JPH04304881A - Equipment for injection - Google Patents

Abstract

PURPOSE:To provide the subject equipment for injection, capable of stably keeping the internal pressure of a pipette when an injection is carried out. CONSTITUTION:One end of an injection gas-feed passage 32 equipped with the first pressure regulator 34 and an electromagnetic valve (A) is connected to a micropipette. To a passage between the electromagnetic valve (A) and the micropipette, a pressure-regulated gas feed-passage 33 equipped with the second pressure regulator 37 and an electromagnetic valve (B) is connected. In this equipment, the electromagnetic valve (A) is opened after the electromagnetic valve (B) is perfectly closed by a controller unit.

Description

[Detailed description of the invention]

0001

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to an injection device, and more particularly to an injection device for injecting a substance to be treated in a pipette into a substance to be treated.

0002

2. Description of the Related Art An injection device is used when, for example, a micromanipulator injects a treatment substance such as a DNA solution into cells to be treated from a pipette. The injection device includes an injection pressure supply means having a first on-off valve and connected to the pipette, and a pressure regulating means having a second on-off valve and connected to the pipette side of the first on-off valve of the injection pressure supply means. and a valve opening/closing means for opening and closing the first opening/closing valve and the second opening/closing valve. Here, the injection pressure supply means is a means for increasing the internal pressure of the pipette and injecting the substance to be treated within the pipette into the substance to be treated such as cultured cells. Further, the pressure regulating means is a means for returning the internal pressure of the pipette to, for example, atmospheric pressure when the injection operation is not performed.

[0003] When the injection device injects the substance to be treated into the substance to be treated, the valve opening/closing means is the second valve opening/closing means.
After outputting a close command to the on-off valve, an open command is output to the first on-off valve. Then, the injection pressure supply means communicates with the pipette, increasing the internal pressure of the pipette, so that the substance to be treated within the pipette is injected into the substance to be treated. Here, if the opening time of the first on-off valve and the internal pressure of the pipette are set constant,
The same amount of treatment material is injected into the material to be treated during each injection operation. After the injection operation is completed, the valve opening/closing means outputs a closing command to the first opening/closing valve and then outputs an opening command to the second opening/closing valve. This returns the internal pressure of the pipette to atmospheric pressure.

0004

In the conventional injection device, it takes a certain amount of time for the first on-off valve and the second on-off valve to complete their opening and closing operations after receiving an operation command. Therefore, during the injection operation, the first on-off valve begins to open before the second on-off valve is completely closed. In this case, a portion of the pressure applied to the pipette from the injection pressure supply means leaks to the pressure regulating means through the second on-off valve, so that the internal pressure of the pipette becomes unstable at the beginning of the injection operation. Therefore, in the conventional injection device,
It is difficult to achieve the injection amount of the treatment substance according to the set value.

An object of the present invention is to provide an injection device that can maintain stable internal pressure of a pipette during injection operation.

[0006]

SUMMARY OF THE INVENTION The injection device of the present invention is a device for injecting a substance to be treated in a pipette into a substance to be treated. This injection device is
an injection pressure supply means having a first on-off valve and for increasing the internal pressure of the pipette and injecting the substance to be treated from the pipette into the substance to be treated; and an injection pressure supply means having a second on-off valve and for adjusting the internal pressure of the pipette. A pressure regulating means and a valve opening/closing means for opening and closing the first opening/closing valve and the second opening/closing valve so that the second opening/closing valve is completely closed when the first opening/closing valve is open. There is. Note that the pressure regulating means is connected to the pipette side of the injection pressure supplying means than the first on-off valve.

[0007]

[Operation] In the injection device of the present invention, at the start of the injection operation, the valve opening/closing means closes the second opening/closing valve and then opens the first opening/closing valve. Here, the valve opening/closing means opens the first opening/closing valve after the second opening/closing valve is completely closed. Further, at the end of the injection operation, the valve opening/closing means opens the second opening/closing valve after the first opening/closing valve is completely closed. Therefore, part of the pressure applied from the injection pressure supply means to the pipette during the injection operation is prevented from leaking to the pressure regulating means through the second on-off valve, so that the internal pressure of the pipette during the injection operation is stabilized. maintained.

[0008]

DESCRIPTION OF THE PREFERRED EMBODIMENTS FIG. 1 is a schematic diagram of a micromanipulator employing an embodiment of the present invention. In the figure, the micromanipulator includes a microscope 2 placed on a base 1, a pair of drive devices 3 placed on the sides of the microscope 2,
4, a control device 5 for controlling the microscope 2 and drive devices 3 and 4, and an injection device 6 as an embodiment of the present invention.

The microscope 2 has an operation table 7 in its center, and a container 8 such as a petri dish containing an object to be processed is placed on the operation table 7. An objective lens 9 is arranged below the operation table 7, and an adapter 1 is attached to the objective lens 9.
A television camera 10 is connected via 0a. The console 7 can be driven horizontally and vertically by a drive mechanism (not shown).

The drive devices 3 and 4 are placed on the base 1, and include a stand 11 and a coarse movement section 1 mounted on the stand 11.
2 and a fine movement section 13 attached to the upper part of the coarse movement section 12. The coarse movement section 12 can move in the vertical and horizontal directions with respect to the table 11 in units of several tens of micrometers by means of a stepping motor (not shown). The fine movement section 13 can move in units of 1 μm in the vertical and horizontal directions using an electromagnetic method. Arms 14 and 15 that are driven by the fine movement section 13 in the vertical and horizontal directions are provided at the end of each fine movement section 13 on the microscope 2 side. A capture needle 16 is attached to the tip of one arm 14. Capture needle 1
6 is a capillary whose tip extends toward the container 8 side. Furthermore, a tube from a suction pump (not shown) is attached to the other end of the trap needle 16 (not shown). The other arm 15 extends horizontally and has a micropipette 17 attached to its tip.

As shown in FIG. 2, the micropipette 17 is mainly composed of a pipette body 18 and a support portion 19 for supporting the pipette body 18. The pipette main body 18 is a cylindrical member with one end formed in a conical shape, and has a suction/discharge port 20 with a diameter of 0.1 to 10 μm at the tip. Further, the other end of the pipette body 18 is open and sealed with a sealing material 22 through which one end of the tube 21 is penetrated. Note that the other end of the tube 21 extends to the injection device 6. The support portion 19 is fixed to the tip of the arm 15. The support part 19 has a holder 23 for holding the pipette body 18. The holder 23 is attached to the arm 15 at a predetermined angle and grips the pipette body 18. Note that the pipette main body 18 supported by the support portion 19 has the suction/discharge port 20 side extending toward the container 8 .

The control device 5 includes a CRT 24, an operation panel 25, and a control unit 26. The operation panel 25 is provided with a joystick and various buttons for commanding movement of the arms 14, 15, etc. In addition, the control unit 26 includes a CPU, ROM, and RAM.
A microcomputer (not shown) is provided. With this microcomputer,
The microscope 2, drive devices 3, 4, CRT 24, etc. are controlled.

The injection device 6 has an operating section 27 on the front side. Moreover, the injection device 6 is
As shown in FIG. 3, it mainly includes a pressurizing device 28 inside. The pressurizing device 28 has a gas introduction pipe 30. A solenoid valve C is arranged on the inlet side of the gas introduction pipe 30. The inlet side of the electromagnetic valve C is connected to an inert gas cylinder 31 arranged outside the injection device 6 . The gas introduction pipe 30 is bifurcated, one side forming an injection gas supply path 32 and the other forming a pressure regulating gas supply path 33. The injection gas supply path 32 includes a first pressure regulator 34 and a solenoid valve A in this order from the upstream side. The first pressure regulator 34 is connected to the gas introduction pipe 30
The pressure required to inject the gas supplied from
It is configured so that it can be set at a pressure of about 300 to 650 kPa (usually about 300 to 650 kPa) and supplied to the solenoid valve A side. Injection gas supply path 3
The other end of 2 is connected to a gas exhaust path 35. The gas discharge path 35 has a membrane filter 36 and is connected to the other end of the tube 21 of the micropipette 17.

The pressure regulating gas supply path 33 has a second pressure regulator 37 and a solenoid valve B in this order from the upstream side. The second pressure regulator 37 controls the gas supplied from the gas introduction pipe 30 to a pressure (usually 100 kP) necessary to prevent capillary action at the suction/discharge port 20 of the micropipette 17.
It is configured such that it can be supplied to the solenoid valve B side. The other end of the pressure regulating gas supply path 33 is connected to a gas exhaust path 35 between the solenoid valve A and the membrane filter 36.
It is connected to the.

The above-mentioned injection device 6 includes a control section 46 as shown in FIG. The control unit 46
It is composed of a microcomputer including a PU 47, a RAM 48, a ROM 49, and an I/O port 50. The input side of the I/O port 50 is connected to the operating section 27 and other input sections. Furthermore, the output side of the I/O port 50 is connected to solenoid valves A, B, and C, pressure regulators 34 and 37, and other output sections. The control unit 46 determines the time c seconds required for the solenoid valve A to completely close after receiving the closing command, and the time a seconds required for the solenoid valve B to completely close after receiving the closing command. I remember.

Next, the operation of the injection device 6 will be explained according to the control flowcharts shown in FIGS. 5 and 6. Here, a case will be described in which a DNA solution is injected into cells.

When a main switch (not shown) of the control device 6 is turned on, the program starts. Step S
1, initial settings are performed. Here, valve A is closed and valve B is opened. When step S1 is completed, the program waits for the operator to input conditions. Here, when the operator inputs the injection pressure to be set in the first pressure regulator 34, the balance pressure to be set in the second pressure regulator 37, and the injection operation time from the operation unit 27, the program moves to step S3. In step S3, the solenoid valve C is opened to introduce inert gas from the gas cylinder 31 into the gas introduction pipe 30. Gas introduction pipe 30
The gas introduced therein flows into the injection gas supply path 32 and the pressure regulating gas supply path 33, and is stored at the pressure according to the conditions input into the first pressure regulator 34 and the second pressure regulator 37, respectively. Since the solenoid valve B is open, the gas (balance gas) stored in the second pressure regulator 37 is supplied into the pipette body 18 via the gas discharge path 35. As a result, the inert gas flows out from the suction/discharge port 20 of the pipette body 18 to the extent that capillarity can be prevented.

When step S3 is completed, the program waits for a key input from the operation section 27. Then, in step S4, the program determines whether or not the injection key has been pressed. Furthermore, in step S5, it is determined whether any other key has been pressed. If the operator does not make any key inputs, the program is in a waiting state.

When the operator turns on the injection key of the operation unit 27, the program moves from step S4 to step S6, where the injection subroutine shown in FIG. 6 is executed. Here, as shown in FIG. 7, the operator operates the micromanipulator in advance to aspirate and hold the cells 50 in the container 8 to the tip of the trapping needle 16, and the pipette body into which the DNA solution is injected. 18 is inserted into the cell 50. Note that the DNA solution is placed in the tube 2 of the pipette body 18.
Pipette main body 1 using a microsyringe from the 1 side end.
It is injected into 8. In this state, pipette body 1
Since the balance pressure from the second pressure regulator 37 is applied to the suction/discharge port 20 of No. 8, the suction/discharge port 20 prevents substances within the cell 50 (for example, cytoplasm) from flowing back due to capillary action.

In step S8 of the injection subroutine, a closing command is output to solenoid valve B. As a result, solenoid valve B starts its closing operation. Next, step S9
Now, the timer value (t) of the control unit 46 is set to 0. Subsequently, in step S10, it is determined whether the t value has reached a second (the time required for valve B to completely close). If the t value is not a second, the program returns t
It enters a waiting state until the value reaches a second. When the t value reaches a second, the program moves from step S10 to step S11. In step S11, an opening command is output to the solenoid valve A. As a result, solenoid valve A starts opening operation,
Inert gas is supplied from the first pressure regulator 34 into the pipette body 18 via the injection gas supply path 32 and the gas discharge path 35 . As a result, the internal pressure of the pipette body 18 increases, and the operation of injecting the DNA solution into the cells 50 from the suction/discharge port 20 is started.

Next, in step S12, it is determined whether a predetermined time b seconds have elapsed since the opening command was output to valve A (ie, whether t=a+b). The predetermined time b here is the time obtained by subtracting the time c required for the solenoid valve A to completely close from the injection time set in step S2. In step S12, the t value is a+b.
If it is not determined that the time is up, the program enters a standby state and continues to inject the DNA solution into the cells 50. If it is determined in step S12 that the t value is a+b seconds, the program moves from step S12 to step S1.
3, and issues a closing command to solenoid valve A. This results in
Solenoid valve A begins its closing operation.

Next, in step S14, it is determined whether c seconds have elapsed since the command to close the solenoid valve A was issued (that is, t
=a+b+c). t value is a+
If it is determined that it is not b+c seconds, the program enters a standby state. Therefore, the operation of injecting the DNA solution continues. When it is determined that the t value is a+b+c seconds,
The program moves from step S14 to step S15. In step S15, a command to open the solenoid valve B is output. As a result, the electromagnetic valve B starts to open, so that the balance gas starts to be supplied from the second pressure regulator 37 into the pipette body 18, and the backflow of substances in the cells 50 due to capillary action at the suction port 20 is prevented. When step S15 ends, the program returns to the main routine of FIG.

The above injection operation will be explained with reference to the timing chart of FIG. In addition, in the chart of electromagnetic valve A and electromagnetic valve B of FIG. 8, the solid line portion shows the state of the command from the control unit 46. Further, the portions indicated by dotted lines indicate the actual opening/closing operation status of the solenoid valves A and B. As can be seen from FIG. 8, in this example,
After solenoid valve B completely closes, solenoid valve A begins to open.
Also, since solenoid valve B starts to open after solenoid valve A is completely closed, gas from first pressure regulator 34 is transferred to second pressure regulator 34.
There is no leakage from the pressure regulator 37 to the outside. Therefore, during the injection operation, the internal pressure of the pipette body 18 is maintained stably. As a result, the volume of the DNA solution injected from the pipette main body 18 into the cells 50 is the same as the set value. Further, since the inert gas supplied from the gas cylinder 31 does not leak out from the second pressure regulator 37 more than necessary, the inert gas can be saved.

When the operator presses any other key on the operation section 27, the program proceeds from step S5 to step S7.
, and the operation corresponding to the key operated by the operator is performed here. When step S7 is completed, the program enters the standby state again.

[Other Embodiments] (a) FIG. 9 is a schematic diagram of another embodiment. Pressure device 28a of this embodiment
is an injection gas supply path 32 configured similarly to the embodiment described above.
and a pressure regulating gas supply path 33, and further includes a sample suction path 51 and a gas discharge path 52 provided in parallel with these supply paths 32, 33. The sample suction path 51 includes a third pressure regulator 53, an ejector 54, and a solenoid valve D in this order from the upstream side. The ejector 54 has a gas exhaust path 54a. The gas discharge passage 52 is a solenoid valve E.
have. In this embodiment, a highly viscous sample such as a DNA solution can be aspirated into the pipette body 18. Here, when the solenoid valves A, B, and E are closed and the solenoid valve D is opened, the inside of the pipette body 18 is caused by the inert gas pressurized by the third pressure regulator 53 and spouted from the gas outlet 54a of the ejector 54. The pressure becomes negative and the suction/exhaust port 2
DNA solution is aspirated from 0. In addition, in this embodiment, when solenoid valve D is opened after closing solenoid valves A, B, and E,
The internal pressure of the pipette body 18 increases rapidly, and inert gas is vigorously jetted out from the suction/discharge port 20 of the pipette body 18. This eliminates clogging of the pipette body 18.

(b) In the above embodiment, the pressurizing device 28
Although the inert gas is supplied from the gas cylinder 31, a compressor may be used instead of the gas cylinder 31.

(c) In the above embodiment, the pressurizing device 28
Although the pressure regulating gas supply channel 33 is provided in parallel with the injection gas supply channel 32, the inlet side of the electromagnetic valve B may be left open without providing such a pressure regulating gas supply channel 33. However, in this case, except during injection operation, the pipette body 1
The internal pressure of 8 becomes atmospheric pressure.

[0028]

Effects of the Invention The injection device of the present invention has the valve opening/closing means as described above. Therefore, according to the present invention, the internal pressure of the pipette can be stably maintained during the injection operation.

[Brief explanation of drawings]

FIG. 1 is a schematic diagram of a micromanipulator employing an embodiment of the present invention.

FIG. 2 is an enlarged partial view of the micromanipulator.

FIG. 3 is a schematic configuration diagram of an embodiment of the present invention.

FIG. 4 is a schematic diagram of the control section of the embodiment.

FIG. 5 is a control flowchart of the embodiment.

FIG. 6 is a control flowchart of the embodiment.

FIG. 7 is a diagram showing the injection operation status by the micromanipulator.

FIG. 8 is a timing chart of the embodiment.

FIG. 9 is a diagram corresponding to FIG. 3 of another embodiment.

[Explanation of symbols]

6 Injection device 18 Pipette body 32 Injection gas supply path 33 Pressure regulating gas supply path 34 First pressure regulator 37 Second pressure regulator A Solenoid valve B Solenoid valve

Claims (1)

[Claims] 1. An injection device for injecting a substance to be treated in a pipette into a substance to be treated, the injection device having a first on-off valve, increasing the internal pressure of the pipette to inject the substance from the pipette into the substance to be treated. For adjusting the internal pressure of the pipette, the pipette has an injection pressure supply means for injecting the treatment substance, and a second on-off valve, and is connected to the pipette side of the first on-off valve of the injection pressure supply means. and a valve opening/closing means for opening and closing the first opening/closing valve and the second opening/closing valve so that the second opening/closing valve is completely closed when the first opening/closing valve is open. An injection device equipped with and.