JP2005333824A - Specimen introduction device and culture treatment apparatus - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a culture treatment apparatus with which an introduction vessel in which a specimen is stored in a sealed state, the vessel is opened in the inside and the specimen in the inside is treated. <P>SOLUTION: The device is the specimen introduction device 55 with which a specimen in a stored state in the introduction vessel 56 composed of an approximately cylindrical tube openable and closable by a cap 57 tightened to a top opening in a treating chamber for subjecting a specimen to be cultured to a predetermined treatment. The device is equipped with a first gripping means 64 for receiving the introduction vessel 56 storing the specimen and gripping the side of the introduction vessel 56, a second gripping means 76 for gripping the cap 57 of the introduction vessel 56 gripped by the first gripping means 64 and a cap opening and closing means 60 for relatively displacing the first gripping means 64 from the second gripping means 76 along the longitudinal shaft of the introduction vessel while relatively rotating the first gripping means 64 or the second gripping means 76 around the longitudinal shaft of the introduction vessel 56. <P>COPYRIGHT: (C)2006,JPO&NCIPI

Description

  The present invention relates to a specimen introduction apparatus and a culture treatment apparatus.

  2. Description of the Related Art Conventionally, bone repair has been performed by filling a bone replacement material in a bone defect caused by bone tumor extraction, trauma, or the like. However, when the bone gradually becomes brittle like osteoporosis, or when the defect is very wide, it is difficult to solve the problem with the above method.

Therefore, in recent years, there has been a demand for a new attempt to collect bone marrow from a patient, artificially proliferate osteoblasts from mesenchymal stem cells contained in the collected bone marrow, and then return them to the patient's body again. ing. In this case, since osteoblasts are proliferated from the bone marrow collected from the patient and returned to the patient's body, bone formation can be activated without causing an immune reaction (see, for example, Non-Patent Document 1). ).
A cell culture device for automatically culturing mesenchymal stem cells is disclosed in Patent Document 1.
Yoshikawa, “Cultivated dermis and bone marrow with bone marrow mesenchymal cells—regenerative medicine with bone marrow mesenchymal cells”, Bioindustry, CMC Publishing Co., Ltd., 2001, Volume 18, No. 7, p. 46-53 JP 2004-119 A (FIG. 3 etc.)

In order to culture a specimen such as a mesenchymal stem cell, the specimen is collected from a patient in a medical institution such as a hospital, transported, and introduced into a culture apparatus. In each of these steps, the specimen must be maintained in a sterile state, and must be kept sealed from the outside until it is introduced into the culture apparatus after being collected.
On the other hand, in the culture apparatus, a specimen is put into a predetermined culture container and a drug such as a predetermined medium or growth factor is added, or a process such as medium exchange is performed. Therefore, the specimen needs to be open to the outside in the culture apparatus.

  However, in order to make the specimen sealed in the outside of the culture apparatus and to open in the culture apparatus, conventionally, there has been a problem that it has been necessary to work manually and the work is troublesome. Moreover, it is not preferable for an operator to enter the culture apparatus because dust is generated. Therefore, there is a need for a device that introduces the specimen into the culture apparatus in a sealed state and opens it inside the culture apparatus.

  The present invention has been made in view of the above-described circumstances, and introduces an introduction container that contains a specimen in a sealed state, opens the inside, and enables a specimen introduction apparatus and a culture process that enable processing on the inside specimen. The object is to provide a device.

In order to achieve the above object, the present invention provides the following means.
The present invention introduces a sample into a processing chamber that performs a predetermined process on a sample to be cultured, and introduces the sample in a state where it is accommodated in an introduction container made of a substantially cylindrical tube that can be opened and closed by a cap fastened to an upper end opening. A first grasping means for receiving an introduction container containing a specimen and grasping a side surface of the tube; and a second grasping means for grasping a cap of the introduction container grasped by the first grasping means. The first gripping means and the second gripping means are relatively displaced along the longitudinal axis while relatively rotating the first gripping means or the second gripping means around the longitudinal axis of the tube. Provided is a sample introduction device including a cap opening / closing means.

  According to the present invention, the introduction container accommodates the sample in the tube and fastens the cap so that the sample is introduced into the processing chamber in a sealed state. The introduction container introduced into the processing chamber grasps the side surface of the tube constituting the introduction container by the operation of the first grasping means, and the cap is grasped by the operation of the second grasping means. In this state, by operating the cap opening / closing means, the first gripping means and the second gripping means are relatively rotated around the longitudinal axis of the tube and are relatively displaced along the longitudinal axis. Thereby, the cap fastened to the tube is loosened, and the introduction container is opened in the processing chamber.

In the above invention, an identification code is attached to a side surface of the tube, an identification code reader is disposed in the processing chamber, the second gripping means is provided to be rotatable around the longitudinal axis of the tube, and the tube It is preferable to provide a moving mechanism for disposing the device opposite to the identification code reader.
By operating the moving mechanism, the tube is disposed opposite to the identification code reader, and the second gripping means is rotated around the longitudinal axis of the tube so that the identification code attached to the side of the tube is opposed to the identification code reader. It becomes possible to read the identification information indicated in the identification code.

In the above invention, the cap opening / closing means may be provided with a sensor for detecting a relative displacement along the longitudinal axis.
By detecting the relative displacement by the operation of the sensor, it becomes possible to more reliably open and close the cap with respect to the tube by the cap opening and closing means.

  Furthermore, according to the culture treatment apparatus including the sample introduction device, the sample can be introduced aseptically in a sealed state by the operation of the sample introduction device, and can be opened inside without manual intervention. it can.

  According to the present invention, the specimen contained in the tube is sealed by fastening the cap and introduced into the processing chamber, and the cap is removed from the tube in the processing chamber by the operation of the first gripping means, the second gripping means, and the cap opening / closing means. By removing and releasing the specimen in the tube, there is an effect that the specimen can be maintained in an aseptic state and the specimen can be processed without mixing dust or the like.

Hereinafter, a sample introduction apparatus, a culture treatment apparatus, and an automatic culture apparatus according to an embodiment of the present invention will be described with reference to FIGS.
As shown in FIG. 1, the automatic culture apparatus 1 according to the present embodiment is sealed by a transparent wall material that can be observed from the outside, and communicates with each other via a shutter 2. And has.

A total of four culture chambers 4 each containing two culture vessels 3 are arranged in both side spaces S11 and S13 of the first space S1, and a transfer robot (transfer) for moving the culture vessels 3 is placed in the central space S12. Mechanism) 5 is provided. In the upper part of the central space S12, there is provided an air purifying unit 6 for sending a clean descending air flow to purify the air in the central space S12.
Each of the four culture chambers 4 is disposed with a space between the two culture chambers 4 arranged side by side with the doors 4a facing each other by disposing the doors 4a toward the central space S12.

As shown in FIGS. 2 and 3, each culture chamber 4 has an opening 4b on one side surface and includes a door 4a that can open and close the opening 4b. A plurality of rail-like tray holding members 4c are provided at corresponding height positions on the left and right side walls toward the opening 4b. 7 can be accommodated in a plurality of stages in the vertical direction. Each culture chamber 4 is maintained at predetermined culture conditions such as a temperature of 37 ± 0.5 ° C., a humidity of 100%, and a CO ₂ concentration of 5%. Note that the tray holding member 4c is not limited to a rail shape, and may be in any form as long as the tray 7 can be supported in a removable manner.

A plurality of, for example, ten culture vessels 3 can be placed side by side on each tray 7. As shown in FIG. 4, each culture vessel 3 includes a vessel body 3a and a lid 3b provided on the upper surface of the vessel body 3a. A protrusion 3c that is hooked by a hand in the space is provided.
An identification code 50 indicating a unique identification code is attached to the culture container.

  Below each culture chamber 4, as shown in FIG. 1 and FIG. 2, a stocker 8 that accommodates a plurality of stages in the vertical direction with an unused culture vessel 3 mounted on a tray 7 is disposed. A lift mechanism (not shown) is provided in the stocker 8, and after the uppermost tray 7 is taken out, the lower tray 7 is sequentially lifted, and the tray 7 to be next taken out is arranged at the uppermost supply position. Can be kept.

  An identification code (not shown) indicating a unique identification code is also attached to each tray. The identification code 50 of the culture container 3 and the identification code of the tray 7 are not associated with each other when accommodated in the stocker 8, and the culture container 3 having the arbitrary identification code 50 on the tray 7 having the arbitrary identification code. Is installed.

  As shown in FIG. 2, the stocker 8 has a door 51 that can be opened and closed on a side surface facing the outside of the first space S <b> 1 opposite to the door of the culture chamber 4. The door 51 is formed in a size that opens one entire side surface of the stocker 8. The culture vessel 3 mounted on the tray 7 can be inserted into the stocker 8 from the outside through the door 51. Further, the stocker 8 is provided with a carry-in port 52 that opens to the door side of the culture chamber 4 at a position corresponding to the uppermost tray 7 in the stocker 8. The transfer robot 5 takes out the unused culture container 3 accommodated in the stocker 8 from the carry-in port 52 together with the tray 7 and moves it to the culture treatment apparatus 30 described later.

  The transfer robot 5 is arranged approximately at the center of the interval between the four culture chambers 4. The transfer robot 5 includes a first arm 5a that can rotate horizontally, a second arm 5b that is connected to the tip of the first arm 5a so as to be rotatable about a vertical axis, and a vertical axis that is connected to the tip of the second arm 5b. A hand 5c that is rotatably mounted around itself and does not have a mechanism for deteriorating the environment in the culture chamber such as a drive unit and a conduction mechanism, and the first arm 5a, the second arm 5b, and the hand 5c can be moved up and down. And an elevating mechanism 5d. As a result, the transfer robot 5 accesses all the trays 7 in the four culture chambers 4 and on the conveyor 9 disposed between the first space S1 and the second space S2 across the shutter 2. It has a horizontal operating range in which the tray 7 can be delivered.

Further, the transfer robot 5 has a vertical movement range that reaches the carry-in port 52 provided in the stocker 8, and takes out the unused culture vessel 3 mounted on the tray 7 from the carry-in port 52, It can be moved to the culture treatment apparatus 30.
Further, the transfer robot 5 is provided with an identification code reader 53 for reading the identification code of the tray 7 and the identification code (not shown) attached to each stage of the culture chamber 4.

  The hand 5c is formed in a flat shape extending in the horizontal direction so that the tray 7 can be placed, and is formed in a thickness dimension that can be inserted into a gap between the trays 7 accommodated in the culture chamber 4. . The hand 5c is lifted from the state inserted in the gap between the trays 7, so that the tray 7 is pushed up from below by the two arms and taken up from the tray holding member 4c, and the tray 7 is stably held. It can be done.

The conveyor 9 includes two endless belts 9a arranged on the left and right sides with a gap larger than the width dimension of the hand 5c of the transport robot 5, so that the tray 7 can be placed over the endless belts 9a. It has become. The transfer robot 5 has access to all the trays 7 in the culture chamber 4 and has a vertical movement range that reaches the carry-in port 52 provided in the stocker 8. The unused culture vessel 3 mounted on 7 can be taken out and moved to the culture treatment apparatus 30.
The belt 9a is not limited to an endless belt.

As shown in FIG. 1, a culture treatment apparatus 30 is configured in the second space S2. The culture treatment apparatus 30 includes, for example, a bone marrow fluid introduction unit 55 (sample introduction device) into which a bone marrow fluid 54 collected from a patient at a medical institution or the like is introduced.
As shown in FIG. 5, the bone marrow fluid 54 is accommodated in a container 56 (for example, a centrifuge tube that can be centrifuged as it is: hereinafter referred to as an introduction container 56), and a lid 57 (cap) is used as an upper opening. By being fastened, it is introduced into the bone marrow fluid introduction part 55 in a closed state. An identification code 58 is affixed to the outer peripheral surface of the introduction container 56. The identification code 58 is assigned an identification code that can identify unique information of the bone marrow fluid 54, for example, the name of the patient who collected the bone marrow fluid 54.

  As shown in FIGS. 6 to 12, the bone marrow fluid introduction unit 55 supplies the bone marrow fluid 54 containing the introduction container 56 into the second space S <b> 2, and the empty introduction container 56 or the cultured cells as a product. Alternatively, the supply / discharge unit 59 that discharges the introduction container 56 enclosing the biological tissue filling body from the second space S2 to the outside, the lid opening / closing mechanism 60 that attaches / detaches the lid 57 of the introduction container 56, and the introduction container 56 are centrifuged. An elevating mechanism 61 that supplies the separator 11 is provided.

  As shown in FIGS. 6 to 9, the supply / discharge unit 59 sends a supply container detainer 62 capable of detaining a plurality of introduction containers 56 containing bone marrow fluid 54 and the introduced introduction containers 56 one by one. A feeding mechanism 63, a grasping hand 64 that receives the introduced introduction container 56, a hand moving mechanism 65 that moves the grasping hand 64 in the horizontal direction, and an empty introduction container 56 from which the bone marrow fluid 54 has been taken out, or A discharge container indwelling unit 66 capable of indwelling a plurality of introduction containers 56 in which cells or biological tissue complements after culture as a product are enclosed is provided.

  The supply container detainer 62 and the discharge container detainer 66 are respectively a horizontal placing part 67 for placing the introduction container 56, and a support rail part disposed above the placing part 67 and supporting the side surface of the introduction container 56. 68, and each of the plurality of introduction containers 56 is arranged in a line with the lid body 57 facing upward. The supply container detainer 62 and the discharge container detainer 66 are arranged in parallel to each other.

  The feeding mechanism 63 includes a horizontally movable slider 69, a linear moving mechanism 70 that horizontally moves the slider 69, and pressing heads 71 and 72 that are mounted on the slider 69 and press the introduction container 56. Two pressing heads 71, 72 are provided on the slider 69, and one pressing head 71 presses the introduction container 56 mounted on the supply container detainer 62 toward the gripping hand 64, and the other pressing head 72. Is configured to press the introduction container 56 mounted on the discharge container indwelling portion 66 so as to separate it away from the gripping hand 64.

  The pressing heads 71 and 72 include pressing arms 71a and 72a, respectively, and the pressing arms 71a and 72a are swung around the horizontal axis by motors 71b and 72b. As shown in FIG. 8, the pressing arms 71a and 72a are swung between a protruding position (solid line) arranged substantially horizontally and a retracted position (chain line) inclined with respect to the protruding position. ing. In the projecting position, the introduction container 56 mounted on the supply container detainer 62 or the discharge container detainer 66 is disposed within the operation locus of the pressing arms 71a and 72a when the linear movement mechanism 70 is operated. It has become. On the other hand, at the retracted position, the movement locus does not overlap with the introduction container 56.

  The gripping hand 64 has two claws 73 that can approach and separate in the horizontal direction, and is configured to sandwich the side surface of the introduction container 56 fed by the feeding mechanism 63 in the diametrical direction. Further, as shown in FIG. 7, the gripping hand 64 is provided with a cradle 74 that supports the bottom surface of the introduction container 56 that has been fed on the placement part 67 of the supply container detainer 62 by the feed mechanism 63. ing. Thereby, the introduction container 56 grasped by the grasping hand 64 is sandwiched between the two claws 73 from the horizontal direction and is supported by the receiving base 74 on the bottom surface thereof.

  Further, as shown in FIGS. 6 and 9, the hand moving mechanism 65 is capable of moving the introduction container 56 gripped by the gripping hand 64 to the horizontal positions A to D. That is, the receiving position B of the introduction container 56 sent from the supply container detainer 62, the delivery position A of the introduction container 56 to the discharge container detainer 66, the opening and closing of the introduction container 56, and the centrifuge 11 described later. It can be arranged as needed at the supply position C and the processing position D such as dispensing for the introduction container 56 with the lid 57 opened.

  In the hand moving mechanism 65, as shown in FIG. 6, optical sensors 75a and reflecting plates 75b are arranged opposite to each other on both ends of the movement locus of the introduction container 56 by the grasping hand 64. When the introduction container 56 does not exist in the movement trajectory by the gripping hand 64, the light emitted from the optical sensor 75a is reflected by the reflection plate 75b and detected by the optical sensor 75a. When it exists in position AD, the detection of the light by the optical sensor 75a is not performed, and, thereby, the presence of the introduction container 56 can be recognized.

As shown in FIGS. 10 and 11, the lid opening / closing mechanism 60 includes a chuck mechanism 76 that grips the lid 57 of the introduction container 56 that is gripped by the gripping hand 64, and a lid 57. A rotating mechanism 77 that rotates the gripped chuck mechanism 76 around the longitudinal axis of the introduction container 56 is provided.
As shown in FIG. 11, the chuck mechanism 76 includes three claws 78 arranged at intervals of 120 °. These claws 78 are configured to be close to and separated from each other toward the central axis. Each claw 78 is provided with an abutting portion 78 a that abuts on the upper surface of the lid 57. In FIG. 10, only one claw 78 is shown for clarity of illustration.

  The rotation mechanism 77 includes a motor 79 and a spline shaft 80 that is rotationally driven by the motor 79. The spline shaft 80 engages the chuck mechanism 76 in the circumferential direction and rotates it around the longitudinal axis, and allows movement along the axial direction of the chuck mechanism 76, that is, vertical movement. The chuck mechanism 76 is always urged downward by the compression spring 81. Reference numeral 82 in the drawing is a sensor. The sensor 82 can detect when the chuck mechanism 76 is displaced upward by a predetermined amount against the pressing force of the compression spring 81. Reference numerals 83 and 84 denote sensors for detecting the rotation angle of the motor 79 and the rotation angle of the spline shaft 80.

  Accordingly, the lid opening / closing mechanism 60 removes the lid 57 from the introduction container 56 by gripping and rotating the lid 57 of the introduction container 56 gripped by the gripping hand 64 with the claws 78, or A lid 57 is fastened to the introduction container 56. Further, the lid opening / closing mechanism 60 releases the side gripping state by the gripping hand 64, and moves the introduction container 56 up and down by the operation of the lifting mechanism 61 in the suspended state in which only the lid 57 is gripped by the chuck mechanism 76. It can be moved in the direction. Furthermore, the introduction container 56 can be rotated around the longitudinal axis in the suspended state by operating the rotation mechanism 77 with only the lid 57 held.

  As shown in FIG. 12, the elevating mechanism 61 is driven by a slider 85 on which the lid opening / closing mechanism 60 is mounted, a linear guide 86 for guiding the slider 85 in the vertical direction, a motor 87, and a motor 87. The endless steel belt 88 is provided. The slider 85 is fixed to the steel belt 88 and is moved in the vertical direction along the linear guide 86 as the steel belt 88 moves. The centrifuge 11 is arranged vertically below the lid opening / closing mechanism 60. The centrifuge 11 receives the introduction container 56 lowered by the elevating mechanism 61, and centrifuges bone marrow cells containing a lot of mesenchymal stem cells from the bone marrow fluid 54. It is supposed to be.

  Further, as shown in FIG. 9, an identification code reader 89 that is arranged to face the side surface of the introduction container 56 is provided in the middle of the raising and lowering by the raising and lowering mechanism 61. When the introduction container 56 in a suspended state in which only the lid body 57 is gripped by the chuck mechanism 76 is rotated around its vertical axis by the operation of the rotation mechanism 77 in the vicinity of the identification code reader 89, it is adhered to the side surface. The identification code 58 can be read by the identification code reader 89.

  The opened introduction container 56 is supplied with PBS (phosphate buffered saline) and stirred by a method such as pipetting, and is then transported to the centrifuge 11 by the lifting mechanism 61 and put in. It has become so. The centrifuge 11 receives the bone marrow fluid 54 received from the bone marrow fluid introduction part 55 and the introduction container 56 containing the PBS mixed solution, and rotates at a low speed, whereby the specific gravity floating in the mixed solution of PBS and bone marrow fluid 54 is suspended. Heavy bone marrow cells are separated from PBS and other body fluids and allowed to settle.

  As shown in FIG. 1, the culture treatment apparatus 30 further applies the above-described unused culture vessel 3 on the tray 7 that has been conveyed by the conveyor 9 from the first space S1 with the shutter 2 opened. A supply / exhaust robot 10 is provided for supplying bone marrow cells collected in the centrifuge 11 or supplying and collecting a medium. In addition, the culture treatment apparatus 30 is provided with an identification code reader (not shown) that reads the identification code 50 of the culture container 3 after bone marrow cells are supplied. The bone marrow fluid identification code 58 and the culture vessel identification code 50 read by these identification code readers are sent to a control device (not shown) and stored in association with each other. In addition, the identification code 50 of the culture vessel 3 and the identification code of the tray 7 are also stored in association with each other by the control device after being introduced into the culture treatment apparatus 30.

Furthermore, the culture treatment apparatus 30 includes four dispensing robots 13 that are equipped with an electric pipette 12 for dispensing various liquids such as serum and reagents, and that can be rotated and moved up and down, and these supply and discharge robots 10 and 10. Three tip supply devices 15 that contain a plurality of disposable tips 14 attached to the tip of the electric pipette 12 of the dispensing robot 13 and can be provided within the operating range of the supply / discharge robot 10 and the dispensing robot 13, and used A chip collection unit 31 that discards and collects the chip 14, a reagent supply device 16 that stores various liquids such as serum and reagents in a plurality of containers, and a microscope 17 that can observe the state of cells in the culture container 3. Culturing between a plurality of storage tanks 18 for storing waste liquids discarded due to each reagent and medium exchange, and between the conveyor 9 and the robots 10 and 13. A horizontal movement mechanism 19 for moving the culture vessel 3 so that the vessel 3 can be delivered, and a mounting table 21 mounted on the slider 20 of the horizontal movement mechanism 19 for placing the received culture vessel 3 thereon. .
Note that the second space S2 is also provided with an air purifier 32 that forms a clean downward airflow in order to purify the air in the second space S2.

  The culture treatment apparatus 30 configured in the second space S2 is arranged at an intermediate position in the height direction, and a first partition wall 33 that vertically partitions the second space S2 into an upper space S21 and a lower space S22. Are divided into three spaces S21, S221, and S222 arranged in the vertical direction by a second partition wall 34 that further partitions the lower space S22 formed by the first partition wall 33 vertically. The first partition wall 33 is disposed at the height of the conveyor 9, and in the upper space S21 above the first partition wall 33, the loading table 21, the supply / discharge robot 10, the arm 13a of the dispensing robot 13, and the XY table 17a of the microscope 17 are provided. The above mechanism part etc. are arranged. This is because these apparatuses are necessary for moving the culture container 3 and need to be accessed from the upper opening of the culture container 3. Note that the upper surface of the reagent supply device 16 is also exposed on the upper surface of the first partition wall 33 in order to open the insertion port 16c of the chip 14 in the upper space S21.

  The first partition wall 33 has a long hole 35 for connecting the mounting table 21 to the horizontal movement mechanism 19 in the lower space S21 in order to move the mounting table 21 in the upper space S21. A through hole 36 for taking out the chip 14 from the chip supply device 15 arranged in the space S221 below the wall 33 and a disposal port 37 for discarding the used chip 14 are formed therethrough. Further, the first partition wall 33 is provided with vent holes 38 penetrating vertically along the side walls 30a and 30b (shaded portions).

  In the space S221 between the first partition wall 33 and the second partition wall 34, as shown in FIG. 13, the main body portion of the dispensing robot 13, the chip supply device 15, the reagent supply device 16, the microscope, and the like. 17 XY table 17 a and lower portions, a horizontal movement mechanism 19, and a duct 40 that connects the waste outlet 37 and the waste container 39 of the chip collection unit 31 are provided. As shown in FIG. 14, the duct 40 has a structure including a flange portion 40 a at the upper end, for example, and is hooked on a hook 44 provided at a lower portion of the first partition wall 33, thereby the first partition wall. What is necessary is just to provide between 33 and the 2nd partition wall 34 so that attachment or detachment is possible. In the vicinity of the side walls 30a and 30b of the second partition wall 34, vent holes 43 penetrating vertically are provided along the side walls 30a and 30b (shaded portions).

  Further, in the space S222 below the second partition wall 34, as shown in FIG. 15, the centrifuge 11, the storage tank 18, the waste container 39, and the exhaust fan 41 are arranged. A filter 42 such as a HEPA filter is provided at the outlet of the exhaust fan 41 so as to clean the exhausted air.

  The supply / discharge robot 10 is a horizontal articulated robot. For example, in the example shown in FIG. 1, a head 10c including two types of electric pipettes 10a and 10b, and two arms 10d and 10e capable of horizontal turning are provided. And an elevating mechanism 10f that elevates and lowers the head 10c. The electric pipette 10a supplies a medium guided from the storage tank 18 through the duct 10g, or performs a pipetting operation. The electric pipette 10b sucks unnecessary medium in the culture vessel 3 or the centrifuge vessel, and discharges it as waste liquid to the other storage tank 18 through the duct 10g.

  The pipetting operation by the electric pipette 10a is performed on the bone marrow fluid 54 infused with PBS and also on the mixed solution of the cells and the medium constituting liquid separated by the centrifuge 11. ing. In the pipetting operation, the tip 14 supplied from the tip supply device 15 is attached to the tip of the electric pipette 10a, inserted into the introduction container 56, and the bone marrow fluid 54 and phosphate buffer stored in the introduction container 56. Aspirate and release are repeated 10 to 20 times for the mixed solution with chlorinated saline or the mixed solution of cells and medium. Thereby, a liquid mixture is stirred uniformly.

  In addition, after the pipetting operation, the supply / discharge robot 10 sucks the mixed solution of the cells and the medium separated by the centrifuge 11 with the electric pipette 10a, and the inside of the culture container 3 mounted on the mounting table 21. Is supplied from the upper opening.

  The used chip 14 once used is removed and collected by the chip collection unit 31. Accordingly, the supply / discharge robot 10 operates in various operating ranges such as the bone marrow fluid introduction unit 55, the mounting table 21, the chip supply device 15, the chip collection unit 31, and the cell supply device (not shown) from the centrifuge 11. It is placed inside.

  Each of the dispensing robots 13 includes a horizontally rotatable arm 13a provided with an electric pipette 12 to which a tip 14 is detachably attached at a tip, and an elevating mechanism 13b for raising and lowering the arm 13a. The dispensing robot 13 supplies a culture medium and various reagents into the culture vessel 3 that has been transported by the horizontal movement mechanism 19. Therefore, the dispensing robot 13 arranges various devices such as the mounting table 21, the chip supply device 15, the chip recovery unit 31, and the reagent supply device 16 on the horizontal movement mechanism 19 within the operation range.

  The tip supply device 15 accommodates a plurality of tips 14 in an arrayed state in a container 15a opened upward with the attachment ports to the electric pipettes 10a, 10b, 12 facing upward. Note: When the robot 13 needs a new tip 14, the tip 14 is attached to the tip of the electric pipette 10a, 10b, 12 simply by inserting the electric pipette 10a, 10b, 12 from above. Yes. The container 15a is attached to the moving mechanism 15b so as to be reciprocated in a direction intersecting the moving direction of the electric pipettes 10a, 10b, and 12 by the supply / discharge robot 10 and the dispensing robot 13. The tip supply device 15 that supplies the tip 14 to the dispensing robot 13 is provided with another moving mechanism 15c that moves the container 15a in a direction orthogonal to the moving direction of the moving mechanism 15b. Thus, the electric pipettes 10a, 10b, and 12 can access all the tips 14 in the container 15a.

The tip collection unit 31 includes a gripping device (not shown) that grips the tip 14 at the entrance of the disposal container 39, and the tip 14 used in the supply / discharge robot 10 and the dispensing robot 13 is inserted into the gripping device. When it is done, it is designed to hold it. In this state, the supply / discharge robot 10 and the dispensing robot 13 move the electric pipettes 10a, 10b, 12 to remove the used tips 14 from the tips of the electric pipettes 10a, 10b, 12 and put them in the disposal container 39. It is collected via the duct 40. The disposal container 39 is detachably disposed in the space S222 and can be replaced as necessary.
When exchanging the duct 40 and the waste container 39, the door may be accessed from the outside of the culture treatment apparatus 30 by opening a door (not shown) provided on the side walls 30a and 30b of the culture treatment apparatus 30.

For example, as shown in FIG. 13, the reagent supply device 16 accommodates a horizontally rotatable table 16 a inside a cylindrical casing, and a cylindrical shape having a fan-shaped bottom shape on the table 16 a. A plurality of the reagent containers 16b are arranged and mounted in the circumferential direction. The inside of the casing is kept cool at a constant temperature. Various reagents etc. are stored in each reagent container 16b. For example, MEM (Minimal Essential, which constitutes a medium necessary for culturing cells)
Medium: Minimum essential medium), DMEM (Dulbecco's Modified)
Such as Eagle Medium), FBS (Fetal Bovine Serum), serum such as human serum, proteolytic enzyme such as trypsin that peels off cells in the culture vessel 3, and cytokines that grow cells during culture A growth factor, a differentiation-inducing factor such as dexamethasone that differentiates cells, an antibiotic such as a penicillin antibiotic, a hormone such as estrogen, and a nutrient such as vitamin are stored.

  An insertion port 16 c through which the dispensing robot 13 inserts the tip 14 at the tip of the electric pipette 12 is provided on the upper surface of the casing of the reagent supply device 16. The insertion port 16 c is disposed within the operation range of the dispensing robot 13. Each reagent container 16b includes an opening (not shown) disposed on the upper surface thereof at a position corresponding to the insertion port 16c. As a result, the dispensing robot 13 displaces the tip 14 at the tip of the electric pipette 12 from above by placing the opening of the reagent container 16b vertically below the insertion port 16c of the casing by rotating the table 16a. The reagent etc. which are inserted in 16b and are stored inside can be aspirated. The two reagent supply devices 16 are provided because a chemical solution such as trypsin common to the sample and a liquid such as serum inherent to the sample are separated and handled.

  The microscope 17 is used when observing the state of cells in the culture vessel 3 and the degree of proliferation or counting the number of cells in the middle of the culture process or when exchanging the medium. It has become. The XY stage 17a, working distance adjustment, magnification change, and the like of the microscope 17 can all be performed by remote control. It may be possible to observe the state of the cells in the culture vessel 3 from the outside of the automatic culture apparatus 1 by arranging an eyepiece lens toward the outside of the second space S2.

  For example, the storage tank 18 stores MEM, PBS, or the like that can be used in common for all the specimens, and supplies it to the reagent container 16b in the reagent supply apparatus 16 as necessary. Yes. In addition, the storage tank 18 may be a waste liquid tank that stores waste medium and the like that are discharged during medium replacement.

  The horizontal movement mechanism 19 includes a slider 20 that can be moved in the horizontal direction by a linear movement mechanism. The mounting table 21 is mounted on the slider 20, and the culture vessel 3 mounted on the mounting table 21 can be moved from the conveyor 9 to the operating range of the dispensing robot 13.

The mounting table 21 includes a holding mechanism (not shown) for mounting and holding the culture vessel 3 transferred from the tray 7 on the conveyor 9. Moreover, you may provide the vibration apparatus (illustration omitted) which gives this culture | cultivation container 3 a vibration. For example, in addition to a device that reciprocally swings the culture vessel 3 within a predetermined angle range, a device that applies ultrasonic vibration or a device that applies horizontal vibration may be employed as the vibration device.
A control device (not shown) is connected to various devices of the automatic culture device 1 according to the present embodiment. The control device controls the order of each process, operation timing, and the like, and records and saves operation history and the like.

The operation of the culture treatment apparatus 30 and the automatic culture apparatus 1 according to the present embodiment configured as described above will be described below.
In order to culture mesenchymal stem cells using the automatic culture apparatus 1 according to this embodiment, first, the bone marrow fluid 54 collected from the patient is introduced into the culture treatment apparatus 30 in a state where it is placed in the introduction container 56. The bone marrow fluid introduction unit 55 is supplied.

  In the bone marrow fluid introduction part 55, the introduced introduction container 56 is temporarily placed in the supply container holding part 62. Then, the feeding mechanism 63 is operated to sequentially feed the gripping hand 64 in the order of introduction. At this time, the pressing arm 71 a provided on the pressing head 71 is disposed at the protruding position, and the linear moving mechanism 70 is operated to press the rearmost introduction container 56, thereby sending it to the gripping hand 64.

  When the gripping hand 64 receives the foremost introduction container 56 containing the bone marrow fluid 54 that has been sent onto the receiving base 74, the gripping hand 64 operates the claw 73 to sandwich the side surface of the introduction container 56. As a result, the introduction container 56 is stably supported by the gripping hand 64 at three points. In this state, by operating the hand moving mechanism 65, the introduction container 56 is disposed at the lid opening / closing position C.

  When the introduction container 56 is disposed at the lid opening / closing position C, the lid opening / closing mechanism 60 is brought close to the lid 57 from above the introduction container 56 by the operation of the lifting mechanism 61. When the elevating mechanism 61 lowers the lid opening / closing mechanism 60 to a position where the contact portion 78 a provided on the claw 78 of the chuck mechanism 76 contacts the upper surface of the lid 57, the chuck mechanism 76 of the lid opening / closing mechanism 60. Is operated, and the lid body 57 is gripped from the side surfaces by the three claws 78.

  In this state, by operating the rotation mechanism 77, the chuck mechanism 76 is rotated counterclockwise around the vertical axis. Since the gripping hand 64 grips the side surface of the introduction container 56 and the chuck mechanism 76 grips the side surface of the lid body 57, the lid body 57 is relatively moved relative to the introduction container 56 by rotating the chuck mechanism 76. The screw that has been fastened together and loosened is loosened.

  The chuck mechanism 76 is always pressed downward by the compression spring 81 and is allowed to move in the vertical direction by the spline shaft 80. Therefore, when the screw is loosened by the operation of the rotation mechanism 77, the chuck mechanism 76 follows the lead of the screw. The lid 57 rises while rotating. At this time, the chuck mechanism 76 holding the lid 57 is also raised against the urging force of the compression spring 81. When the lid 57 is raised to a position where the screws of the introduction container 56 and the lid 57 are completely loosened, the sensor 82 detects this and the operation of the rotation mechanism 77 is stopped. In this state, the elevating mechanism 61 is operated, and the lid opening / closing mechanism 60 in a state where the lid 57 is held by the chuck mechanism 76 is raised, whereby the lid 57 is removed from the introduction container 56, and the introduction container 56 upper openings will be opened.

  At this time, the lid 57 is lifted away from the introduction container 56 by the operation of the elevating mechanism 61, so that the lid 57 is pushed up against the pressing force of the compression spring 81 from the introduction container 56. Since the pressing force is released, the detection state of the sensor 82 is released.

  Next, the opened introduction container 56 is moved to the dispensing position D by the operation of the hand moving mechanism 65. Then, the PBS is supplied from the opened upper opening of the introduction container 56 by the operation of the supply / discharge robot 10 and is agitated by pipetting operation. Then, the upper opening of the introduction container 56 returned to the lid opening / closing position C by the operation of the hand moving mechanism 65 again is closed by the lid 57 held by the chuck mechanism 76 of the lid opening / closing mechanism 60.

  At this time, when the lid 57 is lowered to a predetermined position by the operation of the elevating mechanism 61, the pressing force from the introduction container 56 is generated again, and the sensor 82 senses this. Then, when the lid 57 is rotated clockwise around the vertical axis by the operation of the rotation mechanism 77, the lid 57 is fastened to the screw of the introduction container 56. Then, when the lid 57 is lowered according to the lead of the screw in a state where both are sufficiently fastened, the detection state of the sensor 82 is released again, thereby confirming that the lid 57 is fastened.

The introduction container 56 to which the lid body 57 is fastened is released from the gripping state by the gripping hand 64, is lifted slightly by the lifting mechanism 61 while being gripped by the chuck mechanism 76, and is gripped by the hand moving mechanism 65. With the hand 64 retracted, the lifting mechanism 61 is operated to move vertically downward.
Since the identification code reader 89 is provided in the middle of the downward movement by the elevating mechanism 61, the introduction container 56 rotates around the vertical axis by operating the rotation mechanism 77 within the reading range of the identification code reader 89. Be made. As a result, the identification code 58 provided on the side surface of the introduction container 56 is read by the identification code reader 89 regardless of the position where the identification code 58 is attached. Thereafter, the lifting mechanism 61 is further operated to deliver the introduction container 56 to the centrifuge 11. Then, the centrifuge 11 is operated at 800 to 1300 G for 3 to 5 minutes, for example, so that the cells including the mesenchymal stem cells having a high specific gravity in the mixed solution are collected in a settled state.

  In this state, after the introduction container 56 is taken out of the centrifuge 11 again by the chuck mechanism 76 and raised to above the gripping hand 64, the gripping hand 64 is placed below the introduction container 56 by the hand moving mechanism 65. Then, the introduction container 56 is placed on the cradle 74 of the gripping hand 64 by the lifting mechanism 61. Then, the side surface of the introduction container 56 is grasped by the grasping hand 64, the chuck mechanism 76 is rotated by the operation of the rotation mechanism 77, the lid body 57 of the introduction container 56 is opened, and the introduction container 56 whose upper opening is opened is dispensed. Move to position D.

Then, by operating the supply / discharge robot 10 and the tip supply device 15, the unused tip 14 is attached to the tip of the electric pipette 10b.
That is, the tip supply device 15 operates the moving mechanism 15 b to place the unused tip 14 within the operation range of the supply / discharge robot 10. Then, the supply / discharge robot 10 operates the elevating mechanism 10f to lower the head 10c to receive the unused chip 14 from the chip supply device 15 below the first partition wall 33, and the tip of the electric pipette 10b. Attach to.

  In this state, the supply / discharge robot 10 is operated to insert the tip of the tip 14 of the electric pipette 10b into the introduction container 56 for suction. Thereby, the supernatant liquid in the introduction container 56 is removed, and cells containing a lot of mesenchymal stem cells are left in the introduction container 56.

Next, the specimen remaining in the introduction container 56 is put into the culture container 3 by the supply / discharge robot 10.
Prior to charging, the empty culture container 3 placed on the tray 7 is pushed out from the first space S1 to the second space S2 by the operation of the conveyor 9. The culture vessel 3 on the tray 7 is placed on the placement table 21 by the operation of a transfer device (not shown). Then, the lid 3b of the culture vessel 3 on the mounting table 21 is opened by the operation of a lid opening / closing device (not shown).

  In this state, the supply / discharge robot 10 is operated to supply a predetermined amount of medium constituent liquid such as DMEM or PBS stored in the storage tank 18 from the electric pipette 10a into the introduction container 56. Pipetting is performed by operating the electric pipette 10a while holding the tip 14 at the tip of the electric pipette 10a inserted into the introduction container 56. Thereby, a cell suspension in which cells containing a large amount of mesenchymal stem cells and the medium constituent liquid are uniformly mixed is formed.

  The cell suspension thus produced is sucked into the chip 14 by operating the electric pipette 10a. The sucked cell suspension is put into the culture vessel 3 on the mounting table 21 from the upper opening by operating the supply / discharge robot 10 while being held in the chip 14.

  The empty introduction container 56 from which all of the internal cell suspension has been sucked is moved to the discharge container indwelling portion 66 by the operation of the hand moving mechanism 65 after the lid body 57 is fastened by the lid body opening / closing mechanism 60. Then, in the state where the pressing head 72 having the pressing arm 72a as the retracted position is moved to the position indicated by the chain line in FIG. 6, the gripping hand 64 is operated by the operation of the linear moving mechanism 70 by the pressing arm 72a set to the protruding position. By being pushed away from the head, it is discharged to the outside.

  On the other hand, when the cell suspension is completely put into the culture container 3, the supply / discharge robot 10 inserts and removes the chip 14 into the waste opening 37 formed in the first partition wall 33, Collect. The chip 14 removed at the disposal port 37 is put into a disposal container disposed in the lowest space S222 through the duct 40.

  Next, the culture vessel 3 charged with the cell suspension is moved horizontally along with the mounting table 21 by operating the horizontal movement mechanism 19, and is placed within the operation range of each dispensing robot 13. The dispensing robot 13 operates the electric pipette 12 with the unused tip 14 received from the tip supply device 15 attached to the tip thereof, thereby allowing DMEM, serum, or various kinds of substances from the reagent container 16b of the reagent supply device 16. After a suitable amount of reagent is aspirated, the reagent is conveyed to above the culture vessel 3 and injected into the culture vessel 3. Serum and each reagent are aspirated by exchanging the chip supply device 15 with an unused chip 14 for each aspiration of each reagent and the like. As a result, in the culture vessel 3, the mesenchymal stem cells are present in a mixed state in an appropriate medium. In addition, in order to distribute mesenchymal stem cells uniformly in the medium, the mounting table 21 may be operated to vibrate the entire culture vessel 3.

  Then, the culture vessel 3 that has completed all the treatments is moved to the vicinity of the conveyor 9 by the operation of the horizontal movement mechanism 19, and there again, the operation is performed by the lid body 3 b by the operation of the lid body opening / closing device and the transfer device. With the opening closed, it is returned to the tray 7.

Thereafter, by operating the conveyor 9, the culture vessel 3 placed on the tray 7 is inserted from the second space S2 into the central space S12 of the first space S1.
In this state, by operating the transfer robot 5, the tray 7 is lifted by the hand 5c. When the tray 7 is conveyed to the front of the culture chamber 4, the door 4 a of the culture chamber 4 is opened, and the tray 7 is inserted onto the empty tray holding member 4 c by the transport robot 5. Then, by closing the door 4a again, cells are cultured while maintaining the culture conditions in the culture chamber 4 constant. Needless to say, the order of the cell suspension, the DMEM, the serum, and the various reagents and the aspiration may be changed as appropriate.

In addition, when exchanging the medium and the container, the culture container 3 in the culture chamber 4 is taken out together with the tray 7 by the operation of the transfer robot 5 arranged outside the culture chamber 4 in the same manner as described above. Passed from the first space S1 to the second space S2.
When the culture in one culture vessel 3 is sufficiently performed, it is necessary to enlarge the bottom area of the culture vessel 3. In this case, the culture container 3 taken out together with the tray 7 from the culture chamber 4 is transferred from the first space S1 to the second space S2 by the operation of the transfer robot 5. In the second space S2, trypsin is injected into the culture vessel 3 and the cells in the culture vessel 3 are peeled off. It is put into the machine 11. Then, only necessary ones such as mesenchymal stem cells are collected by the operation of the centrifuge 11, and the supernatant liquid containing trypsin is discarded.

  Then, by the operation of the supply / discharge robot 10 with the chip 14 replaced, the medium constituent liquid is supplied again into the introduction container 56 and stirred by pipetting to form a cell suspension. During this time, the culture container 3 that has been evacuated by sucking the cells is returned to the first space S1 together with the tray 7 with the lid 3b covered, and is moved from the container discharge port (not shown) by the operation of the transport robot 5. One space S1 is discharged outside and discarded.

Then, again by the operation of the transfer robot 5, the unused culture container 3 accommodated in the stocker 8 is taken out from the stocker 8 together with the new tray 7, and is supplied into the second space S2. At this time, the culture container 3 on the tray 7 may be supplied with a large one-size culture container 3, but in the automatic culture apparatus 1 according to the present embodiment, the number of culture containers 3 of the same size is increased. To supply. Thereby, the bottom area required for cell culture can be increased.
Other processing steps are the same as described above.

  And a mesenchymal stem cell is proliferated to sufficient cell number by performing the culture | cultivation process over a predetermined period through several times of culture medium exchanges and container exchanges. Whether or not the sufficient number of cells has been reached is observed or measured by transporting the culture vessel 3 with mesenchymal stem cells adhering to the bottom surface to the microscope 17 by the operation of the feeding and discharging robot 10, and the cell proliferation is increased. The degree is judged. In addition, the culture container 3 of the same sample may be mounted on the tray 7, or the culture containers 3 of different samples may be mixed. In addition, the culture container 3 of the same specimen may be placed on the mounting table 21, or the culture containers 3 of different specimens may be mixed.

  In this way, the automatic culture apparatus 1 according to the present embodiment can automatically cultivate mesenchymal stem cells having a sufficient number of cells from the bone marrow fluid 54 collected from the patient. In addition, after sufficient mesenchymal stem cells are obtained, a biological tissue supplement such as calcium phosphate and a differentiation-inducing factor such as dexamethasone are introduced into the culture vessel 3, and the culture process is continued again. You may decide to manufacture the biological tissue filling body which can be compensated for the defect | deletion part of a biological body.

Thus, according to the bone marrow fluid introduction part 55 and the culture treatment apparatus 30 according to the present embodiment, the bone marrow fluid 54 is accommodated in the introduction container 56 and charged into the supply container detainer 62 in a state of being sealed by the lid 57. By simply opening the lid 57 in the second space S2, the cells inside can be dispensed into the culture vessel 3 and subjected to a predetermined process.
In addition, since the identification code 58 can be read by the identification code reader 89 using the rotation mechanism 77 of the lid opening / closing device 60, the apparatus can be simplified.

  Further, according to the automatic culture apparatus 1 according to the present embodiment, there is no mechanism for taking out the culture vessel 3 in the culture chamber 4. That is, only the tray holding member 4c for supporting the tray 7 in a state where the tray 7 is placed is provided in the culture chamber 4, and all the mechanisms for taking out the culture vessel 3 are arranged outside the culture chamber 4. The transport robot 5 is integrated. The transfer robot 5 can be completely retracted to the outside of the door 4a of the culture chamber 4 after the tray 7 is taken in and out.

Therefore, when the door 4a is closed, there is no mechanism in the culture chamber 4, and no dust is generated as generated by the operation of the mechanism. In addition, the inside of the culture chamber 4 is maintained at a temperature of 37 ± 0.5 ° C., a humidity of 100%, a CO ₂ concentration of 5%, and the like. The problem does not occur. Even when the door 4a is opened, only the tip of the hand 5c of the transfer robot 5 is inserted into the culture chamber 4, so that the rotation mechanism and the sliding mechanism substantially enter the culture chamber 4. There is no. As a result, intrusion of dust into the culture chamber 4 is suppressed, and the cleanliness inside the culture chamber 4 can be increased.
Incidentally, the culture chamber 4, CO ₂ incubator, a multi-gas incubator, incubator or as cool box, etc., may be constituted by one or a combination is used in the culture.

  Furthermore, according to the culture treatment apparatus 30 and the automatic culture apparatus 1 according to the present embodiment, the second space S2 of the culture treatment apparatus 30 is partitioned into the upper space S21 and the lower space S22 by the first partition wall 33. The upper space S21 is provided with an air purifier 32 that generates a clean downdraft. The first partition wall 33 is provided with a vent 38 in the vicinity of the side walls 30a and 30b. The first partition wall 33 is formed with through holes 36, 37 and the like for penetrating various devices in addition to the vent hole 38. By ensuring a sufficiently larger cross-sectional area such as 37, the airflow can be passed through the vent 38.

  Accordingly, the clean airflow descending in the upper space S21 is directed toward the side walls 30a and 30b near the first partition wall 33, and is circulated to the lower space S22 via the vent hole 38. As a result, the airflow that has swept the dust floating in the upper space S21 downward does not stay in the corners near the side walls 30a, 30b of the upper space S21, and smoothly flows to the lower space S22. Will be allowed to.

  Furthermore, according to the culture treatment apparatus 30 and the automatic culture apparatus 1 according to this embodiment, the upper space S21 in which the culture container 3 with the lid 3b opened is moved is necessary for the movement of the culture container 3. Only the mounting table 21, the XY table 17 a of the microscope 17, the supply / discharge robot 10 that needs to be accessed from the upper opening of the culture vessel 3, the electric pipette 12 of the dispensing robot 13, the light source portion of the microscope 17, etc. are arranged. The mechanism part is arranged in the lower space S22. Therefore, the generation of dust in the upper space S21 is minimized, and the possibility of dust being mixed into the culture vessel 3 is reduced.

  In particular, devices that are likely to generate dust, such as the centrifuge 11, the waste container 39, the exhaust fan 41, and the like, are the lowest in the lower space S22 divided by the second partition wall 34. The dust generated in the space S222 does not flow into the upper space S21. Furthermore, the air in the space S222 is sucked by the exhaust fan 41, and after dust is removed by the HEPA filter 42, it is discharged to the outside of the culture treatment apparatus 30. Therefore, the cleanliness of the upper space S21 is maintained at a very high cleanliness.

  In addition, since the second partition wall 34 is also provided with the vent holes 43 along the side walls 30a and 30b, the airflow including dust flowing in from the upper space S21 does not spread into the space S221 smoothly. It will be distributed toward the space S222.

  Furthermore, the waste container 39 containing the used chip 14 to which the medium and cells are attached is detachable, and the cleanliness of the lower space S22 is restored to a high state by replacing it as necessary or periodically. be able to. Further, the duct 40 through which the used chip 14 is passed when being discarded into the disposal container 39 can be removed or replaced or cleaned as necessary or periodically, thereby contributing to improvement in cleanliness.

Furthermore, since the automatic culture apparatus 1 according to the present embodiment includes the air cleaning unit 6 above the central space S12 where the transfer robot 5 is installed, the automatic culture apparatus 1 is always in the central space S12 where the transfer robot 5 exists. Cleanliness is maintained. Therefore, even when the door 4a of the culture chamber 4 is opened, it is possible to minimize the inflow of dust into the culture chamber 4.
Therefore, according to the automatic culture apparatus 1 according to the present embodiment, it is possible to reduce the possibility that cells in culture are contaminated with dust and the like, and it is possible to culture healthy cells.

  In addition, this invention is not limited to the structure shown in the said embodiment. That is, the shape and number of the culture chamber 4, the form and number of the transfer robot 5, the supply / discharge robot 10 and the dispensing robot 13, the form and number of various apparatuses, etc. are not limited at all, and can be arbitrarily selected according to the application conditions. Can be set to

  In addition, the cell suspension was prepared by supplying PBS to the introduced bone marrow fluid and stirring, but instead of this, red blood cells may be removed by introducing a hemolytic agent and performing hemolysis treatment. .

As growth factors, in addition to cytokines, for example, substances that contribute to growth such as concentrated platelets, BMP, EGF, FGF, TGF-β, IGF, PDGF, VEGF, HGF, and combinations thereof are employed. You may decide to do it. In addition to penicillin antibiotics, any antibiotics such as cephem, macrolide, tetracycline, fosfomycin, aminoglycoside, and new quinolone can be employed as the antibiotic.
The automatic culture apparatus according to the present invention is not limited to the culture of bone marrow mesenchymal stem cells. You may culture the cell extract | collected from the various structure | tissue of the biological body, and the established cell line.

  Moreover, as a biological tissue filling material, instead of calcium phosphate, any material may be used as long as it is a material having an affinity for biological tissue, and a bioabsorbable material is more preferable. In particular, porous ceramics having biocompatibility, collagen, polylactic acid, polyglycolic acid, hyaluronic acid, or a combination thereof may be used. Further, a metal such as titanium may be used. The biological tissue filling material may be granular or block-shaped.

It is a perspective view which shows the automatic culture apparatus which concerns on one Embodiment of this invention. It is a longitudinal cross-sectional view which shows roughly the 1st space of the automatic culture apparatus of FIG. It is a top view which shows roughly the 1st space of the automatic culture apparatus of FIG. It is a perspective view which shows an example of the culture container used in the automatic culture apparatus of FIG. It is a front view which shows an example of the introduction container which introduce | transduces a cell into the automatic culture apparatus of FIG. It is a top view which shows the bone marrow fluid introduction part which introduces the introduction container of FIG. It is a front view of the bone marrow fluid introduction part of FIG. It is a figure explaining the supply discharge part of the bone marrow fluid introduction part of FIG. It is a front view which shows the hand movement mechanism of the bone marrow fluid introduction part of FIG. FIG. 6 is a structural diagram showing a lid opening / closing mechanism of the bone marrow fluid introduction part of FIG. 5. It is a figure which shows the nail | claw of the chuck mechanism of the cover body opening / closing mechanism of FIG. It is a front view explaining the raising / lowering mechanism carrying the cover body opening / closing mechanism of FIG. It is a perspective view which shows the apparatus on a 2nd division wall by removing the 1st division wall of the culture processing apparatus of the automatic culture apparatus of FIG. It is a perspective view which shows the apparatus which removed the 1st and 2nd division wall of the culture processing apparatus of the automatic culture apparatus of FIG. 1, and was installed in the lowest space. It is a longitudinal cross-sectional view which shows the attachment structural example of the duct connected to the waste container of the culture processing apparatus of the automatic culture apparatus of FIG.

Explanation of symbols

S2 Second space (processing room)
30 Culture treatment equipment 54 Bone marrow fluid (specimen)
55 Bone Marrow Fluid Introduction Unit (Sample Introduction Device)
56 Introduction container 57 Lid (cap)
58 Identification code 60 Lid opening / closing mechanism (cap opening / closing means)
61 Lifting mechanism (moving mechanism)
64 gripping hand (first gripping means)
76 Chuck mechanism (second gripping means)
82 Sensor 89 Identification code reader

Claims (4)

A sample introduction device that introduces a sample in a processing chamber that performs a predetermined process on a sample to be cultured while being accommodated in an introduction container composed of a substantially cylindrical tube that can be opened and closed by a cap fastened to an upper end opening. ,
First gripping means for receiving an introduction container containing a specimen and gripping a side surface of the tube;
Second gripping means for gripping the cap of the introduction container gripped by the first gripping means;
A cap that relatively displaces the first gripping means and the second gripping means along the longitudinal axis while relatively rotating the first gripping means or the second gripping means around the longitudinal axis of the tube. A sample introduction device comprising an opening / closing means. An identification code is attached to the side of the tube,
An identification code reader is disposed in the processing chamber,
The second gripping means is provided rotatably around the longitudinal axis of the tube;
The sample introduction device according to claim 1, further comprising a moving mechanism for arranging the tube so as to face the identification code reader.   The sample introduction device according to claim 1, wherein the cap opening / closing means includes a sensor that detects a relative displacement along the longitudinal axis.   A culture treatment apparatus comprising the sample introduction device according to any one of claims 1 to 3.

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