JP3906424B2 - Dispensing method for chemicals, etc. - Google Patents

Description

The present invention relates to a dispensing apparatus for chemicals used for dispensing various reagents in physicochemical tests and the like, and for quantification and qualitative analysis of chemicals and other target liquids, particularly when producing protein crystallization reagent kits. The present invention relates to a dispensing method suitable for reagent dispensing.

  Protein crystallization, which is a main field of application of the present invention, is performed by a combination of reagents such as a precipitant, a buffer, and an additive. Among them, when dispensing a precipitant with high viscosity, there are problems that reproducibility cannot be obtained if it is performed manually and that it takes time to prepare the reagent. In addition, various plates are often used in crystallization experiments, and the volume is as small as 0.5 μl. As an advantage of using a dispenser, it is important that reagent preparation for protein crystallization can be automated with high reproducibility, and that detailed studies can be carried out efficiently, especially in cases related to bio-related and drug discovery research. Many.

  In the preparation of each solution for crystallization, conventionally, a mixture of undiluted solutions having different viscosities of several kinds of solutions is weighed by a manual volume. For this reason, reproducibility is low, time and effort are required, and mistakes are likely to occur. In addition, it is necessary to check the reagent after preparation with a pH meter or the like. Conventional liquid handlers widely used in this field use a single syringe as shown in FIG. 5 and employ an air gap to avoid mixing of filling water and reagents in the line.

  That is, this syringe configuration is for dispensing various kinds of chemical solutions with a single needle (nozzle), and as shown in the figure, a valve is provided on the head side of the syringe 51 for sucking and discharging chemical solutions such as water and reagents. A transfer tube 53 is connected via 52, and a needle (nozzle) 54 serving as a suction and discharge port for a chemical solution is connected to the tip of the transfer tube 53 to form a chemical solution suction / discharge (dispensing) line.

  As shown in the steps A to D in FIG. 3, the suction and discharge (dispensing) work by the above apparatus is in the discharge position in which the piston of the syringe 51 is stroked to the deepest part in the initial state A. The tube 53 and the needle 54 are preliminarily filled with water (purified water) 55 as a cleaning liquid, and the needle 54 is inserted into a reagent container (bottle) 57 containing a reagent 56.

In the reagent suction step B, the syringe 51 is operated in the suction direction to suck the reagent 56 in the reagent container 57. The time between the water 55 and the reagent 56 in the line are formed air formic cap 58 to move the line along with the water 55 and the reagent 56 has a structure in which both are not mixed (contamination).

  In the subsequent dispensing step of C to the crystallization plate 59, the syringe 51 is stroked in the discharge direction, and the reagent is sequentially predetermined from the needle 54 into a large number of wells (for example, 6 × 12 = 72) of the reagent kit plate 59. Are discharged (dispensed) in predetermined order. At this time, the dispensing of the reagent is finished on the leading end side from the air gap 58 in the line, the dispensing is completed as shown in FIG. 4D, and the operation returns to the initial step A.

  In this way, by forming an air gap in the line, the technology for preventing the contamination of different chemicals such as water and reagent is also applied to dispensing devices for analyzers such as blood and urine as shown in Patent Document 1. in use. In addition, an apparatus shown in Patent Document 2 is known in which a water injection line for applying a dispensing pressure is connected to the middle of a line between a reagent and a needle (nozzle) without using an air gap.

JP-A-6-213907 JP 2003-302410 A

In the method of forming an air gap in the chemical liquid suction / dispensing line as in the apparatus shown in FIG. 5 or Patent Document 1 described above to prevent contamination between the chemical liquids, the reagent suction is performed so as not to break the air gap. In this case, there is a problem that the flow rate cannot be generated, and even if the flow rate condition is determined, the condition setting cannot be reproducible due to the contamination of the inner wall of the transfer tube.
Further, the above method has a drawback that the flow rate condition needs to be set every time because the volume that can be dispensed is adjusted by changing the volume of the transfer tube.

  In the method disclosed in Patent Document 2, it is understood that mixing of the mixed water and the next reagent can be suppressed by mutual diffusion of the washing water and the reagent. ) Side is not washed, so mixing the needle and the previous reagent adhering to the upstream side with the next reagent is inevitable, and the needle-like reagent suction port itself is different unless it is replaced for each reagent. There is a drawback that mutual contamination is unavoidable.

  In addition to the reagent aspiration / dispensing line, all of the above-mentioned patent documents separately form a line for supplying cleaning liquid and water, and further installing a syringe and other actuators for aspiration / discharge in the other line. There are also disadvantages such as complicated mechanisms.

The present invention is intended to provide a dispensing method for solving the above-described problems of the prior art, and is different at the proximal end side and the distal end side of the line for aspirating / dispensing chemical liquid and other target liquids , respectively. Via the valves 17 and 18 for preventing the mixing of the target liquid, the cleaning liquid 2 that mainly sucks and fills the cleaning liquid into the line and cleans the line by the suction and discharge of the cleaning liquid, and the target liquid other than the cleaning liquid is sucked Then, the dispensing syringe 3 for sucking and filling the target liquid into the line and the syringe and dispensing the target liquid is connected, and the needle 14 for dispensing and sucking and discharging the target liquid is connected to the tip of the line. in the method for sucking-dispensing of drug solution from the needle 14 by the suction and discharge operation movement of the operating and syringe of the valves 17 and 18,
A. An initial step of filling the needle 14 and the line with the cleaning liquid in a state where the syringes 2 and 3 are respectively stroked to the discharge side;
B. A step of aspirating the cleaning syringe 2 to suck the chemical solution from the needle 14, storing the cleaning solution in the line in the cleaning syringe 2, and filling the chemical solution in the tip side of the dispensing syringe 3;
C. A step of aspirating the dispensing syringe 3 to store the chemical in the syringe;
D. The dispensing syringe 3 is discharged,
E. Completing the dispensing; and
F. A step of discharging the cleaning syringe 2 by discharging the cleaning syringe 2 while the dispensing syringe 3 is stopped in the discharging state, and discharging the remaining chemical solution in the line;
G. A step of suctioning the cleaning syringe 2 to suck the cleaning liquid from the needle 14 and filling the cleaning liquid in the line and the cleaning syringe 2;
H. Further, a process of cleaning the cleaning syringe 2 and the line by discharging the cleaning syringe 2;
It is characterized by consisting of .

  According to the present invention configured as described above, at least two syringes for washing and dispensing are used, filling water in the line is removed with the other syringe, and the line is filled with a reagent. Since the method is adopted, there is no need to use an air gap, and there is an advantage that an arbitrary flow rate can be set for reagent dispensing.

Flow rate condition reproducibility can be ensured without causing flow rate obstruction due to dirt on the inner wall, and the transfer volume can be adjusted by fixing the transfer tube volume and changing the syringe volume or operating speed. Therefore, it is possible to speed up the continuous dispensing into a large number of containers, and it is also suitable for producing a large amount of reagent set.
In addition, since the syringe to the needle can be arranged in a single line, the configuration is simple, and it does not take up space in the apparatus, and is excellent in operability and other handling characteristics.

Next, the structure of a liquid handler and a syringe for preparing a reagent kit related to the apparatus of the present invention will be described.
FIG. 1 shows an overall view of a Gilson liquid handler used for preparing the reagent kit of the present invention. In this example, two syringes 2 and 3 are supported on a box-shaped body case 1 for sucking and discharging drugs. The dispensing unit 4 is installed. The space on the front side of the main body case 1 is a rectangular work space 6. Above this work space 6, a horizontal arm 7 that horizontally moves in the left-right (X-axis) direction by an actuator (the actuator itself is not shown). Is protruded in front of the main body case 1. In the work space 6, a plate for preparing a reagent and other containers are arranged in accordance with each dispensing operation.

  The lateral arm 7 is provided with a groove-shaped guide 9 in the front-rear (Y-axis) direction, and is vertically guided in the vertical (Z-axis) direction, which is guided by the guide 9 and moves in the Y-axis direction. A column 11 protrudes upward, and a slider 13 with an actuator that moves up and down by an actuator along a vertical groove-shaped guide 12 is attached to a side surface of the column 11. A needle 14 for dispensing a chemical solution is attached to the slider 13 in the vertical direction, and a transfer tube 16 connected to the above-described syringe 3 is connected to the proximal end side of the needle 14.

  A drive device (actuator) for horizontally moving the horizontal arm 7 and the vertical column 11 is built in the main body case 1, and an actuator for driving the two syringes 2 and 3 is built in the dispensing unit 4. Yes. Except for the fact that two syringes are installed, the liquid handler is substantially the same as a conventionally known one.

  The two syringes of the liquid handler used in this embodiment are divided into a syringe 14 for washing the needle 14 and transfer tube 16 and a syringe 3 for dispensing chemicals as shown in FIG. Valves 17 and 18 are provided. The syringe heads of both the syringes 2 and 3 and the valves 17 and 18 are communicated with each other by a flow path 19. The flow path 19 including the valves 17 and 18, the transfer tube 16, and the needle 14 are used for sucking and discharging chemicals ( A series of lines for dispensing) are formed. When each of the valves 17 and 18 operates the dispensing cylinder to suck and dispense the reagent, the two kinds of chemical liquids (cleaning liquid and reagent) do not cause mixing (contamination) in the suction / discharge line. Is automatically opened and closed.

  As shown in the operation process diagrams of A to I in FIG. 2, the syringe structure is configured such that in the process A, the pistons 21 and 22 of the syringes 2 and 3 are respectively stroked to the syringe head side, The lines of the transfer tube 16 and the needle 14 are filled with a cleaning liquid (purified water). This filling liquid (water) is for preventing the generation of bubbles (air gap) in the line and ensuring the stability of the flow rate and the reagent dispensing amount.

  Then, as shown in Step B, the syringe 2 for cleaning liquid suction is operated in the suction direction while the syringe 3 for dispensing is stopped in the discharge (push-out) completion state, and the cleaning liquid and the reagent 24 in the reagent container 23 are driven by the needle 14. , The cleaning liquid in the line is stored in the syringe 2 and the reagent 24 is filled in the line to be filled. At this time, the suction by the cleaning syringe 2 may be a minimum amount for filling the line with the reagent, and the cleaning liquid is stored in a state of being mixed with the reagent.

  Subsequently, as shown in Step C, the dispensing syringe 3 is operated in the suction direction to store a sufficient amount of the reagent 24 in the syringe 3. The stored reagent 24 is sequentially dispensed into each well (concave portion) of the reagent plate 26 similar to that shown in FIG. 3 by causing the syringe 3 to act in the pushing direction in the D step, and as in the E step. The dispensing to each plate 26 is completed.

  Next, in the state where the dispensing syringe 3 is stopped in the extruding state in the F step, the syringe 2 is pushed out to push out the cleaning liquid and the residual reagent in the line is discharged, and then the syringe 2 is sucked in the G step. By operating and suctioning the cleaning liquid 28 from the cleaning liquid container 27 into the syringe 2 and all lines, the line and the syringe 2 are filled with the cleaning liquid. After that, the reagent 2 remaining in the line is diluted with water by repeating extrusion and suction of the syringe 2 at least twice as in the H and I steps, and the cleaning in the syringe 2 and the line is completed. Reset to A. Such a dispensing method is repeated for each chemical solution constituting the reagent, and the operation control of the liquid handler itself is performed by a personal computer or other control device based on a predetermined program. The microcomputer unit 32 has a built-in control program for controlling the next suction / discharge operation based on the detection signal of the chemical liquid suction / discharge operation by the syringe of the liquid handler.

  FIG. 3 is a block diagram showing an example of the control device of the device of the present invention. The controller 31 includes a control microcomputer unit 32, an input interface 33 connected to the microcomputer unit 32, and an output interface 34. On the input interface 33 side, flow meters 35a and 35b of the two syringe valves 17 and 18 and syringe stroke timers 36a and 36b of the syringes 2 and 3 are connected as input units (detection units).

  The other output interface 34 side is connected with syringe motors 37a and 37b, which serve as actuators for operating the syringes 2 and 3 as output units, and solenoids 17a and 18a for the syringe valves. By using the chemical flow rates in the valves 17 and 18 as input signals, the syringes 2 and 3 and the operating parts of the valves 17 and 18 are actuated, whereby the chemical liquid is quantitatively sucked and discharged. The movement of the needle 14 in the works base 6 of the liquid handler is controlled by a driving actuator by a three-dimensional driving program set so as to be linked with the dispensing operation.

  FIG. 4 shows a control flowchart of the dispensing operation by the syringe and the control device. After the control is started, if the drug solution remains in the syringe as a result of the check, the drug solution is discharged and washed while controlling the flow rate. After the cleaning or when there is no remaining chemical, suction of the chemical into the syringe is started.

  At that time, when the stroke speed of the syringe is maintained at a predetermined speed, suction is performed as it is, and when the speed is exceeded, the stroke speed and the flow rate control are sequentially performed to perform discharge and washing. The operation control detection unit is not limited to a flow meter or a timer. For example, a syringe stroke amount detection switch, a liquid amount detector provided on the side of the chemical solution container, or the like can be used. is there.

  Table 1 shows a setting example of the flow rate (unit ml / min) when various chemical solutions are dispensed by the apparatus of the present invention. As described above, in the present invention, in order to dispense a highly viscous chemical solution, a flow rate corresponding to the viscosity of each chemical solution is set, and generation of bubbles during dispensing can be prevented, and quantitative determination can be performed with high reproducibility. It was. An example of setting the flow velocity is given below, but it is also possible to set the flow velocity with a parameter quantified for each item.

  As described above, conventionally, a single syringe is used, and an air gap is used as a boundary area between the filling water (cleaning liquid) in the circuit and the chemical liquid in order to avoid mixing the filling water (cleaning liquid) and the reagent in the line. Because the reagent is aspirated so as not to break the air gap, the flow rate cannot be obtained, and even if the flow rate condition is determined, the condition setting cannot be reproducible due to the contamination of the inner wall of the transfer tube. In order to adjust the volume that can be dispensed by changing the volume of the liquid, the flow rate condition must be set each time.

  According to the above-described apparatus and method configured as described above, since two syringes are used, the cleaning liquid is removed with the other syringe, and the line is filled with the reagent, there is no air gap. The reproducibility of the flow rate condition can be ensured without being disturbed by dirt on the inner wall. Moreover, since the volume of the transfer tube is fixed and the dispensing volume can be adjusted by changing the volume of the syringe, there is an advantage that the flow rate condition setting is simple.

  In addition, when considering the application field using the liquid handler of the above example, it is possible to dispense several kinds of highly viscous solutions, as well as small and large volume solutions by changing the syringe size It is possible to deal with dispensing. Therefore, it can be used for quantifying solutions (solvents) in the field of organic chemistry and synthesis, or for quantifying solutions that are difficult to handle (high viscosity, high volatility, etc.).

  The apparatus and method of the present invention can be used for dispensing reagents and chemicals into test containers in various physical and chemical tests.

It is a whole perspective view which shows the outline | summary of the liquid handler provided with the apparatus of this invention. It is a schematic diagram which shows the structure of the apparatus of this invention, and the process of a chemical | medical solution dispensing operation. It is a block diagram which shows the structural example of the control apparatus of this invention apparatus. It is a control flowchart of this invention apparatus. It is a schematic diagram which shows the structure of the conventional dispensing apparatus and the process of the chemical | medical solution dispensing operation | work.

Explanation of symbols

2 Syringe for washing 3 Syringe for dispensing 14 Needle (nozzle)
16 Transfer tube 17, 18 Valve 19 Flow path 21, 22 Piston 23 Reagent container (bottle)
24 Reagents 26 Plate 27 Cleaning solution container (bottle)
28 Purified water (cleaning solution)
31 Controller 35a, 35b Flow meter (detector)
36a, 36b Timer (detection unit)

Claims (1)

The cleaning liquid is mainly fed into the line through valves (17) and (18) for preventing mixing of different target liquids at the base end side and the distal end side of the line for sucking and dispensing chemical liquids and other target liquids. A syringe for cleaning (2) that sucks and fills and cleans the line by sucking and discharging the cleaning liquid, and sucks the target liquid other than the cleaning liquid, sucks and fills the target liquid into the line and the syringe, and dispenses the discharged liquid. dispensing connects the syringe (3), connect the needle (14) for dispensing and intake and out of the target liquid to the tip of the line, each valve (17), and the operation of (18) syringes in the method by the suction and discharge operation movement of the aspirating-dispensing of chemical liquid from the needle (14),
A. An initial step of filling the needle (14) and the line with the cleaning liquid in a state where the syringes (2) and (3) are respectively stroked to the discharge side;
B. The washing syringe (2) is aspirated to suck the chemical solution from the needle (14), and the washing solution in the line is stored in the washing syringe (2) and is placed in the line on the tip side from the dispensing syringe (3). A step of filling the chemical solution;
C. A step of aspirating the dispensing syringe (3) to store the chemical in the syringe;
D. Dispensing the dispensing syringe (3),
E. Completing the dispensing; and
F. A step of discharging the cleaning syringe (2) by discharging the cleaning syringe (2) in a state where the dispensing syringe (3) is stopped in the discharging state, and discharging the remaining chemical solution in the line;
G. A step of suctioning the cleaning syringe (2) to suck the cleaning liquid from the needle (14) to fill the cleaning liquid in the line and the cleaning syringe (2);
H. Further, a step of cleaning the cleaning syringe (2) and the line by operating the discharge of the cleaning syringe (2),
Dispensing method for chemicals consisting of