JPH01228453A - Automatic partial sampling-and-delivery unit - Google Patents

Abstract

PURPOSE:To provide the title unit so designed that a partial sampling-and- delivery unit is detachably fitted, via a support, on a moving means capable of moving said unit in three-axis direction and a partial delivery nozzle part and a partial sampling-and-delivery pipette part are made to be capable of relative vertical movement, thereby ensuring the partial sampling-and-delivery operation in higher efficiency. CONSTITUTION:The objective partial sampling-and-delivery unit for a variety of solutions on a culture vessel such as a microplate with 96 wells, etc., used for cell culture, etc. This unit 1 is detachably fitted, via a support 2, on a moving means so as to be capable of partial sapling-and-delivery operation on each any well on a microplate. Said unit 1 is also provided with a partial delivery nozzle part X equipped with nozzle(s) 3 for delivering solution to one or more wells and a partial sampling-and-delivery pipette part Y equipped with pipette chip(s) 4 for suction and delivery of solution in one or more wells, and fitted on said support 2 via elevator means 7, 32 for making relative vertical motion for said parts X and Y. Further, said part Y is equipped with means 29, 32 for individually operating plungers 28 by which each pipette chip 4 makes suction and delivery operations.

Description

DETAILED DESCRIPTION OF THE INVENTION Field of Industrial Application The present invention relates to an automatic sorting and dispensing device, and more specifically, it is used in automatic cell culturing devices, cell secretion testing devices, etc.
Dispensing and separating various solutions into the wells of cell culture plates such as 6-well microplates, 24-well microplates, and 6-well microplates is performed.

Conventional technology Conventionally, as this type of automatic preparative dispensing device, for example, Cetus' Provet and Beckman's Biomec 1 have been used.
000 etc. are provided, and in these devices,
As shown in FIG. 8, pipettes P are attached in parallel to the lower end surface of the head H which is moved up and down by an elevating mechanism, and the tip C of this pipette P is placed in the well W of the culture microplate T, and the solution is aliquoted. Note work is being done.

The culture microplate T is, for example, a 96-well microplate, as shown in FIG.
96 (+2×8=96) circular holes (wells W) with a diameter of 7 mm are formed, 12 horizontally and 8 vertically, and the wells W are arranged in parallel at a pitch of 9 mm.

As shown in Figure 1O (1) (I [XII [)], the external dimensions of the 24-well, 12-well, and 6-well microplates are almost the same as the above-mentioned 96-well, and the size of the well W (diameter and depth) ) is changing.

The above-mentioned conventional automatic preparative dispensing apparatus which the present invention aims to solve has the following problems.

■Each pipettor is divided into a preparative pipettor section and a fixed dispensing nozzle section. Therefore, for example, when replacing the culture supernatant during cell culture in a 96-well microplate, the old culture solution must be replaced. After dispensing and discarding with a preparative pipettor, a new culture solution is supplied using a fixed dispensing nozzle.
It was necessary to replace the pipettor head. On the other hand, unlike the above-mentioned method, when a new culture solution is supplied using a preparative pipetter, the preparative pipetter operation is repeated, resulting in a problem of prolonging the working time.

(2) Regarding preparative dispensing pipettes, there are commercially available pipettes that have multiple cylinders and whose pipette depth members can be replaced (installed and removed). However, since the cylinders are raised and lowered, the plungers are driven, and other operations are performed in conjunction with multiple cylinders at the same time, some wells of a culture microplate (for example, wells where bacteria have grown) Even if one does not wish to carry out a preparative dispensing operation, it has been impossible to cancel the dispensing operation only for that well. Furthermore, it was also impossible to attach the pipette tip member to only a portion of the cylinder. Therefore, if it is not desired to dispense into such a part of the well, the dispensing operation must be performed using a single-head pipette, which increases the working time.

■Furthermore, in a preparative dispensing pipettor, when a pipette tip member is attached to the tip of the cylinder, variations in the inner shape of the pipette tip member may cause poor frictional engagement with the outer part of the cylinder tip, making it impossible to attach the pipette depth member or There was a problem in that the pipette tip member easily fell during work. When the pipette tip member falls during work, and the tip falls between the culture microplate to be worked on and the pipetter, there is a possibility that both the pipette function and the microplate will be damaged.

Means for Solving the Problems The automatic sorting and dispensing device according to the present invention solves the problems of the conventional device described in 2.
．． 24.12.A device capable of fractionating and dispensing various solutions to various culture vessels such as 6-well microplates, and moving the fractionating and dispensing device in three axes (XSY, Z) directions. It is attached to a moving means such as a robot arm via a support, and is capable of performing fractional dispensing to each arbitrary well of the microplate, and also for discharging a solution into one or more wells. a dispensing nozzle part, 1
A dispensing pipette section for aspirating and discharging the solution from one or more wells is provided, and the dispensing nozzle section and the dispensing pipette section are provided with an elevating means that allows the dispensing nozzle section and the dispensing pipette section to move up and down relative to each other. an automatic preparative dispensing device, wherein the preparative dispensing pipette section is provided with a means for individually operating a plunger that performs a suction operation on each pipette tip. This is what we provide.

In the above-mentioned apparatus, the dispensing nozzle part includes a single nozzle part, such as three nozzles, and a multi-nozzle part, such as 12 nozzles in one row, and by controlling the supply of solution to the nozzle part, the single nozzle part is capable of dispensing into 1g wells with one nozzle, and the multi-nozzle section allows dispensing into multiple (1g) wells.
It is possible to dispense into multiple wells at the same time using two nozzles.

Further, each of the preparative dispensing pipette parts includes a pipette tip, a cylinder that engages with the pipette tip, a lifting means for moving the cylinder up and down, and a pipette inside the cylinder for aspirating and discharging the solution with the pipette tip. A plunger is slidably inserted therethrough, and a plurality of songle bivet mechanisms equipped with elevating means such as air cylinders are installed in parallel to move the plunger up and down, and the plunger moves the piston rod of the air cylinder. The air cylinders can be raised and lowered individually so that suction and discharge operations can be performed using any pipette tip, and the common cylinder block of these air cylinders is operated by a single lifting mechanism to interlock a plurality of parallel air cylinders. It has a structure that allows it to be raised and lowered.

Further, each nozzle installed in the dispensing nozzle section is inclined with respect to the vertical direction, and a cleaning nozzle is provided above each nozzle for discharging cleaning liquid.

By having the above-described configuration, the present invention enables continuous switching between dispensing work using a dispensing nozzle and dispensing work using a dispensing pipette. Furthermore, by providing a single nozzle part and a multi-nozzle part in the dispensing nozzle part, it is possible to perform dispensing work to one well with one nozzle, and also to perform dispensing work to one well with many nozzles. It is possible to perform dispensing operations on wells at the same time. Similarly, in the preparative dispensing pipette section, it is possible to perform preparative dispensing with one pipette tip or with multiple pipette tips at the same time. Dispensing work can be carried out and work efficiency can be improved.

EXAMPLES Hereinafter, the present invention will be explained in detail with reference to examples shown in the drawings.

The automatic sorting and dispensing device l moves in three axes (X, Y, Z) directions via a bracket (not shown) attached to a support body consisting of an outer frame 2 with an opening at the bottom and a cross-sectional frame shape. is removably attached to a robot arm (not shown) that can
Therefore, a dispensing nozzle 3 and a dispensing pipette tip 4, which will be described later, to be installed in the automatic sorting and dispensing device I are positioned corresponding to an arbitrary well (not shown) of a cell culture microplate, It allows for preparative dispensing work.

As shown in FIG. 1, this automatic preparative dispensing apparatus I has a dispensing nozzle section X disposed on the right side of the figure, and a preparative dispensing pipette section Y disposed on the left side.

In the dispensing nozzle section X, an air cylinder 7 is attached via a bracket 6 to a support plate 5 fixed to the national body 2 of the support. The central part of a horizontal connecting plate 8 is fixed to the lower end of the operating rod 7a protruding from the lower surface of the air cylinder 7, and slide guide blocks 10 are connected to both sides of the connecting plate 8 via slide pars 9. The slide guide block 10 is fixed to both upper surfaces of a block plate 11, and the block plate 11 is raised and lowered by an air cylinder 7. The block plate 11 is provided with a solution supply path and a cleaning liquid supply path, respectively, and a nozzle body 1 to which a dispensing nozzle 3 and a cleaning nozzle 12 are attached.
3 are installed at predetermined intervals. That is, the fourth
As shown in the figure, a single nozzle section 14A with three dispensing nozzles 3 installed in the first row from the outside, and a single nozzle section 14A with three dispensing nozzles 3 installed in the second row.
Multi-nozzle section 14B13 with book dispensing nozzle 3 installed
A multi-nozzle section 14C in which the same <12 dispensing nozzles 3 are installed in each row is arranged. A solution supply line 15 is connected to the solution supply path of each nozzle body 13, and a cleaning liquid supply line 16 is connected to the cleaning liquid supply path. These supply lines 15, 16 for the single nozzle portion 14A are individually connected to lines on the supply source (not shown) side via electromagnetic valves 17 for opening and closing flow paths, and each nozzle 3.
12 are individually supplied with solutions and cleaning fluids. - direction,
The 12 supply lines 15 of the multi-nozzle section 14A are connected to the opening/closing solenoid valve 17 via a common line, and simultaneously supply solution and cleaning liquid to the 12 nozzles 3.12.

The multi-nozzle section 14C is also similar to the multi-nozzle section 14B. Therefore,
In the multi-nozzle section 14B114C, two dispensing nozzles 3 (1') are used to simultaneously dispense to 12 wells.

Therefore, the dispensing operation corresponding to each well of the culture microplate can be performed individually or simultaneously for 12 wells, thereby shortening the working time.

Furthermore, all the dispensing nozzles 3 attached to the block plate 11 are attached so that their tips are at the same height, and these dispensing nozzles 3 are attached to the vertical direction as shown in FIG. It has an inclination of 30° or more (45° in this embodiment). By tilting in this way, the dispensing nozzle 3 can discharge the solution R along the wall surface of the well W of the culture microplate T, as shown in FIG. In this case, in order to prevent the discharged solution R from directly reaching the bottom of the well, the cell colony S on the bottom is dispensed through the wall of the well W.
The force acting on the colony S can be significantly reduced, making it possible to perform dispensing operations without destroying the colony S.

Each of the cleaning nozzles 12 is installed above each dispensing nozzle 3, and the cleaning nozzle 12 discharges a cleaning solution to clean the culture solution remaining on the side surface of the dispensing nozzle 3. .

In the preparative dispensing pipette section Y disposed on the left side in FIG. The ball screw 23 is connected to a vertical ball screw 23 installed in a vertical direction, and the ball screw 23 is rotated in forward and reverse directions by a pulse motor 20. A ball nut 24 is screwed into the ball screw 23, a slide block 25 is fixed to the pole nut 24, and a cylinder block 2 is attached to the slide block 25.
6 is fixed, and the cylinder block 26 is moved up and down by the pulse motor 20.

The cylinder block 26 has 12 cylinder chambers 2 inside.
6a, and a Vislon rod 27 is fitted into each cylinder chamber 26a. Each Vislon rod 27 protruding from the lower surface of the cylinder block 26 connects a plunger 28, and each plunger 28 is slidably fitted into a tip engaging cylinder 29. Additionally, a spring 30 is attached within the cylinder 29 to urge the plunger 28 upward. Each cylinder 2 above
The inner circumferential surface of the upper part of the separating/dispensing pipette tip 4 is fitted onto the outer circumferential surface of the lower end of the pipette tip 9 through frictional engagement.

Connectors 31A13113 connected to the air lines 30A and 30B are attached to the upper and lower sides of each cylinder chamber 26a of the cylinder block 26, and the cylinder chambers
The Vislon rod 27 is moved up and down by supplying and discharging air to the space defined by the Vislon rod 27 of 6a.

The supply and discharge of air to each cylinder chamber 26a can be controlled separately, so that the blunt gears 28 can be individually raised and lowered via the Vislon rod 27, and suction and discharge operations can be performed using any pipette tip 4. ing.

Each of the above-mentioned tip engaging cylinders 29 is individually moved up and down by an air cylinder 32 fixed to the national body 2. That is, similarly to the cylinder 26, the lower end of the piston rod 33 fitted in the 12 cylinder chambers 32a partitioned in the cylinder block is connected to the housing 34,
The housing 34 is connected to the cylinders 29, so that the cylinders 29 are individually moved up and down. At this time, a housing 34 fixed to the cylinder 29 is slidably fitted and guided via a ball bush 37 along a guide bar 36 supported by a support plate 35 fixed to the national body 2.

When attaching the pipette tip 4 to the cylinder 29,
As shown in FIG. 6 (I rod 33
is lowered, a predetermined cylinder 29 is lowered, and the cylinder 29 is fitted into the pipette tip 4 by pushing it into the pipette tip 4 from above. At this time, since the cylinders 29 can be lowered independently, it is possible to deal with variations in the shape of the individual pipette tips 4, and it is possible to increase the mounting accuracy of the pipette tips 4.

Further, in order to prevent the pipette tip 4 from falling during the operation, a holder 41 is provided to hold the body of the pipette tip 4. The holder 41 has an abbreviated shape as shown in the figure, and its upper end is fixed to the cylinder 29 and suspended.
As shown in Figure (II), the lower end pressing portion 41a is pressed against the body of the pipette tip 4. The holder 41 is attracted by an electromagnetic force when energized by a solenoid holder 43 provided at the bottom of a solenoid holder 42 attached to the cylinder 29, as shown in FIG. ing. Further, a tip removal plate 44 is installed above the attachment part of the pipette tip 4, and the bar 44 is attached to the piston rod 46 of the air cylinder 45 which is fixed to the national body 2.
They are connected via 7. Therefore, the air cylinder 4
By lowering the piston rod 46 of No. 5, the plate 44 is lowered and brought into contact with the upper end surface of the pipette tip 4, and the pipette tip 4 is pushed down and removed from the cylinder 29.

The air supply/discharge line of each air cylinder of the preparative/dispensing pipette section Y is as shown in FIGS. 1 and 3, and the air control member 50. An air control member 54 that supplies air to each of the twelve cylinder chambers 32a is fixedly installed on the support body 2. That is, the connectors 51A and 151B attached to the air control member 50 are connected to the connectors 31A and 31B of the cylinder chamber 26a via the lines 30A and 30B, and similarly the connectors 55A and 55B attached to the air control member 54 are connected to the lines 53A and 53B. It is connected to the connector 52A152B of each cylinder chamber 32a via.

In the preparative dispensing pipette section Y configured as described above, the 12 parallel pipette tips 4 are connected to the engaged cylinder 2.
9 are raised and lowered individually by the air cylinder 32,
Any pipette tips 4 can be raised and lowered individually, and all pipette tips 4 (12 tips 4 in one row) can be raised and lowered in one piece. Furthermore, the plunger 28 that performs suction and discharge operations of the pipette tip 4 can be operated up and down by a pulse motor 20, a ball screw 23, a pole nut 24, a cylinder block 26, and a piston rod 27. I can do it. At this time, the driven plunger 28 supplies compressed air from the connector 31A into the corresponding cylinder chamber 26a,
The piston rod 27 is lowered. In this case, the piston rod 27 moves up and down in conjunction with the up and down movement of the cylinder block 26. Therefore, the plunger 28 connected to the piston rod 27 moves up and down to perform the necessary suction and discharge operations. On the other hand, the plunger 28 that you do not want to drive is
Even if the connectors 31A and 31B of the corresponding cylinder chamber 26a are released and the pulse motor 20 is driven to lower the cylinder block 26, the piston rod 27 is not moved to the uppermost position by the action of the spring 30 in the cylinder 29. I am keeping it. In this state, even if the cylinder block 26 moves up and down, the plunger 28 will not move.
The plunger 28 does not perform suction or discharge operations. In this way, in this preparative/dispensing pipette section Y, any pipette tip 4 can be operated individually, and
A plurality of pipette tips 4 or all pipette tips 4 can be operated integrally. Therefore, a plurality of pipette tips 4 can be used to simultaneously carry out sorting and dispensing operations corresponding to each well of the culture microplate.

The above-mentioned preparative dispensing pipette section Y and the dispensing nozzle section X
is moved up and down relative to. That is, by raising and lowering the dispensing nozzle part The tip is set at a higher position than the tip of the pipette tip 4. The tip of the dispensing nozzle 3 can be moved by the air cylinder 7 to a position higher than the tip height of the pipette tip 4 (a position below the tip) by the depth of the well of the culture microplate from the tip of the pipette tip 4. It is set as follows. In this way, the preparative dispensing pipette part Y and the dispensing nozzle part
Therefore, when performing a continuous dispensing operation using the dispensing nozzle 3 after performing preparative dispensing work using the pipette tip 4, the block plate 11 is lowered by the air cylinder 7, and the dispensing By lowering the tip of the injection nozzle 3 below the tip of the pipette tip 4, continuous dispensing work can be performed using the dispensing nozzle 3.

That is, simply by driving the air cylinder 7, it is possible to easily switch between preparative dispensing work and continuous dispensing work, and the time required for switching can be significantly shortened.

In addition, as mentioned above, during the operation of either the dispensing nozzle section It is possible to perform tasks corresponding to each well of the microplate, improving work efficiency.

Furthermore, in this device, for example, as shown in FIG.
A preparative dispensing pipette tip 4 (similarly, a dispensing nozzle 3) provided for the wells of a 96-well microplate,
It can be used for wells W of a 6-well microplate T, and in this case, as shown in the figure, two pipette tips 4 (also nozzles 3) are used for one well W to perform aliquots. Able to do note work. In this way, with this device, the nozzle tip to be used can be freely selected.

Furthermore, in this device, the dispensing nozzle 4 is tilted to dispense the culture solution R onto the wall surface of the well W as shown in FIG.
can prevent collapse.

For example, the tip of the nozzle 3 is positioned at least 1 inch away from the center of each well of a 96 microplate in the liquid ejection direction and 1 mm or more in the vertical direction, and the nozzle inclination angle is 45°.
, the nozzle height was set to 1 mm from the top of the well, and a colony (cells were 5P210
- OK cells) were formed using a nozzle 3 with an inner diameter of 0 and 5 mmφ and a dispensing speed of 75 μρ/sec.
When 50 μNjl was dispensed, no colony collapse was observed.

It should be noted that the present invention is not limited to the above-mentioned embodiments, but is shown in FIG. 7 (I
As shown in FIG.
The plunger 28° may be driven by the plunger drive unit 70. That is, the body 7 of the plunger drive section 70
5 is fixed to a base (not shown), etc., a pulse motor 20° is fixed to the national body 75, and the pulse motor 2
0' operates the air cylinder 26°, and one end of a flexible tube 72 is connected to the piston rod 27′ of the air cylinder 26°.

On the other hand, the pipette movable part 71 moves its outer body 73 to
It is attached to a robot arm 74 that moves in the Y and Z directions. An air cylinder 32° for raising and lowering the cylinder 29' is attached to the body 73, and the pipette tip 4 is engaged with the lower end of the cylinder 29', and the cylinder 29' is moved up and down.
The upper end of the plunger 28', which is slidably fitted within the plunger 9', is connected to the other end of the flexible tube 72.

In FIG. 7, members corresponding to those in the embodiment described above are indicated by the same reference numerals with (゛) attached thereto, and their explanations will be omitted. Furthermore, in reality, the pipette movable section 71 is also provided with the dispensing nozzle section X within its national body 73;
Since the configuration is the same as that of the previous embodiment, illustration and explanation will be omitted.

If the plunger drive unit consisting of a pulse motor, air cylinder, etc. is separated from the movable part moved by a robot arm, etc., the weight of the movable part can be reduced. I can do it.

Effects of the Invention As is clear from the above explanation, the automatic sorting and dispensing device according to the present invention can be used for cell culture in 96-well microplates, 24-well microplates,
When performing preparative dispensing work on wells such as a 6-well microplate, it is extremely easy to switch between work using a partial pressure nozzle and work using a preparative pipette tip. Since it is possible to work with each well and preparative pipette tip individually or with multiple pipette tips at the same time, it is possible to perform work facing each well, which also improves work efficiency. It can be suitably used in automatic cell culturing devices, cell secretion testing devices, etc.

In addition, with this device, only the pipette tip can be attached individually to the cylinder pipette tip.
It is possible to shorten working time. Moreover, since the cylinders can be operated individually in response to variations in the shape of the pipette tip during attachment, the pipette tip can be attached reliably. Moreover, since the pipette tip and the cylinder are pressed and engaged by the holder, it is possible to reliably prevent the tip from falling off during the operation.

On the other hand, the pipette tip can also be automatically removed using an air cylinder, making the pipette tip replacement work more efficient.

Furthermore, in this device, the dispensing nozzle is tilted to discharge the solution along the well wall surface, which provides various operational advantages such as not destroying the colony when there is a cell colony on the bottom of the well. It is something that you have.

[Brief explanation of the drawing]

FIG. 1 is a front view showing an embodiment of the automatic sorting and dispensing device according to the present invention, FIG. 2 is a right side view of FIG. 1, FIG. 3 is a left side view of FIG. 1, and FIG. Figure 1 shows the arrangement of the dispensing nozzle and the dispensing pipette tip as seen from the bottom side, and Figure 5 (I The drawing shown in FIG. 6 ([XIIXI) is a drawing showing the state in which the pipette tip 4 is attached to and removed from the cylinder 29, and the drawing shown in FIG.
1) is a perspective view showing a modification of the present invention, FIG. 7 (II) is a sectional view of FIG. Figure 9 is a perspective view showing a 96-well microplate, Figure 1O (IXIIXII[) is a plan view showing a 24-well microplate, a 12-well microplate, and a 6-well microplate, respectively, and Figure 11 is a device of the present invention. FIG. 2 is a schematic diagram illustrating a usage example of the method, in which two nozzles are used for one well. 1...Automatic preparative dispensing device, 2...Kokutai, 3...Dispensing nozzle, 4...Pipette tip, 7...Air cylinder, 14A...Single nozzle section, 14B, 14C...Multi nozzle section, 17... Solenoid valve, 20... Pulse motor, 24... Slide block, 26... Cylinder block, 27... Piston rod, 28... Plunger, 29... Cylinder, 30
...Spring, 32...Air cylinder, 40...Holder, 43
・・Solenoid, Patent Applicant Sumitomo Electric Industries, Ltd. Agent Patent Attorney: Mr. Maeda and 2 others Figure 4 Figure 5 Figure 6 (1) Figure 8 Figure 10

Claims (1)

[Claims] (1) A device for dispensing various solutions into culture containers such as 96-well microplates used for cell culture, etc., in which the support of this dispensing and dispensing device can be attached to and detached from the moving means. a dispensing nozzle section that is attached to the microplate, and is equipped with a nozzle for discharging a solution into one or more wells, so as to be able to perform preparative dispensing to each arbitrary well of the microplate; Alternatively, a preparative dispensing pipette section equipped with a pipette tip for aspirating and discharging solutions from multiple wells is provided, and the dispensing nozzle section and the preparative dispensing pipette section are moved up and down relative to each other. The pipette is attached to the support via a lifting means that allows the pipette to move, and the pipette section is further provided with means for individually operating a plunger that causes each pipette tip to perform suction and discharge operations. Automatic preparative dispensing device.