGS. 2 to 4 and the display screen 71 in FIG. 5 can be switched by operating corresponding tabs.

  In the display screen 71 of FIG. 5, a pull-down menu 72 is operated to specify a desired patient ID, or a search screen for searching the patient ID is displayed by operating the search button 73. ID can be specified. On the display screen 71, the designated patient ID (“125856” in the example of FIG. 5) is displayed in the pull-down menu 72, and the cell ID, input date, input amount, and required cell amount of the cultured cell of the patient are displayed. , Tolerance, forecast completion date, and critical points are displayed.

For example, the cell ID, the input date, the input amount, the required cell amount, the allowable amount, and the critical point are input by the user when starting the cell culture, and the predicted completion date is scheduled based on the information. Predicted by the creation unit 32. Specifically, as shown in FIG. 5, the input date is “October 31, 2008”, the input amount is “1 × 10 ⁵ cells = 2 μl”, and the required cell amount is “1 × 10 5”. ^{When “8} pieces = 2 ml” and the allowable amount is “1 × 10 ⁶ pieces = 2 μl”, the schedule creation unit 32 predicts the prediction completion date as “November 31, 2008”.

  Further, the schedule creation unit 32 determines whether or not to modify the schedule based on the result of measuring the cell amount of the cultured cells at the critical points “November 5, 2008” and “November 12, 2008”. to decide.

  Next, prediction and correction of the prediction completion date by the schedule creation unit 32 will be described with reference to FIG.

  FIG. 6 shows a change in the amount of cells when cell culture is started under the conditions shown in FIG. In FIG. 6, the vertical axis represents the amount of cells, and the horizontal axis represents the date.

For example, when culturing is started with the input amount “1 × 10 ⁵ cells = 2 μl” on the input date “October 31, 2008”, the schedule creation unit 32 is registered in the database of the culture data storage unit 34. With reference to the growth rate or the like, an increase in the number of cultured cells is predicted as indicated by a prediction line L1 shown in FIG. Then, the schedule creation unit 32 predicts that the required cell amount “1 × 10 ⁸ cells = 2 ml” will reach the prediction completion date “November 31, 2008”.

  When the amount of cultured cells is measured at the first critical point “November 5, 2008”, the amount of cells is less than the amount of cells predicted by the prediction line L1. The schedule creation unit 32 sets the prediction completion date “December 10-15, 2008” in accordance with the prediction line L2 obtained by extending the straight line according to the increase of the cultured cells from the input date to the first critical point. To correct.

  After that, when the cell volume of the cultured cells was measured at the second critical point “November 12, 2008”, the cell volume was a cell volume V2 larger than the cell volume predicted by the prediction line L2. The schedule creation unit 32 sets the prediction completion date “December 3-5, 2008” according to the prediction line L3 obtained by extending the straight line according to the increase in the number of cultured cells from the first critical point to the second critical point. To "".

  Here, the reason that the corrected prediction completion date is 3 days is that the allowable amount is set with respect to the required cell amount, and the schedule creation unit 32 determines the required cell amount according to the allowable amount. From the date predicted to reach the lower limit value to the date predicted to reach the upper limit value of the required cell amount according to the allowable amount, the prediction completion date range is predicted. In addition, as a result of measuring the cell amount of the cultured cell at the critical point, when the cell amount is the cell amount predicted by the prediction line, the schedule creation unit 32 does not correct the prediction completion date.

  As described above, the schedule creation unit 32 predicts and corrects the prediction completion date, and obtains a candidate for the scheduled surgery date and time based on the information regarding the facility and the doctor on the prediction completion date.

  FIG. 7 shows the schedule of the scheduled operation date and time (doctor available date and time, facility available date and time, and scheduled surgery date and time) on the predicted completion date “December 3-5, 2008” determined at the second critical point. ing.

  In the schedule storage unit 33, the date and time for which treatment is possible for each doctor ID is stored as information about the doctor, and the reservation status for each operating room ID is stored as information about the facility. In FIG. 7, hatching indicating that the doctors with the doctor IDs “a564”, “a533”, and “a511” can perform the operation is performed on the doctor available date and time, and the operating room IDs “OR0101”, “OR0104” ”And“ OR0201 ”are hatched on the facility available date and time indicating that the operating room is available date and time.

  The schedule creation unit 32 obtains a date and time that is a date and time that can be operated by the doctor and that can be used by the operating room as a candidate for a scheduled operation date and time. In FIG. 7, hatching indicating that it is a candidate for the scheduled surgery date and time is performed at 13 to 15 o'clock on December 3, 7 to 9 o'clock on December 4, and from 24 o'clock on December 4 to 6 o'clock on December 5 Has been.

  As described above, the schedule creation unit 32 corrects the prediction completion date based on the culture amount of the cultured cell for each critical point after starting the cell culture, and from the prediction completion date range obtained at the critical point, Based on the information about the facility and the doctor, a candidate for the scheduled operation date can be obtained. And the schedule preparation part 32 displays the candidate of scheduled surgery date and time on the display apparatus 14, and a user selects the scheduled surgery date and time from these candidates.

  Note that the schedule creation unit 32 may perform matching based on constraint conditions such as operation time to rank the scheduled operation date and time, and present the optimal operation date ranking for the operation to the user. In the example of FIG. 7, “December 4 24 o'clock to December 5 o'clock 6 o'clock”, which has the longest scheduled operation date, is given the highest priority, and the next scheduled operation date is “December 3, 13:00 to 15:00 Can be presented to the user as “priority ranking 3rd”, “December 4 7: 00-9: 00” having the shortest scheduled surgery date and time. In addition, the schedule creation unit 32 can make a doctor's wish, a wish of an operation time zone, and the like a matching constraint condition.

  Next, FIG. 8 is a flowchart for explaining processing for creating and correcting a schedule for a predetermined cultured cell in the management apparatus 13 of FIG.

  For example, when the user of the cell culture management system 11 stores a cell to be newly cultured in the culture chamber of the cell culture device 12, the process starts. In step S11, the schedule creation unit 32 displays information on the cultured cell. An input screen for input is displayed on the display device 14. When the user operates the input device 15 and inputs information about the cultured cells such as the cell ID, the input date, the input amount, the required cell amount, the allowable amount, and the critical point, the schedule creation unit 32 displays the information. And the process proceeds to step S12.

  In step S12, the schedule creation unit 32 instructs the search unit 35 to search for the cell ID acquired in step S11, and the search unit 35 searches the database in the culture data storage unit 34 to find the corresponding growth rate. That is, the proliferation rate registered in association with the cell ID acquired in step S11 is acquired and supplied to the schedule creation unit 32. Then, in accordance with the prediction that the input amount of cultured cells will grow according to the growth rate, the schedule creation unit 32 calculates the number of days required until the cultured cell reaches the required cell amount, and adds the number of days to the input date. Thus, the prediction completion date is predicted, and the process proceeds to step S13.

  In step S <b> 13, the schedule creation unit 32 obtains the predicted completion date range as described with reference to FIG. 6 according to the allowable amount for the necessary cell amount. Furthermore, as described with reference to FIG. 7, the schedule creation unit 32 extracts a candidate for the scheduled surgery date and time from the predicted completion date range based on the information regarding the facility and the doctor. And the schedule preparation part 32 displays the candidate of a scheduled operation date on the display apparatus 14, and determines what was selected by the user from the candidate of a scheduled operation date as a scheduled operation date. The schedule creation unit 32 causes the schedule storage unit 33 to store information on the cultured cells acquired in step S11 together with the determined scheduled surgery date.

  After the process of step S13, the process proceeds to step S14, and the schedule creation unit 32 determines whether to confirm the state of the cultured cell according to the critical point of the cultured cell stored in the schedule storage unit 33, and the culture The process waits until it is determined that the state of the cell is confirmed. Then, when the date specified by the critical point of the cultured cell is reached, in step S14, the schedule creation unit 32 determines that the state of the cultured cell is confirmed, and the process proceeds to step S15.

  In step S15, the schedule creation unit 32 causes the control unit 31 to execute a cell state observation process (the process of the flowchart of FIG. 12 described later). In the cell state observation process, the observation mechanism 22 captures an image of the cultured cell according to the control of the control unit 31, and the image processing unit 36 performs image processing on the image of the cultured cell to determine the cell amount of the cultured cell and creates a schedule. To the unit 32.

  After the cell state observation process of step S15, the process proceeds to step S16, and the schedule creation unit 32 determines whether or not to correct the scheduled operation date based on the cell amount of the cultured cells obtained by the image processing unit 36. judge.

  For example, the schedule creating unit 32 is on the prediction line (for example, L1 or L2 in FIG. 6) when the cell amount of the cultured cells obtained by the image processing unit 36 in the immediately preceding step S15 and the prediction completion date are predicted last time. It is determined that the scheduled operation date / time is not to be corrected if the cell volume is consistent with the cell amount, and it is determined that the scheduled operation date / time is to be corrected otherwise.

  In step S16, when the schedule creation unit 32 determines to modify the schedule, the process proceeds to step S17, and the schedule creation unit 32 determines the cell volume of the cultured cell at the critical point, the proliferation rate of the cultured cell, and the necessary. The prediction completion date range is predicted again based on the tolerance for cell volume. Furthermore, the schedule preparation part 32 calculates | requires the candidate of an operation scheduled date and time based on the information regarding the facility and doctor in the prediction completion date range estimated again.

  After the process of step S17, the process proceeds to step S18, and the schedule creation unit 32 displays the scheduled surgery date / time candidate obtained in step S17 on the display device 14, and the scheduled surgery date / time is selected from the scheduled surgery date / time candidates. The schedule is corrected to the one selected by the user and the schedule stored in the schedule storage unit 33 is updated.

  After the process of step S18 or when the schedule creation unit 32 determines in step S16 that the scheduled surgery date and time are not to be corrected, the process returns to step S14, and the same process is repeated thereafter.

  As described above, the cell culture management system 11 can determine the optimal operation date. That is, in the cell culture management system 11, the scheduled operation date and time is determined based on the schedule of the facility and the doctor on the prediction completion date predicted from the input amount and the growth rate. Therefore, the cell culture state, the facility and the doctor are determined. The date and time that is convenient for both of these schedules can be determined as the scheduled operation date and time.

  In addition, the amount of cultured cells is measured at the critical point, and the scheduled surgery date and time are corrected based on the measurement result and the schedule of the facility and doctor, so the scheduled surgery date and time can be accurately predicted, for example, The occurrence of a change from the scheduled surgery date determined at the last critical point is suppressed. Thereby, since a schedule before a certain period can be secured, a sufficient medical system can be secured. In addition, by setting a plurality of critical points, the prediction accuracy of the prediction completion date can be improved.

  In addition, by setting an allowable amount with respect to the required cell amount, it is possible to widen the range for selecting the scheduled surgery date and time, and to determine the desired scheduled surgery date and time. In addition, by setting an acceptable amount for the required cell amount, even if the final cultured cell amount has increased or decreased relative to the required cell amount, the scheduled operation date and time can be changed if it is within the allowable amount. Can be avoided and surgery can be performed.

  Even if the amount of cells measured at the critical point is slightly increased or decreased with respect to the predicted amount of cells, if the amount of increase or decrease is within the allowable range, the previous schedule is not changed. May be maintained. As a result, frequent schedule changes can be avoided, and the burden on the doctor and patient can be reduced.

  As described above, the cell culture management system 11 can secure a schedule in a certain period of time in a more appropriate cell state than before, so that it is possible to give merit to both the patient and the doctor.

  Next, a method for measuring the amount of cultured cells will be described with reference to FIGS.

  For example, as shown in FIG. 9, the observation mechanism 22 captures a plurality of images (five images in the example of FIG. 9) at different positions in the Z-axis direction by the phase difference observation method. The cell region and the three-dimensional volume of the cell image are calculated from these images.

  First, the image processing unit 36 calculates a variance value of luminance at the same position (the same XY coordinate) where the Z-axis position is different for all pixels on a predetermined image captured by the observation mechanism 22.

  That is, as shown in FIG. 9, the image processing unit 36 obtains a point P (point of coordinates (X, Y) = (200, 300)) from each of the five images from the Z-axis position 1 to the Z-axis position 5. Luminance values are acquired, and a variance value of those luminance values is calculated. For example, the luminance value at point P in the image at Z-axis position 1 is 150, the luminance value at point P in the image at Z-axis position 2 is 170, the luminance value at point P in the image at Z-axis position 3 is 165, and the Z-axis position When the luminance value at the point P in the image 4 is 164 and the luminance value at the point P in the image at the Z-axis position 5 is 162, the image processing unit 36 calculates the variance value 55.2.

  Next, when the variance value of a certain coordinate is equal to or less than a predetermined threshold, the image processing unit 36 estimates that no cell exists at that coordinate.

  Here, in general, in the phase difference image, the luminance of the phase difference has a characteristic that the portion having a larger phase change has a characteristic that the luminance value tends to be maximized around the cell contour. Therefore, the image processing unit 36 first obtains a rough position of the cell outline based on the luminance dispersion value calculated as described above, and the presence of the cell in the area of the cell outline and in the vicinity of the cell outline. Assuming a region, three-dimensional extraction of the cell contour is performed.

  The image processing unit 36 detects the maximum luminance value in the Z-axis direction for all the pixels on a specific image. The maximum luminance value corresponding to the detected pixel position is defined as the cell height (Z-axis position). Further, the image processing unit 36 maximizes the change in the luminance value of the phase image in a part where the phase difference is not extremely increased due to the influence of the intracellular organ or the like in the cell region obtained from the dispersion value. A part is extracted and made into cell height (Z-axis position).

  The image processing unit 36 can obtain a three-dimensional image of a cell as shown in FIG. 10 by performing the above-described processing.

  FIG. 10A shows one of a plurality of microscopic images having different Z-axis positions, and FIG. 10B shows the cell height on the line shown in the image of FIG. 10A. FIG. 10C shows a three-dimensional image of the cell.

  Furthermore, the image processing unit 36 integrates the entire area displayed in the three-dimensional plot from the three-dimensionally detected image as shown in FIG. 10C based on the magnification of the microscope and the Z-axis position thereof, thereby obtaining the cell volume. Is calculated. Then, the image processing unit 36 can obtain the amount of cells in the observation region from the calculated volume data and cell density.

  The observation region may be the entire culture region, or the amount of cells in the entire culture vessel may be estimated using data in a certain observation region as a representative point.

  Next, in the following, the process for obtaining the cell amount will be described in more detail.

  FIG. 11 is a block diagram illustrating a detailed configuration example of the observation mechanism 22 and the image processing unit 36 of FIG.

  In the cell culture device 12, a culture container 81 in which cultured cells to be observed are accommodated is placed on the sample stage 82, and the observation mechanism 22 controls the culture cells in the culture container 81 according to the control of the control unit 31. An image is captured, and the image is supplied to the image processing unit 36 via the control unit 31.

  The culture vessel 81 is, for example, a petri dish, a flask, a well plate, or the like, and cultured cells and a medium are accommodated in the culture vessel 81. The sample stage 82 is at least partially made of a translucent material, and the position of the culture vessel 81 placed on the upper surface thereof can be adjusted in the horizontal direction.

  The observation mechanism 22 includes a light source 83, a collector lens 84, a mirror 85, a field lens 86, a ring diaphragm 87 (aperture diaphragm), a condenser lens 88, an objective lens 89, a phase ring 90, an imaging lens 91, a lens driving unit 92, and An imaging unit 93 is provided.

  As shown in FIG. 11, the illumination light emitted from the light source 83 becomes parallel light by the collector lens 84 and is guided downward by the mirror 85 in FIG. 11. The illumination light reflected by the mirror 85 is collected by the field lens 86 and enters the ring diaphragm 87. The ring diaphragm 87 is a disk having a ring-shaped opening, and uses illumination light as a ring-shaped diaphragm light. The ring diaphragm 87 is disposed at the front focal position of the condenser lens 88. The condenser lens 88 condenses the light beam that has passed through the ring diaphragm 87. Thereby, the culture vessel 81 placed on the sample stage 82 is irradiated with illumination light from above.

  The objective lens 89 transmits the direct light (0th order light) transmitted through the culture vessel 81 and the diffracted light generated according to the phase object (cultured cells) in the culture vessel 81. The phase ring 90 is disposed at a position that is optically conjugate with the ring stop 87 at the rear focal plane position of the objective lens 89. The phase ring 90 transmits a part of the diffracted light generated in the cultured cells and causes a delay (or advance) in the phase of the transmitted light. Further, the imaging lens 91 forms an enlarged image of the cultured cell on the imaging unit 93 based on the direct light and the diffracted light.

  Thus, the microscopic optical system of the observation mechanism 22 is configured, and the light transmitted through the phase ring 90 interferes with the light that has passed through other than the phase ring 90, so that light and darkness is formed on the imaging surface via the imaging lens 91. It is imaged as a difference. Thereby, phase difference observation of a cultured cell can be performed.

  The lens driving unit 92 drives the objective lens 89 in the optical axis direction (vertical direction in FIG. 11) to adjust the focus of the microscopic optical system.

  The imaging unit 93 captures an image formed by the microscopic observation system and generates microscopic image data. The imaging unit 93 includes an imaging device, an analog front end that performs gain adjustment and A / D conversion on an output signal of the imaging device, and an image processing unit that performs various types of image processing. In FIG. 11, illustration of individual components of the imaging unit 93 is omitted.

  The image processing unit 36 functions as an area separation unit 102, a focus position calculation unit 103, and a three-dimensional information calculation unit 104 when a processor (not shown) executes a program stored in the memory 101.

  Next, processing performed in the region separation unit 102, the in-focus position calculation unit 103, and the three-dimensional information calculation unit 104 will be described with reference to FIGS.

  FIG. 12 is a flowchart for explaining the cell state observation process in step S15 of FIG. FIG. 13 shows an example of a microscopic image and a separation state of a cell region and a non-cell region. FIG. 14 shows an example of the relationship between the height of the contour surface of the cultured cell and the focus position. It is shown.

  In the following description, as shown in FIG. 13A, it is assumed that adherent cells and floating cells exist in the culture solution in the culture vessel 81, and the image processing unit 36 performs culture in the field of view. The cell volume is calculated.

  In step S <b> 21, the image processing unit 36 controls each block of the observation mechanism 22 via the control unit 31 to capture a plurality of microscope images having different focal positions in the same visual field range, and stores data of each microscope image. 101.

  Specifically, the control unit 31 turns on the light source 83 to illuminate the culture vessel 81 and drives the imaging unit 93 to capture a microscope image. Thereafter, the control unit 31 causes the lens driving unit 92 to shift the position of the objective lens 89 in the optical axis direction (Z-axis direction), and then repeats the imaging of the microscope image under the same imaging conditions. For example, a plurality of microscope images obtained by sectioning the coordinates Z1 to Z5 from the bottom direction of the culture vessel 81 in FIG. 13A are acquired. Thereby, a plurality of microscope images having different focal positions in the same field of view are generated (see FIG. 13B). Each microscope image data is associated with focal position information indicating the focal position of the objective lens 89 when the image is captured.

  In step S22, the region separation unit 102 of the image processing unit 36 uses the luminance values of the plurality of microscope images captured in step S21 to obtain a luminance dispersion value for each pixel of the shooting screen corresponding to the microscope image. For example, the region separation unit 102 calculates a variance value of luminance for all the pixels at the coordinates (x, y) on the photographing screen from a plurality of luminance values indicated by the pixels at the coordinates (x, y) of each microscope image. .

  In step S23, the region separation unit 102 separates the cell region where the cultured cell is located from the non-cell region where the cultured cell is not located, based on the luminance dispersion value calculated in step S22.

  As an example, the region separation unit 102 compares the luminance dispersion value at the coordinates (x, y) of the shooting screen with a threshold value for region determination. Here, at the location where the cultured cells are located on the imaging screen, a change in luminance occurs between images due to a change in the focal position. For this reason, the variance value of the brightness is increased at the location where the cultured cells are located. On the other hand, in places where cultured cells are not located on the imaging screen, there is almost no change in brightness between images even if the focal position changes. Therefore, the luminance dispersion value is extremely small at the location where the cultured cells are located.

  Accordingly, when the luminance dispersion value at the coordinates (x, y) is equal to or greater than the threshold, the region separation unit 102 determines the pixel at the coordinates (x, y) as a cell region. On the other hand, when the luminance dispersion value is less than the threshold value, the region separation unit 102 determines that the pixel at the coordinates (x, y) is a non-cell region. Then, the region separation unit 102 separates the cell region and the non-cell region on the photographing screen by performing the above determination on each coordinate of the photographing screen. In addition, the state which isolate | separated the cell area | region and the non-cell area | region on the imaging | photography screen is typically shown to FIG. 13B.

  In addition, the region separation unit 102 performs a grouping process for grouping the pixels of the cell region and a labeling process for associating identification information (for example, an identification number) with each grouped cell region on a one-to-one basis.

  In step S24, the region separation unit 102 sets a brightness threshold using the brightness value in the non-cell region separated in the process of step S23.

  As an example, the region separation unit 102 selects an arbitrary frame from among a plurality of microscope images captured in step S21. Then, the region separation unit 102 obtains the above-described luminance threshold by averaging the luminance values of the respective pixels in the non-cellular region in the selected frame. The brightness threshold obtained in step S24 is used by the focus position calculation unit 103 when determining the focus position at each pixel of the cell region.

  In step S <b> 25, the in-focus position calculation unit 103 obtains in-focus positions at each pixel in the cell area of the imaging screen. This focus position indicates the outline of the cell. That is, when a phase difference image (microscope image) is used, there is a characteristic that the luminance of the phase difference becomes larger as the phase change is larger. Therefore, generally, the contour portion of the cell has the maximum luminance value.

  For example, the focus position calculation unit 103 obtains a focus position at each pixel in the cell region of the imaging screen by executing first to third calculation processes described below.

  In the first calculation process, the in-focus position calculation unit 103 selects a target pixel to be processed from the cell area of the shooting screen. Then, the in-focus position calculation unit 103 acquires the luminance value corresponding to the target pixel from the data of each microscope image captured in step S21.

  In the second calculation process, the in-focus position calculation unit 103 compares each luminance value acquired in the first calculation process with the luminance threshold set in step S24. Here, in the pixel in which the luminance change due to the phase difference appears in the bright direction, the relative luminance value becomes the largest when the objective lens 89 is in the in-focus position. On the other hand, in the pixel in which the luminance change due to the phase difference appears in the dark direction, the luminance value becomes the smallest when the objective lens 89 is at the in-focus position.

  Therefore, when each luminance value acquired in the first calculation process is equal to or higher than the threshold value, the focus position calculation unit 103 performs the following process. In this case, the in-focus position calculation unit 103 obtains the maximum point of the luminance value in the spatial direction using the luminance value acquired in the first calculation process. Then, the focus position calculation unit 103 determines that the focus position of the objective lens 89 capable of acquiring the microscope image indicating the maximum point is the focus position at the target pixel. Thereby, the in-focus position in the pixel where the luminance change due to the phase difference appears in the bright direction can be obtained.

  On the other hand, when each luminance value acquired in the first calculation process is less than the luminance threshold, the in-focus position calculation unit 103 performs the following process. In this case, the in-focus position calculation unit 103 obtains the minimum point of the brightness value in the spatial direction using the brightness value acquired in the first calculation process. Then, the focus position calculation unit 103 determines the focus position of the objective lens 89 that can acquire the microscope image indicating the minimum point as the focus position at the target pixel. As a result, the in-focus position at the pixel where the luminance change due to the phase difference appears in the dark direction can be obtained.

  In the second calculation process, the in-focus position calculation unit 103 extracts the microscope image having the maximum (or minimum) luminance value at the position of the target pixel, and the objective lens 89 corresponding to the extracted microscope image. The focal position may be the in-focus position. Alternatively, the focus position calculation unit 103 obtains an interpolation curve for interpolating each luminance value at the position of the target pixel, and the focus position of the objective lens 89 based on the maximum point (or the minimum point) on the interpolation curve. May be obtained by calculation.

  In the third calculation process, the in-focus position calculation unit 103 changes the target pixel to be processed to another pixel and repeats the second calculation process, so that the objective lens 89 is aligned at each pixel in the cell region. Find each focal position. Then, the focus position calculation unit 103 generates a focus position map indicating the focus position of the objective lens 89 for each pixel in the cell region of the imaging screen.

  Thus, in step S25, by executing the first to third calculation processes, the focus position calculation unit 103 obtains the focus position at each pixel in the cell region of the imaging screen.

  In step S <b> 26, the three-dimensional information calculation unit 104 obtains the height of the contour surface of the cultured cell (the position of the contour surface of the cultured cell in the optical axis direction) based on the focus position of the objective lens 89. That is, the three-dimensional information calculation unit 104 uses a well-known basic optical equation (lens Gaussian equation) from the in-focus position of the objective lens 89 at each pixel obtained in step S25 to the pixel position. The height of the contour surface of the corresponding cultured cell is obtained. Thus, the three-dimensional information calculation unit 104 can obtain three-dimensional distance image data (point cloud data) indicating the height of the contour surface of the cultured cell. In addition, in the example of FIG. 12, the relationship between the height of the contour surface of a cultured cell and a focus position is shown in FIG.

  In step S27, the three-dimensional information calculation unit 104 determines whether each cell region grouped in step S22 is a floating cell or an adherent cell based on the height of the contour surface of the cultured cell obtained in step S26. Determine if it belongs.

  Specifically, first, the three-dimensional information calculation unit 104 acquires the bottom surface position of the culture vessel 81. For example, the three-dimensional information calculation unit 104 regards the bottom surface position of the culture vessel 81 as the bottom surface position of the contour surface in all cell regions in the imaging screen (see FIG. 14). This is because the position of the contour surface corresponding to the above condition is considered to correspond to the outer edge of the adherent cell attached to the bottom surface of the culture vessel 81.

  Then, the three-dimensional information calculation unit 104 determines that the cell surface group whose contour surface height is separated from the bottom surface position of the culture vessel 81 by a certain distance (threshold value h) or more is a floating cell (see FIG. 14). As a result, the three-dimensional information calculation unit 104 can automatically identify floating cells and adherent cells in the imaging screen.

  In step S28, the three-dimensional information calculation unit 104 estimates the total volume of adherent cells included in the imaging screen. Specifically, first, the three-dimensional information calculation unit 104 excludes, from the processing target, cells determined as floating cells in step S <b> 27 among the cell regions on the imaging screen. Next, the three-dimensional information calculation unit 104 obtains the unit pixel area in consideration of the magnification of the objective lens 89. Then, the three-dimensional information calculation unit 104 estimates the volume of each cultured cell by integrating the height of each pixel included in the cell region (adherent cell) to be processed. Thereafter, the three-dimensional information calculation unit 104 generates data of the total volume of adherent cells included in the imaging screen and records it in the memory 101.

  As described above, in the cell culture management system 11, the in-focus position of the pixel in the cell region is obtained using a plurality of phase difference observation images having different focal positions, and the three-dimensional shape of the cultured cell is determined based on the in-focus position. To get. Therefore, the observation mechanism 22 can acquire three-dimensional information of cultured cells without damaging the cells by irradiation with excitation light or staining. Moreover, since the observation mechanism 22 observes the cultured cells in the culture chamber of the cell culture apparatus 12, the cultured cells are not damaged by a change in environmental conditions during observation.

  In addition, the image processing unit 36 can obtain the three-dimensional information of the cultured cell by narrowing down the cell region in the photographing screen in advance using a plurality of phase difference observation images. It is possible to reduce the calculation load when obtaining. Furthermore, the image processing unit 36 can identify the floating cells and adherent cells and estimate the volume of the cultured cells based on the three-dimensional information of the cultured cells, and further enhance the functionality of the cell culture management system 11. It becomes possible.

  In particular, since the cell culture management system 11 can acquire a growth curve indicating the temporal transition of the total volume of adherent cells for each critical point, for example, more accurate results can be obtained for stacked cells. Observation and evaluation of the culture state can be performed.

  For example, for a patient who has already cultured cells in the cell culture management system 11, when the desired operation date and the required cell amount are specified, the schedule creation unit 32 is stored in the culture data storage unit 34. On the basis of the past culture results, it is possible to calculate the cell input time to reach the required cell amount on the desired surgery day.

  In the present embodiment, since the allowable amount for the required cell amount is set, as shown in FIG. 6, the predicted completion date range is obtained as three days, and the scheduled operation date / time is determined within the range. However, for example, when the allowable amount for the necessary cell amount is not set, the predicted completion date is obtained as one day, and the scheduled time for performing the surgery can be determined within the day.

  Furthermore, in addition to predicting the prediction completion date with reference to the culture rate stored in advance in the culture data storage unit 34, the cell culture rate is determined at the initial stage when cell culture is started (pre-check), The prediction completion date may be predicted based on the result. Moreover, it is good also considering the active state of a cultured cell with the cell amount of a cultured cell as a judgment material in a critical point.

  In addition to acquiring the value entered by the user, the cell ID and input amount of the cultured cell can be detected using, for example, a culture container with an IC tag recording the cell ID, It can be obtained by measuring the weight of the culture vessel carried into the chamber with a sensor.

  The series of processes described above can be executed by hardware or can be executed by software. When a series of processing is executed by software, a program constituting the software executes various functions by installing a computer incorporated in dedicated hardware or various programs. For example, it is installed from a program recording medium in a general-purpose personal computer or the like.

  1 is connected to the computer, and the computer includes a program stored in a ROM (Read Only Memory), a hard disk, a nonvolatile memory, and the like. A program or the like stored in the storage unit is loaded into a RAM (Random Access Memory) and executed by a CPU (Central Processing Unit). Thereby, the series of processes described above are performed.

  These programs are stored in advance in a storage unit, or via a communication unit such as a network interface, or a magnetic disk (including a flexible disk), an optical disk (CD-ROM (Compact Disc-Read Only). Memory, DVD (Digital Versatile Disc, etc.), magneto-optical disk, or drive that drives removable media such as semiconductor memory can be installed in the computer.

  The program executed by the computer may be a program that is processed in time series in the order described in this specification, or in parallel or at a necessary timing such as when a call is made. It may be a program for processing. Further, the program may be processed by one CPU, or may be processed in a distributed manner by a plurality of CPUs.

  Further, in this specification, the system represents the entire apparatus constituted by a plurality of apparatuses.

  The embodiment of the present invention is not limited to the above-described embodiment, and various modifications can be made without departing from the gist of the present invention.

It is a block diagram which shows the structural example of one Embodiment of the cell culture management system to which this invention is applied. It is a figure which shows the operation screen 41 operated when inputting or displaying the information regarding a patient. It is a figure which shows the operation screen 51 operated when inputting or displaying the information regarding a facility. It is a figure which shows the operation screen 61 operated when inputting or displaying the information regarding a doctor. It is a figure which shows the display screen 71 which displays the information regarding a cultured cell. It is a figure explaining the prediction and correction of the prediction completion date by the schedule preparation part. It is a figure which shows the schedule of an operation scheduled date. It is a flowchart explaining the process which produces and corrects a schedule. It is a figure explaining the process which calculates | requires the three-dimensional image of a cell from several images in the position where Z-axis directions differ. It is a figure which shows the three-dimensional image of a cell. 3 is a block diagram illustrating a detailed configuration example of an observation mechanism 22 and an image processing unit 36. FIG. It is a flowchart explaining a cell state observation process. It is a figure which shows an example of a microscope image, and the separation state of a cell area | region and a non-cell area | region. It is a figure which shows the example of a relationship between the height of the contour surface of a cultured cell, and a focus position.

Explanation of symbols

  11 cell culture management system, 12 cell culture device, 13 management device, 14 display device, 15 input device, 21 culture mechanism, 22 observation mechanism, 23 medium exchange mechanism, 24 passage mechanism, 25 solution addition mechanism, 31 control unit, 32 schedule creation unit, 33 schedule storage unit, 34 culture data storage unit, 35 search unit, 36 image processing unit, 41 operation screen, 51 operation screen, 61 operation screen, 71 display screen, 81 culture vessel, 82 sample stage, 83 Light source, 84 collector lens, 85 mirror, 86 field lens, 87 ring diaphragm, 88 condenser lens, 89 objective lens, 90 phase ring, 91 imaging lens, 92 lens drive unit, 93 imaging unit, 101 memory, 102 region separation unit , 10 Focusing position calculating unit, 104 three-dimensional information computing unit

Claims (4)

In a cell culture management system for culturing cells and managing the culture schedule,
Storage means for storing information about facilities and doctors;
Predicting means for predicting the arrival date when the cells are cultured and proliferated, and reach the necessary cell amount necessary for the operation using the cells;
A cell culture management system comprising: a determination unit that determines a scheduled operation date and time for performing an operation using the cell based on information on the facility and a doctor on the arrival date predicted by the prediction unit. Observation means for observing the culture state of the cell at a predetermined time after the start of the culture of the cell;
The cell culture management system according to claim 1, further comprising: a correcting unit that predicts the arrival date again according to the observation result by the observation unit, and corrects the scheduled operation date according to the arrival date. . The predicting means predicts a range of arrival dates in which the cell volume of the cells falls within an allowable range specified as an allowable range for the required cell volume,
The determination means obtains a candidate for the scheduled operation date and time based on information on the facility and doctor in the range of the arrival date and presents it to the user, and is designated by the user from the candidates for the scheduled operation date and time The cell culture management system according to claim 1 or 2, wherein a scheduled surgery date and time is determined as the scheduled surgery date and time. A culture data storage unit for storing information indicating a culture rate for each cell;
Identification information for identifying the cells, and information acquisition means for acquiring input amount information indicating a cell amount of the cells input when starting culture,
Search means for searching the culture data storage unit based on the identification information acquired by the information acquisition means, and acquiring a culture rate of the corresponding cells; and
The said prediction means predicts the said arrival date based on the culture | cultivation rate of the said cell searched by the said search means, and the said input information acquired by the said information acquisition means. 4. The cell culture management system according to any one of 3.

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