JPS5953029B2 - Tissue culture device equipped with centrifugal tube transport mechanism - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to an apparatus for automatically culturing living tissues such as cells.

Generally, tissues are cultured in culture containers such as petri dishes, and a large number of these culture containers are placed in a tissue culture device.

In conventionally known manual tissue culture, it is often necessary to take out the culture vessel from the inside of the culture apparatus into the outside air in order to observe the culture state of the tissue or perform various operations necessary for subculture.

Therefore, the tissue has a specific atmosphere within the culture device,
Due to environmental conditions such as temperature and humidity, mutations may occur when taken out into the outside air, and there is also a risk of contamination by bacteria in the atmosphere.

In order to solve these difficulties, the applicant has developed a tissue culture device that dynamically performs all the necessary culture operations inside the device under a constant atmosphere.

With this device, culture operations are performed as follows.

That is, the tissue to be subcultured is diluted with a culture solution, suspended, and poured into a culture container such as a petri dish, and cultured stationary in a culture device maintained in a specific atmosphere.

Then, after a predetermined period of time has elapsed, the culture container is transported to an observation position within the apparatus, and the culture state of the tissue is inspected using a microscope.

If it is confirmed that the tissue has grown to the full capacity of the culture container, move it to the dispensing position in the device, aspirate the culture solution in the container with a pipette, and then remove the tissue in the container from the buffer solution. Wash by injection and discard the buffer by aspirating with a pipette.

Next, in order to bring the tissue implanted on the bottom of the culture container into a state where it is about to be released from the bottom of the culture container, an enzyme solution such as 1 to lipsin is injected and left for a predetermined period of time.

After this time has elapsed, the enzyme solution is sucked out from the culture container with a pipette, then the culture solution is again injected into the culture container, and the culture solution is continuously sucked and drained with a pipette to vibrate and agitate the tissue. completely released from the bottom of the culture vessel and suspended in the culture medium.

Furthermore, this floating tissue is transferred to a centrifuge tube with a pipette, and the culture solution and tissue are separated using a centrifuge.

As a result, tissues with high specific gravity adhere to the bottom of the centrifuge tube,
The culture solution becomes a supernatant, which is discarded by tilting the centrifuge tube.

Then, the culture solution is poured into the centrifuge tube again, and the culture solution is stirred by the pipette's sucking and discharging operation similar to that described above to uniformly suspend the tissue in the culture solution within the centrifuge tube. Dispense it into equal parts to complete one culture.

Then, subculture of the tissue is performed by performing such culture operation multiple times for each of the tissues contained in a large number of culture containers.

The reason why centrifugation is performed during the above-mentioned culture operation is as follows.

In other words, the enzyme solution that is injected to bring the tissue implanted on the bottom of the culture container to a state immediately before being released cannot be completely discarded by suction with a pipette, and a small amount of the enzyme solution is deposited on the bottom of the culture container. remain attached or between tissues.

In this state, the tissue cannot be dispensed into two new culture vessels while suspended in the culture solution that is subsequently injected.

Therefore, by centrifuging the tissue suspended in the culture solution, the remaining enzyme solution is also separated from the tissue.

Note that at the same time, some tissues that died during culture are also separated from the tissues to be dispensed.

In a tissue culture apparatus with such a configuration, a centrifuge tube is mounted on a rotary table, and each rotation of the rotary table is It is conceivable to perform various operations depending on the position.

However, in one rotational position the mounted centrifuge tube must be conveyed to the rotor of the centrifuge, which forms a rotational system separate from the carousel, and vice versa.

The rotor must be stopped during this transport, but
The stoppage time of the rotor must be shortened as much as possible.

The reason for this is that the total number of tissues to be processed during subculture in the culture device doubles with each generation, so the centrifuge must be operated almost continuously at the end of subculture. This is because the need arises.

Therefore, the number of generations that can be subcultured is determined by the time period during which the rotor must be stopped.

An object of the present invention is to provide a tissue culture device that allows centrifuge tubes to be efficiently transported between a rotary table and a rotor of a centrifuge.

In order to achieve this object, the present invention provides a rotary table on which a plurality of centrifuge tubes can be mounted at equal intervals on the same circumference, a central opening is formed in the rotary table, and the centrifugal tubes are inserted into the central opening. A holding member is provided that detachably holds the centrifuge tubes by displacing them in the radial direction; The centrifuge tube is disposed coaxially with the rotor and configured to detachably hold the centrifuge tube by vertically displacing the centrifuge tube below the central opening of the rotary table; a claw engageable with the centrifuge tube. and a first drive part that displaces this pawl inward in the radial direction of the rotary table from the centrifuge tube attachment/detachment position on the rotary table to a position directly above the centrifugal tube attachment/detachment position on the rotor; The present invention is characterized in that it is provided with a centrifugal tube conveying mechanism having a second drive portion that vertically displaces the tube to the tube attachment/detachment position.

BRIEF DESCRIPTION OF THE DRAWINGS In order to make other objects, features and advantages of the invention clear, the invention will be described with reference to the illustrated preferred embodiments.

FIG. 1 is a plan view showing the centrifugal tube rotating operation table of the tissue culture apparatus according to the present invention in correspondence with various operation mechanisms;
The figure is a partial cross-sectional view of the rotary operating table taken along the line II--II of FIG. 1, and showing the attached mechanisms in relation to each other.

In the figure, 1 is a centrifuge tube rotating operation table, 2 is a centrifuge that works with the rotary operation table 1 to centrifuge the cultured tissue in the centrifuge tube, and 3 is a tube for inserting or removing the centrifuge tube into the centrifuge 2. This is a mechanism that opens the lid of the centrifuge 2 and positions the rotor of the centrifuge 2.

In the present invention, the rotary operating table 1 is disposed coaxially above the centrifuge 2.

In this rotary operating table 1, a circular rotary plate 5 having an opening 4 in the center is rotatably arranged on a plate 6, and two fixed rollers 7 which abut the outer surface of the rotary plate 5 and two fixed rollers 7 in the radial direction Centering is performed using two moving rollers 8 that are movable to
and is connected to a drive motor 12 outside the culture atmosphere via a plurality of pairs of spur gears 11.

The holder 1 is installed at a plurality of positions on the rotating plate 5, in the illustrated example, at eight positions spaced apart from each other by equal distances in the circumferential direction on the same radius.
A holding mechanism 15 is provided to tiltably hold the centrifugal tube 14 inserted into the centrifuge tube 3.

Although only four retaining mechanisms 15 are shown in FIG. 1 for simplicity, there is actually one additional retaining mechanism 15 between each of the illustrated retaining mechanisms.

In each of these holding mechanisms 15, a holding member 15a that engages with the flange portion 13a of the cylindrical holder 13 from both upper and lower sides is fixed to a pivot shaft 15b, and this pivot shaft 15b is supported by a bearing member 15C mounted on the rotary plate 5. , furthermore, the axis 15b
A disk 15e having a pin 15d implanted therein is attached to one end of the disk.

In the centrifuge tube disposal position, the pin 15d functions to rotate the holding member 15a about the pivot 15b in cooperation with the lifting rod 16 shown in phantom in FIG.

On the other hand, when the holding member 15a returns, the shaft 15b and the bearing member 15
This is done by a spring member (not shown) inserted between C and C.

The same applies to the disposal of the supernatant.

A cam follower 17 is fixed to the holding member 15a in order to tilt the holder 13 and the centrifugal tube 14, and the free end of the cam follower 17 is brought into contact with an annular cam 18 fixed to the plate 6 below the rotating plate 5. .

This annular cam 18 has a cam shape that allows the centrifuge tube 14 to stand upright only in its lifting and lowering positions and in its feeding and dispensing positions.

It is preferable to use a curved plate having a substantially L-shaped cross section as the cam follower 17, and it is preferable that the engagement between the cam follower 17 and the annular cam 18 is ensured by the aforementioned spring member. .

A microswitch 19 that senses the origin of the rotary plate 5, that is, the feeding position and generates a signal, and a microswitch 20 that senses the index position and generates a signal are arranged in the rotary plate drive section. The output signal of the switch is transmitted to, for example, a computer (not shown).

Note that the microswitches 19 and 20 may be directly connected to the drive motor of each operating mechanism so that necessary operations can be performed for each side-exit position.

In the illustrated state of the rotating plate 5, position A refers to the centrifugal tube supply mechanism 21, position B refers to the supernatant liquid disposal mechanism 22, and position C refers to the supernatant liquid disposal mechanism 22.
refers to the culture medium supply mechanism 23, position refers to the supply and dispensing mechanism (not shown), and position E refers to the tube disposal mechanism 24.
correspond to each other.

Note that the tube transport mechanism 25 that corresponds to position F will be described in detail later.

In this example, both the rotary plate 5 and the opening 4 are circular, but they can also be formed into any angular shape.

Further, three or more positioning rolls 7.8 may be provided as desired.

The centrifuge 2 disposed below the rotary operation table 1 has a drive motor 27 fixed to a frame 26 outside the culture atmosphere via a casing 26a, and an output shaft 27 of the drive motor 27.
A rotor 28 connected to H and capable of simultaneously centrifuging four centrifugal tubes 14, and a fixed casing 2 of this rotor 28.
9, and a lid member 30 that opens and closes the upper end opening of the casing 29.
and a housing 31 fixed to the frame 26 with the rotor 28 and casing 29 housed therein.

The rotor 28 connects a rotating element 28b to the end of the output shaft 27a via a rotating arm 28a, fixes a centrifugal tube holding member 28C to the rotating element 28b, and fixes the centrifugal tube holding member 28C to the rotating arm 28H.
One depression 2 spaced apart from the axis of the shaft 27H on the upper end surface of the
8d.

The centrifugal tube holding member 28C holds the centrifugal tube 14 in a vertical position as shown by the imaginary line in the figure when the rotating element 28b is stopped, and holds the centrifugal tube 14 in a horizontal position by centrifugal force when rotating.

Further, the recess 28d cooperates with the mechanism section 3, which will be described in detail, to determine the absolute position of the rotating element 28b when the centrifugal tube 14 is transported.

Note that 32 is a pipe for discharging the culture solution and water droplets spilled inside the casing 29 to the outside of the centrifuge 2;
33 is a sealing member that eliminates contact between the culture atmosphere with 100% humidity and the drive motor 27 to increase the durability of the motor.

The mechanism section 3, which positions the rotor 28 of the centrifuge 2 and also opens and closes the lid member 30 of the centrifuge, is a rod to which step motors 34 and 35 are connected via a cam mechanism 36 and a speed reduction mechanism 37, respectively. 38, a pin 40 projecting downward through a disk 39 attached to the lower end of the rod, and a lid member 30 provided at the middle of the rod 38.
It consists of an opening/closing mechanism 41.

The cam mechanism 36 connects the rotating cam 3 to the output shaft 34a of the motor 34.
6a, and also a cam follower 36 at the upper end of the rod 38.
b is fixed, and the rod 38 is configured by fixing the cam 3.
6a, it descends by a certain amount, and after the end of this downward stroke, it is raised by a restoring spring (not shown) in the rod guide member 43 fixed to the plate 42.

The operating state of the motor 34 is input to the computer by a microswitch 44 for sensing the origin of the cam 36a and a microswitch 45 for sensing the index position.

These microswitches 44, 45 may be directly connected to other operating mechanisms, as in the case of microswitches 19 and 20 described above.

The speed reduction mechanism 37 includes a spur gear 37a fixed to the output shaft 35a of the motor 35 and a spur gear 37 fixed to the upper end of the rod 38.
b to rotate the rod 38 about its axis.

In order to sense this rotational position of the rod 38, a microswitch 47 for sensing the origin and a microswitch 48 for sensing the index position are provided on the support plate 46 in association with the spur gear 37b, and these are operated in the same manner as described above. Connect to other circuits.

A pin 40 protruding downward from the disk 39 is pressed downward by a spring member 40a at a position where it can engage with the peripheral edge of the disk 39, that is, the aforementioned recess 28d, and the tip of this pin 40 is When not engaged with the recess 28d,
It comes into contact with the upper end surface of the rotating arm 28a and moves on that surface.

Therefore, when the pin 40 is not engaged with the recess 28d, it is maintained at a somewhat elevated position against the force of the spring 40a, and only when it is engaged with the recess 28d, it projects downward to the lower limit position.

The opening/closing mechanism 41 of the lid member 30 includes a guide member 41b that is fixed to a plate 49 and has a guide groove 41a extending diagonally upward and to the left in the illustrated position, and a cam member 41C fixed to the lower end of the guide member 41b. , a pin 41d rotatable in the same plane relative to the rod 38, a cam follower 41f provided with a long hole 416 that cooperates with the pin and the guide groove 41a, and a cam follower 41f that presses the cam follower downward. It is constituted by a spring member 41g.

The cam follower 41f is connected to lid members 41h and 30 that close an opening (not shown) provided in the plate 49 and an opening (not shown) provided in the frustoconical rotor cover 50.

This causes rod 38 to be moved by actuation of motor 34.
When the pin 41d descends, the pin 41d moves into the fixed guide groove 41a.
, move diagonally to the lower right position in the figure, and press pin 4 again.
The cam follower 41f that engages with the pin 41d rotates in the direction of movement of the pin 41d, thereby opening each lid member 30, 41h.

When the lid members 30 and 41h are closed, the convex portion 41j of the cam follower 41f engages with the recess 41 of the cam member 41C, so that the lid members 30 and 41h are in contact with the surfaces of the rotary cover 50 and the plate 49, respectively. However, when they are released, the engagement between the convex portion 41j and the concave portion 41 is released and the convex portion 41j rides on the flat portion 411 of the cam member 41C. It is opened while being separated from the surface of the plate 49.

Therefore, when the lid members 30, 41h are opened and closed,
Dust on the plate 49 and cover 50 does not fall into the centrifuge 2.

On the other hand, the lid members 30, 41h are closed by raising the rod 38 by the rotation of the motor 34 and the extension of the spring member 41g, which is the reverse operation described above.

Further, when the rod 38 is lowered, a pin 40 is also lowered, and this pin 40 positions the rotor 28 in order to take out a centrifuge tube placed on the rotor 28 or to attach a centrifuge tube to the rotor. Since the pin 40 and the cover 28d do not necessarily engage with each other simply by lowering the rod, the pin 28d is rotated 360 degrees around the axis of the rod 38 by the operation of the motor 35, thereby completely engaging the two, and the rotor 28 Determine the absolute position of

In the figure, reference numerals 51 and 52 denote frames that are connected to plates 6, 42, 49, support plate 46, etc., and reliably maintain each component in a predetermined position.

In the present invention, the centrifugal tube engaging portion 53 of the transport mechanism 25
As shown in FIG.
is fixed to the lower end of a vertical lifting shaft 54.

As shown in FIG. 3, the lifting shaft 54 passes through the sleeve 55.

A bearing 56 of the shaft 54 is fixed to the lower end of the sleeve 55.

A groove 57 extending in the axial direction is formed in the sleeve 55, and by engaging a pin 58 fixed to the shaft 54 in the groove 57, the shaft 54 and the sleeve 55 can be relatively displaced in the axial direction. It is structured so that it rotates integrally in the state.

A disk 59 is integrally fixed to the outer periphery of the sleeve 55, and a protrusion 59a fixed to the disk 59 is constantly brought into contact with a roller 61a fixed to a small diameter disk 61 by a spring 60 (see FIG. 5). The latter disc 61 is fixed to the output shaft of a reversible drive motor 62 for rotation.

The motor 62 is connected to the frame 51.52 (see Figure 2).
A mounting base 63 is fixed to the mounting base 63, a support pedestal 64 is fixed to the mounting base 63, and the device is mounted on the support pedestal 64.

Note that bearings 65 and 66 of the sleeve 56 are provided on the mounting base 63 and the support base 64, respectively, and one of the bearings 66
By engaging the boss of the disc 59 with the sleeve 55
position in the axial direction.

A sleeve-like rack member 67 is attached to the shaft 54 within the sleeve 55.
are connected so as to be integrally displaced in the axial direction in a relatively rotatable state.

The rack member 67 meshes with a pinion 68 (see FIG. 4) above the sleeve 55, and the pinion 68 is fixed to the output shaft of a reversible motor 69 for driving up and down.

This motor 69 is connected to another pedestal 7 fixed to the support pedestal 64.
Attach to 0.

As shown in FIG. 5, the rotation angle of the shaft 54 is limited to a constant angle between the solid line position and the chain line position.

In order to reliably prevent further rotation of the disk 59, the support base 64 is provided with a pair of stoppers 7 that engage with the protrusion 59a.
1.72 can be placed.

Furthermore, three microswitches 73 to 73 are mounted on the support base 64.
75, and these microswitches 73 to 75 can be configured to be opened and closed by a switch operating member 76 fixed to the disc 59 at the stop position, first and second rotational positions of the disc 59, respectively.

The conveyance mechanism 25 shown in FIGS. 3 to 5 rotates and raises and lowers the claw 53C and therefore the centrifugal tube 14 that engages with the claw in the following manner.

First, in order to rotate the centrifugal tube 14 by the transport mechanism 25, the rotation drive motor 62 is activated.

The rotation of the motor 62 is transmitted to the disc 61 and the roller 61a.

The roller 61a slides on the side surface of the protrusion 59a of the disc 59 and rotates the protrusion 59a counterclockwise in FIG. 5 from the solid line position to the chain line position.

When the motor 62 is rotated in the opposite direction from this chain line position,
Since the protrusion 59a is pressed against the roller 61a by the spring 60, it follows the roller 61a from the chain line position to the solid line position and returns clockwise due to the spring force of the spring 60.

The rotation of the protrusion 59a within this limited range is due to the rotation of the disc 61.
, is transmitted to the centrifugal tube engaging portion 53 via the sleeve 55, the groove 57, the pin 58 and the shaft 54 as rotation within a certain range.

Note that, during this rotation, relative rotation occurs between the shaft 54 and the sleeve-like rack member 67.

Next, in order to move the centrifugal tube 14 up and down by the transport mechanism 25, the lifting drive motor 69 is activated.

The rotation of the motor 69 is transmitted to the pinion 68, and is transmitted to the rack member 67 that meshes with the pinion 68 as its vertical movement.

Therefore, since the rack member 67 and the shaft 54 are integrally displaced in the axial direction, the centrifugal tube engaging portion 53 moves up and down.

In order to transport the centrifuge tube 14 from the rotary operating table 1 to the centrifuge 2, the transport mechanism 25 is caused to move up and down and rotate in the following order.

Here, the claw 53C of the engaging portion 53 is connected to the centrifugal tube holder 13.
The explanation will be given starting from the initial position, which is slightly below the upper end portion 13b.

As shown in Fig. 6, the claw 53C is moved from the initial position 53-1 (solid line position in Fig. 5), and the claw 53C narrows the holder 13 to a position 53-2 (dotted line position in Fig. , see Figure 6a).

From this position 53-2, the claw 53C is raised to a position 53-3 where the claw 53C pushes up the upper end 13b of the holder 13 and the centrifugal tube 14 is slightly lifted from the rotary plate 5 of the rotary operating table 1 (the third (See Figure 6b).

From position 53-3, the claw 53C is connected to the centrifuge tube 14, and the centrifuge tube 14 is connected to the centrifuge 2.
A position 53 reaching directly above the holding member 28C of the rotor 28
Rotate in the opposite direction to the initial rotation until -4.

From position 53-4, the flange located below the claw 53C at the upper part of the holder 13 engages the holding member 28C, and the centrifugal tube 14 is slightly lower than the position where it is held within the rotor 28. 53-5 (see Figure 6C).

From the position 53-5, the claw 53C is slightly rotated in the same direction as the rotation from the position 53-3 to the position 53-4, and reaches the position 53-6 directly below the initial position 53-1.

At this position 53-6, the claw 53C and the centrifugal tube 14 no longer engage with each other.

Finally, the claw 53C is raised and returned from the position 53-6 to the initial position 53-1.

Then, the rotary operation table 1 and the rotor 28 of the centrifuge 2 are
The transport mechanism 25 is rotated by a predetermined angle so that the same operation can be performed at the new centrifuge tube receiving position and delivery position, respectively.

Moreover, when returning the centrifuge tube 14 from the centrifuge 2 to the rotary operating table 1, the above-mentioned operations may be performed in the reverse order.

Note that the vertical direction and rotational direction of the transport mechanism 25 are switched by switching the motors 69 and 62 between forward and reverse rotations, respectively.

Stopping the rotation at positions 53-2, 53-4, and 53-6 is controlled by a logic circuit including microswitches 73 to 75, such as the circuit shown in FIG.

In addition, the switching and stopping of lifting at positions 53-1, 53-3, and 53-5 are controlled by a similar logic circuit,
A microswitch for this purpose can be arranged in association with the lifting drive motor 69.

The input/output levels of each logic element in the logic circuit of FIG. 7 are shown in the illustrated static state, and those in a state where a drive command is given are shown in parentheses.

In order to mechanically limit the rotation angle of the claw 53C by the motor 62, a guide groove is formed in the projection 59a of the disc 59 as shown in FIG. 8 instead of the example shown in FIG. It is also possible to arrange the roller 61a of the disc 61 in the guide groove.

Further, as shown in FIG. 9, a pair of protrusions 59 are provided on the disc 59.
The rollers 61a of the disc 61 may be guided between the protrusions 59a by disposing the protrusions 59a slightly opposite each other.

In either case, the disc 59 is rotated by the roller 61a in both rotational directions.
The spring 60 required to return the disk 59 clockwise in the example of FIG. 5 is no longer necessary.

The tissue culture apparatus according to the present invention carries out various dispensing operations for culture as described below while transporting the centrifuge tube 14 between the rotary operating table 1 and the centrifuge 2 using the transport mechanism 25 configured as described above. .

In order to facilitate understanding, the explanation will be given from a state in which the centrifuge 2 has previously completed centrifugation of tissues injected into the four centrifugation tubes 14.

Based on the fact that the centrifuge 2 has stopped, the step motor 34 of the mechanism section 3 is activated and the rod 38 is lowered.

As a result, the lid members 30 and 41h are opened to allow the passage of the claw 53C and the shaft 54 of the transport mechanism 25, and the tip of the pin 40 comes into contact with the end surface of the rotary arm 28a.

Next, the pin 40 is engaged with the recess 28d by the operation of the step motor 35, and the absolute position of the rotor 28 is determined.

After the positioning of the rotor 28 is completed, the holding mechanism 15 of the rotary operating table, which is waiting at the position shown in the figure without holding any centrifugal tube 14, receives a predetermined centrifugal tube 14 after centrifugation from the transport mechanism 25. At the same time, at position A, the empty centrifuge tube 14 is removed from the centrifuge tube supply mechanism 21.

Further, the centrifuge tube supply mechanism 21 functions to drop the centrifuge tube 14 into the holder 13 in cooperation with a centrifuge tube stock section (not shown).

After both operations are completed, the drive motor 12, which is a step motor, rotates and the operating table 1 is rotated 178 times.

Therefore, positions A and F on the rotary plate face the supernatant liquid disposal mechanism 22 and tube supply mechanism 21, respectively, but the centrifuge tube at position A has not yet been filled with culture medium, and
Since the centrifuge tube 14 from the centrifuge 2 is held at position F, each mechanism does not perform any work at this rotational position of the console 1.

In addition, position G, which does not face any mechanism in the illustrated state, faces the transport mechanism 25, but the transport mechanism 25 also does not operate at this rotational position.

Position A by subsequent <178 rotations of the console.

F faces the culture solution supply mechanism 23 and disposal mechanism 22, respectively, and positions G and E respectively face the tube supply mechanism 21.
and facing the transport mechanism 25.

At this time, at positions G and E, centrifuge tubes are supplied and centrifuge tubes are delivered in the same way as described above, and at position F, the supernatant after centrifugation is discarded, but at position A, the supernatant liquid is discarded. Since the centrifuge tube 14 is still empty, no culture solution is injected.

Note that when transferring the second centrifuge tube in the centrifuge 2 to position E at this rotational position of the operation console, the 2/2/
Before the completion of 8 rotations, for example after completion of 178 rotations, it is necessary to transmit a signal from the microswitch 20 to the step motor 35 to rotate the rotor 28 by 174 rotations, and to time the operation of the transport mechanism 25 in advance.

The same applies to subsequent deliveries of centrifuge tubes.
By associating the rotor 28 with each other so that it completes 174 revolutions every 278 revolutions of the operating console 1, the specificity of the centrifuge tube that is transferred from the operating console 1 to the centrifuge 2 and returned to the operating console 1 after centrifugation is improved. can be maintained.

Further, the supernatant liquid disposal mechanism descends from the upper position in the same way as the lifting rod 16 of the tube disposal mechanism 24, and presses the pin 15d of the holding mechanism 15 to rotate the centrifuge tube around the pivot 15b. Centrifuge tube rotation mechanism (not shown) and centrifuge tube 1
In order to prevent the centrifuge tube from falling off and contaminating the tissue, the centrifuge tube is provided with a cam mechanism 22a that moves in an arc while engaging with the side surface of the centrifuge tube as the centrifuge tube rotates, and a waste liquid tube 22b.

The centrifuged cultured tissue adheres to the bottom of the centrifuge tube 14, while dead tissue, culture fluid, etc. are located above the attached tissue as a supernatant, so the centrifuge tube 14 is rotated by the above-mentioned mechanism. Then, the supernatant liquid etc. flows out into the waste liquid pipe 22b, and the tissue to be dispensed remains in the centrifuge pipe.

Furthermore, at the 378-rotation position of the operating table 1, position A faces the culture medium supply and dispensing mechanism, and the empty centrifuge tube 14 is in an upright state.

The cultured tissue that has been stirred and aspirated from the Petri dish into the pipette is injected into this centrifuge tube by the operation of a distributor (not shown).

Further, position F is opposite to the culture solution supply mechanism 23 and is located in the centrifugal tube 1.
A new culture solution is injected into the chamber 4 via a culture solution switching device (not shown).

Note that during such rotation of the rotary operating table 1, the centrifugal tube 14 stands upright only in the rotational position facing the supply and dispensing mechanism and the rotational position facing the transport mechanism 25.
In other rotational positions, the annular cam 18 and the cam follower 17 cooperate to keep the culture tissue tilted, so that there is very little risk that the cultured tissue will be contaminated by fallen bacteria, dust, etc.

After 178 rotations, the operating table moves to the 172 rotation position, and position F faces the supply and dispensing mechanism, and the cultured tissue injected with new culture solution is stirred and dispensed by the newly installed pipette. .

This stirring with a pipette is performed by alternately sucking and discharging a fixed amount of the culture solution in the centrifuge tube by means of a pump provided in the dispensing mechanism. As a result, the tissue attached to the bottom of the centrifuge tube 14 is uniformly dispersed in the culture solution. Floating on.

Also, for dispensing, aspirate the tissue in the centrifuge tube into a pipette,
This is done by equally injecting the solution into two empty petri dishes on a dispensing operation table (not shown).

Further, position E faces the disposal mechanism 22, and positions H and D face the tube supply mechanism 21 and the lifting mechanism 25, respectively, and undergo the same operations as described above.

Subsequent rotation causes position G to face the cultured tissue supply mechanism, and the tissue in the petri dish is transferred to the centrifuge tube, and culture solution is injected into the centrifuge tube at position E by the culture solution supply mechanism 23.

With a subsequent 1/8 rotation of the operating table 1, the centrifuge tube 14 at position F is discarded by the tube disposal mechanism 24, and the cultured tissue at position E is dispensed into two empty petri dishes and held in position. The supernatant liquid in the centrifuge tube 14 is discarded by the mechanism 22.

Further, an empty centrifugal tube 14 is supplied to each of positions B and C, and the centrifugal tube force "5+" of the fourth centrifugal tube is delivered from the centrifuge 2.

The centrifuge 2, which has handed over all the centrifugation tubes after centrifugation to the operating table 1, remains at that position without indexing until it receives and removes the cultured tissue before centrifugation after the next 178 revolutions of the operating table.

The disposal mechanism 24 includes a lifting rod 16 operated by a drive source (not shown), a lid opening/closing rod 16a branched from the rod 16, and a lid 2 provided at the waste pipe port 24a.
4b, and a link 24C connected to the lid 24b,
This drive source is actuated based on a signal from the microswitch 20 or the computer generated by position F reaching this mechanism 24.

Therefore, the rods 16 and 16a are lowered together,
The centrifuge tube 14 is rotated by pressing the pin 15d and the link 24C, respectively, and the lid 24b is opened in the direction of the arrow in the figure.

Due to this rotation of the centrifuge tube 14, only the centrifuge tube is attached to the holder 13.
It is pulled out and falls inside the waste pipe.

After the tube is discarded, the centrifuge tube holder 13 and lid 24b return to their original positions by raising the rods 16, 16a.

The reason why the centrifuge tube 14 is discarded every time it is used and then renewed by the tube supply mechanism 21 is that a new sterilized centrifuge tube is always used because the cultured tissue is extremely easily contaminated with bacteria. This is to eliminate as much as possible the adhesion and propagation of germs and to enable pure tissue culture.

Through the subsequent 178 revolutions, the centrifuge tube 14 at position A is transferred to the rotor 28 in the centrifuge 2 as described above, the empty centrifuge tube at position H is injected with cultured tissue from the petri dish, and the centrifuge tube at position is injected with culture medium.

Mounting of the centrifuge tube 14 into the centrifuge 2 is performed by the transport mechanism 25 performing a reverse operation of transferring the centrifuge tube from the centrifuge 2 to the holding mechanism 15 .

At the subsequent one rotation position of the console 1, the centrifuge tube at position E is discarded, the tissue after centrifugation is dispensed from the centrifuge tube at position, and the supernatant liquid is discarded at position B.

After that, by 178 rotations of the operating table, the centrifuge tube at position G is attached to the rotor 28, which has been indexed in the same manner as described above, in the centrifuge 2, and the cultured tissue from the petri dish is injected into the centrifuge tube at position C. The culture solution is then injected into the centrifuge tube at position B.

Subsequently, at <178 revolutions, the centrifuge tube at position is discarded, and the cultured tissue at position B is dispensed into two empty petri dishes.

By this dispensing, the tissue, which was cultured in four petri dishes before centrifugation, is divided into a total of eight petri dishes and cultured stationary again.

At the 1178th rotation position of the operating table, the third centrifuge tube at position H is installed into the centrifuge 2, and at the 1278th rotation position, the centrifuge tube at position B is discarded, and at 1378 rotation, the third centrifuge tube at position C is discarded. The centrifuge tube is attached to the centrifuge 2 and the rotating operation table 1
The operation on the centrifuge tube held above is completed.

At this point, the rod 28 of the mechanism 3 detects the operating state of the transport mechanism 25 or rises based on a signal from a computer or the like to disengage the pin 40 and the recess 28d, and the lid member 30, Close 41h.

Thereafter, the rotary operating table 1 is rotated until the microswitch 19 for detecting the origin is activated and then stopped, and the centrifuge 2 is activated again based on a signal from the microswitch 19 or the computer.

Regarding the cultured tissue newly handed over to centrifuge 2, approximately 10
After the centrifugation for 1 minute is completed, the centrifugation tube in the centrifuge 2 is replaced with a centrifugation tube into which the cultured tissue has been newly injected, again in the same manner as described above.

As is clear from the detailed description above, according to the present invention, the rotary operation table and the centrifuge are arranged in a vertical relationship, and the centrifuge tube can be efficiently transported between them, thereby reducing the wasted time of the centrifuge. This makes it possible to reduce the amount to

[Brief explanation of drawings]

FIG. 1 is a partially sectional plan view showing the essential parts of the tissue culture apparatus according to the present invention, FIG. 2 is a sectional view taken along line II-II in FIG. 1, and FIG. 3 is an example of a centrifugal tube transport mechanism. 4 is a plan view thereof, FIG. 5 is a sectional view taken along line v-V in FIG. 3, and FIGS. 7 is a diagram showing an example of the control circuit, and FIGS. 8 and 9 are schematic diagrams showing modifications of the rotation angle limiting mechanism shown in FIG. 5. be. 1...Rotary operation table, 2...Centrifuge, 4
...Opening, 5...Rotating plate, 14...
...Centrifugal tube, 15...Same holding mechanism, 25...
...Conveyance mechanism, 28...Rotor, 53C.
...Centrifugal tube engaging claw, 62, 69...Motor.

Claims (1)

[Claims] 1. As a tissue culture device, a tissue to be cultured is placed in a culture container that is placed stationary inside the device filled with a predetermined atmosphere, and when the tissue is sufficiently cultured, it is injected into the culture container. The tissue is suspended in a liquid that is aspirated from the culture vessel and injected into a centrifuge tube, and the injected liquid is dispensed into two new culture vessels in the centrifuge tube.The tissue to be discarded and the tissue to be discarded A rotary table is provided on which a plurality of centrifuge tubes can be mounted at equal intervals on the same circumference, and the rotary table has a central opening and the centrifuge tubes are separated in the radial direction within the central opening. A holding member is provided to detachably hold the centrifuge tubes by displacement; a centrifugal separator having a rotor capable of mounting a plurality of centrifuge tubes at equal intervals on the same circumference is disposed coaxially below the rotary table. The rotor is configured to detachably hold the centrifuge tube by vertically displacing the centrifuge tube below the central opening of the rotary table; a claw that can engage the centrifuge tube; a first drive part that displaces the centrifuge tube inward in the radial direction of the rotary table from a centrifuge tube attachment/detachment position on the rotary table to a position directly above the centrifuge tube attachment/detachment position on the rotor; 1. A tissue culture device comprising: a centrifugal tube transport mechanism having a second driving portion for vertically displacing the tissue.