WO2004063755A1

WO2004063755A1 - Automatic analyzer with sample container and cleaning method of sample container

Info

Publication number: WO2004063755A1
Application number: PCT/JP2003/000062
Authority: WO
Inventors: Yoshiyuki Shoji; Toshiaki Yokobayashi
Original assignee: Hitachi High-Technologies Corporation
Priority date: 2003-01-08
Filing date: 2003-01-08
Publication date: 2004-07-29
Also published as: JPWO2004063755A1; JP4154390B2

Abstract

An automatic analyzer where a sample container is cleaned through at least two processes, and a cleaning method of a sample container. At first, the sample container is brought into contact, in the vicinity of the opening thereof, with the vicinity of the discharging hole of a cleaning vessel thus forming a vessel section, i.e. a cleaning space where a liquid can be reserved. Surface of the vessel section includes the surface of the sample container but does not include the opening part. Cleaning liquid is injected into the vessel section, a specified quantity of the cleaning liquid is reserved and then a part of the sample container (excluding the opening part) is cleaned. Subsequently, a specified gap is formed between the sample container and the cleaning vessel under a state where a specified quantity of the cleaning liquid is reserved in the vessel section. The reserved cleaning liquid flows out from the gap and cleans the opening part before being discharged to the discharging hole of the cleaning vessel.

Description

Specification

Automatic analysis with a sample container, and a method for cleaning the sample container

The present invention relates to an automatic analyzer provided with a sample container, and a method for cleaning a sample container. For example, the present invention relates to a nucleic acid extracting device provided with a nucleic acid capturing chip having a built-in nucleic acid capturing solid phase, and a method for cleaning a nucleic acid capturing chip. Background art

Japanese Patent Application Laid-Open No. H11-1266664 describes a method for automating nucleic acid extraction using a nucleic acid capturing chip containing a silica-containing solid phase. Then, the solid phase to which the nucleic acid is bound and the inside of the chip are washed, and the waste liquid is discharged. At this time, the cleaning liquid adheres to the outer wall of the chip.

The amount of the elution reagent used in the elution step performed after the washing is very small. For this reason, if the washing solution remains on the chip and mixes with the elution reagent, the concentration of the eluent changes, disturbing the nucleic acid recovery rate.

In addition, the cleaning liquid adhering to the outer wall of the chip collects at the tip of the chip with the passage of time to form a liquid droplet. The liquid droplets scatter when the nucleic acid capturing chip is moved, causing contamination.

An object of the present invention is to prevent generation of contamination and to efficiently wash a sample container. DISCLOSURE OF THE INVENTION.

The present invention provides an automated analysis system that cleans sample containers in at least two steps. This is a method for cleaning the device and the sample container. First, the vicinity of the opening of the sample container and the vicinity of the discharge hole of the washing container are brought into contact with each other to form a vessel which is a washing space capable of storing liquid. The surface of this vessel includes the surface of the sample container, but does not include the opening. A cleaning solution is injected into the container, a predetermined amount of the cleaning solution is stored, and a part of the sample container (a portion not including the opening) is washed. Next, a predetermined gap is formed between the sample container and the cleaning container while a predetermined amount of the cleaning liquid is accumulated in the vessel. The accumulated cleaning liquid flows out of the gap, cleans the opening, and is discharged into the discharge hole of the cleaning container.

In the present invention, portions other than the opening are cleaned first, and then the vicinity of the opening is cleaned. For this reason, the sample contamination near the opening does not diffuse to the entire sample container via the cleaning liquid. Also, by defining the volume of the cleaning liquid and the shape of the gap, the flow of the cleaning liquid flowing out of the gap can be adjusted. By controlling this flow, the cleaning liquid always reaches the specific location even if the opening has a structure that is difficult to wash, such as irregularities. Therefore, even a sample container having a structure that is difficult to wash can be reliably washed every time.

Further, the present invention is an automatic analyzer and a cleaning method for discharging a sample to a discharge hole isolated from the outside, and thereafter cleaning the sample container and the discharge hole. First, the vicinity of the opening of the sample container and the vicinity of the discharge hole of the washing container are brought into contact with each other to form a vessel capable of storing liquid. At this time, the opening of the thought container is placed in the discharge hole isolated from the container. Next, the sample in the sample container is discharged to the discharge hole, and the cleaning liquid is injected into the container. The order of sample discharge and washing liquid injection is not specified. Next, a gap is formed between the sample container and the washing container, and the accumulated washing liquid flows into the discharge hole. The cleaning liquid flowing out of the gap cleans the opening and the discharge hole. In the present invention, the sample is discharged to the discharge hole, which is a region isolated from the outside. This isolated area is cleaned while maintaining the isolated state with the cleaning liquid. Since the droplets of the sample generated by the discharge are present only inside the isolated area and do not diffuse outside, the danger of contamination due to the scattering of the sample droplets can be avoided. For this reason, for example, even when fluid resistance such as a solid phase is present in the sample container and the discharge of the sample is obstructed, the sample can be forcibly discharged by high pressure.

Hereinafter, the above and other novel features and effects of the present invention will be described with reference to the drawings. BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 is a schematic diagram of a cleaning unit in one embodiment. FIG. 2 is an explanatory diagram of cleaning unit operations a to f in the cleaning step of one embodiment. FIG. 3 is a schematic view of a liquid dispensing channel in one embodiment. FIG. 4 is an explanatory diagram of an operation of removing a chip from a nozzle in one embodiment. FIG. 5 is a schematic view of a chip for capturing nucleic acids according to one embodiment. FIG. 6 is a plan view of an apparatus for purifying nucleic acid according to one embodiment. FIG. 7 is a schematic external view of an apparatus for purifying nucleic acid in one embodiment. FIG. 8 is a block diagram showing a configuration of an electric system in one embodiment. FIG. 9 is a flowchart of a cleaning step in one embodiment. BEST MODE FOR CARRYING OUT THE INVENTION

Hereinafter, an apparatus for purifying nucleic acid according to an embodiment of the present invention will be described with reference to FIG. 1 to FIG. .

FIG. 6 is a plan view of the nucleic acid purification device, and FIG. 7 is a schematic external view of the nucleic acid purification device. The nucleic acid purification apparatus 100 includes two arms 16 and 33 that can move in the horizontal direction (X direction). One arm 16 holds a nozzle holder 17 for holding a dispensing nozzle 36 (not shown), It is provided so that it can move in the horizontal direction (Y direction) along the length direction. The other arm 33 has a nozzle holder 34 holding a reagent discharge nozzle 39 (not shown) so as to be movable in the horizontal direction (Y direction) along the length of the arm 33. The reagent discharge nozzle 39 is provided with a plurality of nozzles, a nozzle capable of mounting a disposable tip and a dispenser nozzle capable of directly discharging a reagent without using a disposable tip. The nozzle holders 17 and 34 can operate in the vertical direction (Z direction) with respect to the corresponding arms 16 and 33.

On the work surface 5 of the main body stand, two tip racks 14a and 14b on which a large number of unused dispensing tips 15 are placed are set in a predetermined area. As shown in FIG. 3, these tip racks 14a and 14b have holes into which each dispensing tip 15 can be inserted, and the tip of the dispensing tip is a work surface. 5 or a box with a height that does not touch the bottom of the chip black. Each chip black 14a can hold up to 96 dispensing tips 15, and 14b can hold up to 48 dispensing tips 15.

Further, on the work surface 5, a chip rack 30 on which a large number of unused nucleic acid capturing chips 31 are placed is set in a predetermined area. The shape of the tip rack 30 is the same as that of the tip rack 14 described above. In this example, a maximum of 96 nucleic acid capturing chips 31 can be held on the chip rack 30.

On the work surface 5, the sample rack 12 is set in a predetermined area. The sample rack 12 can contain a sample to be processed, that is, a sample containing nucleic acids. In this embodiment, each sample rack 12 can hold a maximum of 48 sample containers 13.

Also, on the work surface 5, container racks 23a and 23b holding many unused processing containers 24a and 24b are set in a predetermined area. Container rack Each of 23 a and 23 b can hold a maximum of 64 processing containers 24. Further, the container racks 23a and 23b have a temperature control function, and can heat and cool the processing containers 2424b. By heating, nucleic acids can be efficiently eluted from the nucleic acid capturing carrier. In addition, the cooling prevents the solution containing the eluted nucleic acid from evaporating, and can be stored for a certain period of time. On the work surface 5, a container storage rack 25 holding many unused purified product containers 26 is set in a predetermined area. The purified product container 26 collects the purified nucleic acid-containing liquid for each sample. In this embodiment, the container rack 25 has a maximum of eight large-sized refined product containers 26a, 96 small-sized product containers 26b, and 96 holes in one rack. Holds up to two containers 26 c.

Further, the work surface 5 is provided with a chip remover 27a shown in FIG. The chip remover 27a removes the dispensing tip 15 and the nucleic acid capturing tip 31 connected to the dispensing nozzle 36 from the nozzle. Also, it is the home position of dispensing nozzle 36.

Further, the work surface 5 is provided with cleaning sections 18 and 28. The washing unit 18 also has a function of discharging unnecessary liquid from the nucleic acid capturing chip 31.

FIG. 1 shows the details of the cleaning section 18. The washing section 18 is composed of a washing water inlet 101, a water storage tank 102, a washing tank 103, a waste liquid hole 104, and a bypass flow passage 105.

The washing water inlet 101 is a flow path for injecting the washing liquid into the main washing section, and is connected to a washing liquid supply pump (not shown).

The water reservoir 102 is a portion for temporarily storing a cleaning liquid in order to remove liquid droplets attached to the tip of the nucleic acid capturing chip. At the lower part of the water storage tank 102, a flow path for discharging the cleaning liquid is provided. From this flow path, the cleaning liquid is supplied to the cleaning tank 103. Supply. This flow path does not require an on-off valve, and the cross-sectional area of this flow path and the amount of reagent supplied from the cleaning liquid inlet 101 balance each other so that the liquid level in the water storage tank 102 is determined. It has become.

The washing tank 103 is a vessel for washing the outside of the nucleic acid capturing chip 31 and treating the drainage. The inner side surface of the washing tank 103 has a substantially rectangular parallelepiped shape, and its cross-sectional area is larger than the outer diameter of the nucleic acid capturing chip 31. The bottom of the cleaning tank 103 has a tapered shape tapered downward, and has a discharge hole 104 at the tip. The washing tank 103 can be inserted without contacting the nucleic acid capturing chip 31 with the inner side surface thereof. When the tapered portion of the nucleic acid capturing chip 31 comes into contact with the tape-like bottom surface of the washing tank 103, it is possible to prevent the washing solution from being discharged to the waste liquid hole 104. As a result, the cleaning liquid can be stored in the cleaning layer 103. At this time, the distal end portion 48 of the nucleic acid capturing chip 31 is in a state of entering the discharge hole 104.

The waste liquid hole 104 is a discharge hole through which the waste liquid can be discharged to the outside of the cleaning unit, and has a cylindrical shape having a cross-sectional area larger than a tip end 48. Thus, when the nucleic acid capturing chip 31 is inserted into the washing tank 103, the distal end portion 48 fits in the waste liquid hole 104.

The bypass flow path 105 is a flow path having one end connected to the inside of the washing tank 103 and the other end connected to the waste liquid hole 104. Excess cleaning reagent in the cleaning tank 103 is discharged to the waste liquid hole 104 through this channel. By the arrangement of the main flow path, the amount of the cleaning reagent stored in the cleaning tank 103 can be regulated. In addition, there is a high possibility that the sample is contaminated in the vicinity of the tip end 48 where the sample is sucked and discharged. However, if the level of the cleaning liquid when full is higher than the expected contamination site, the contamination of the chip can be surely cleaned.

In addition, the stand 100 has a washing solution potter 19, an eluent potter 20, and dilution 3 000062

Various reagent containers such as a liquid bottle 21 and a binding promoter 22 are arranged (not shown). The washing solution potter 19 contains a washing solution for washing the solid phase in the nucleic acid capturing chip 31. The eluent bottle 20 contains an eluent for eluting the nucleic acid bound to the solid phase. The diluent pottor 21 contains a diluent, and the binding promoter bottle 22 contains a solution of a binding promoting substance that promotes binding of nucleic acid to a solid phase.

The peripheral structure of the dispensing nozzle 36 will be described with reference to FIG. The dispensing nozzle 36 is connected to a pump for suction, discharge, stirring, reagent discharge, and air discharge. The syringe pump 10 is a pump for performing suction / discharge and stirring in the dispensing tip 15 and the nucleic acid capturing tip 31. The syringe pump 80 is a pump that discharges the solid phase washing reagent to the nucleic acid capturing chip 31 and is connected in plurality. In addition to the syringe pump, a pump having a liquid sending function such as a bellows pump and a peristaltic pump can be used. The discharge pump 87 is a pump for discharging the remaining liquid in the solid phase of the nucleic acid capturing chip 31 to the outside of the chip. In addition, a flow path connecting the discharge pump 87 and the chip is connected to a valve 82 for opening to the atmosphere and a filter 86 for preventing intrusion of substances from the outside.

The attachment / detachment of the dispensing tip 15 and the nucleic acid capturing tip 31 to / from the dispensing nozzle 36 will be described with reference to FIG.

The tip is mounted by lowering the nozzle and fitting the tip to the tip of the nozzle on the tip racks 14a, 14b, 30. For chip removal, a chip remover 27a is used. The chip remover 27a has a plate-like member at a predetermined height position. A slit 55 having a width smaller than the outer diameter of the tip heads 52 and 54 and larger than the outer diameter of the dispensing nozzle 36 is formed on this plate-shaped member. Chip removal should be done in slit 55 With the heads 52, 54 of each chip lower than the height at which they are located, the nozzle 36 is moved horizontally so as to enter the slit. Next, the nozzle holder 117 is raised to bring the heads 52, 54 into contact with the lower surface of the plate-shaped member. Then, the tips 15 and 31 fall out of the dispensing nozzle 36 due to the further rise of the nozzle holder. The dropped chips fall into a chip disposal port 50 (not shown) and are collected in a collection box (not shown). Disposal of tips for dispensing reagents is performed in the same manner using a tip remover 27b.

The configuration of the electrical system of the nucleic acid purification device will be described with reference to FIG. The personal computer (PC) 60 as an operation control unit has a keyboard 61 as an operation panel for inputting operation conditions and specimen information, and a CRT as a display device for displaying input information and warning information. 62, a mechanism control section 65 for controlling each mechanism section of the apparatus, and the like. The mechanism control section 65 controls four stepping motors 71 to 74, two AC servomotors 75 and 76, a bellows pump 80, and two solenoid valves 81 and 82. The stepping motor 71 drives the syringe 10 of the syringe pump to perform a suction / discharge operation. The stepping motor 72 drives the syringe 32 of the syringe pump 80 to perform a suction / discharge operation. In the stepping mode 73, the nozzle holder 17 is moved horizontally and vertically. The stepping motor 74 moves the nozzle holder 34 horizontally and upward and downward. The AC supporter 7 '5 moves the arm 16 horizontally, and the AC supporter 7 6 moves the arm 33 horizontally. Each mechanism of the nucleic acid purification apparatus operates according to a predetermined program.

The configuration of the nucleic acid capturing chip 31 will be described with reference to FIG.

The nucleic acid capturing chip 31 has an inner diameter such that the head 54 is airtightly fitted to the tip of the dispensing nozzle 36. In the lower part, the inside diameter gradually increases toward the tip 48. An opening for sucking and discharging the sample exists at the tip of the tip portion 48. The inclination toward the front end portion 48 is substantially the same as the angle of the bottom surface of the cleaning tank 103. For this reason, by pressing the nucleic acid capturing chip 31 under the washing tank 103, the washing solution can be prevented from flowing out to the waste liquid hole 104.

The chip is made of a transparent or translucent synthetic resin. A disc-shaped blocking member 40b for preventing the solid phase from flowing out is inserted into the tip end side by press-fitting, and a disc-shaped blocking member 40a is inserted on the head 54 side. Is provided. These blocking members 40a and 4Ob have a large number of holes through which liquids and gases can easily pass, and the holes are large enough to prevent the outflow of the solid phase. Polypropylene is used as the material of the blocking members 40a and 4Ob. Since this material can reduce non-specific adsorption of proteins, nucleic acids, and the like, the influence on the purity and yield of nucleic acids is small. The blocking member 40a has a plurality of projecting insertion guides 37 on the lower surface side for facilitating insertion into the chip 31. The room sandwiched between the blocking members 40a and 40b is filled with flint glass (Wako Pure Chemical Industries) powder 44 as a solid phase. This print glass has a high silica content having a nucleic acid capturing effect.

In the nucleic acid capture chip 31, the cross section of the internal space containing the solid phase is preferably larger than the opening for aspirating and discharging the sample. This is because the dead volume (sample that does not touch the solid phase) during suction and discharge can be reduced. It is desirable that the tip for suction and discharge is thin. This is because the sample remaining at the bottom of the conical sample container can be suctioned and discharged. It is desirable that the thickness of the opening for sucking and discharging the sample be thin. This is because the sample liquid droplets and the cleaning liquid droplets adhering to the vicinity of the opening are reduced, and the generation of contaminants can be reduced. In addition, the solid phase has a thin flat plate shape that has a thickness in the chip major axis direction. For example, a disk shape is desirable. This is because clogging of the solid phase can be reduced and the volume of the solid phase can be increased. For this reason, the tip of the nucleic acid-capturing chip 31 is shaped like a combination of a large column containing a solid phase and a small column with an opening. Its longitudinal cross section is substantially convex and has an opening at the tip. For this reason, the substantially convex valley may be contaminated by the sample due to the suction and discharge of the sample from the opening.

Next, the operation of purifying nucleic acids will be described with reference to FIG.

Before starting the operation of purifying a nucleic acid-containing sample, a sample from which nucleic acid is to be extracted is put into a sample container 13, held by a sample rack, and set in a sample area on the apparatus 100. Tip rack 14 a with dispensing tip 15, tip rack 30 with nucleic acid capturing tip 31, tip rack 91 with reagent dispensing tip 94, each dispenser reagent pot 19 a, 19 b, 19 c, 19 d, 19 e (not shown), processing containers 24 a, 24 b, and containers 26 a, 26, 26 c for purified products After setting in the place, operation by the refining device 100 is started.

First, the first reagent (degrading enzyme solution, specifically ProK) is injected into the reaction vessel. Specifically, the nozzle holder 3 4 is operated, the reagent discharge nozzle 3 9 moves to the chip rack ₉ 1 above, fitting the 1st reagent dispensing tip 9 4 reagent discharge nozzle 3 9 . Next, the attached reagent dispensing tip 94 is moved onto a reagent bottle 92 containing a reagent such as the first reagent and lowered into the bottle. Next, a predetermined amount of the first reagent is sucked into the dispensing tip 15 by causing the syringe pump 32 to perform a suction operation. Next, after moving onto the container 24a, the reagent sucked into the dispensing tip 15 is discharged into the container 24a. Then, the reagent dispensing tip 94 is moved to the tip removing device 27b, and the used reagent dispensing tip 94 is removed. Next, the first reagent and the nucleic acid-containing sample are mixed in a reaction vessel. This elutes the nucleic acid. Specifically, the nozzle holder 17 is operated to move the dispensing and stirring nozzle 36 onto the tip rack 14a, and the first dispensing tip 15 is fitted to the dispensing and stirring nozzle 36. Combine. Move the dispensing and stirring nozzle 36 to the first sample container 13 on the sample rack 12 and lower the dispensing tip 15 into the sample container. A predetermined amount of the sample is sucked into the dispensing tip 15 by the suction operation of the syringe pump 10. Move the dispensing tip 15 from which the sample was inhaled to the first processing container 24 a on the container rack 23 a and move the dispensing tip 15 into the processing container 24 a. Dispense the entire volume. After the discharge, the whole amount of the discharged liquid is again sucked into the same dispensing tip 15 and further discharged to the first processing container 24a one or more times. Thereby, the first reagent and the sample are mixed.

Next, the second reagent (the dissolving reagent, specifically, guanidine hydrochloride solution) is mixed with the mixed sample in the reaction vessel. This elutes the nucleic acids. Specifically, the arm 16 and the nozzle holder 17 are operated to move the dispensing tip 15 to the position of the tip remover 27a, which is the standby position. Next, the arm 33 and the nozzle holder 34 are operated, and the reagent discharge nozzle 39 is moved to the container 24a from which the aforementioned sample is discharged. Next, the syringe pump 32 is operated to discharge a prescribed amount of the second reagent as the dispenser reagent. Then, the arm 33 and the nozzle holder 34 are operated again, and the arm 33 and the nozzle holder 34 are moved to the position of the chip remover 27 b which is the standby position. Next, the waiting dispensing tip 15 is again moved to the first processing container 24a on the container rack 23a, and all the samples in the processing container 24a are separated. Suction into the injection tip 15 and discharge to the first processing container 24a one or more times. This mixes the nucleic acid-containing sample with the second reagent. Then, remove the dispensing nozzle 36 from the tip Move to 2 7a and remove the used dispensing tip 15 from the dispensing nozzle 36 according to the removal operation described above. Next, the dispensing nozzle 36 is returned to the position of the cleaning section 18, and after a predetermined amount of pure water is discharged from the dispensing nozzle 36, a small amount of air is sucked into the tip of the dispensing nozzle 36.

Next, the third reagent (binding promoter, principally diethylene glycol dimethyl ether) is mixed with the mixed sample in the reaction vessel. Thereby, the eluted nucleic acid is changed to a state of binding to the nucleic acid capturing carrier. Specifically, the arm 33 and the nozzle holder 34 are operated, and the reagent discharge nozzle 39 is moved to the container 24a from which the above-described sample is discharged. Next, a predetermined amount of the third reagent, which is the dispenser reagent, is discharged. Next, the arm 33 and the nozzle holder 3-4 are operated again, and moved to the position of the tip remover 27 b which is the standby position. Next, the waiting dispensing tip 15 is moved onto the first processing vessel 24a on the container rack 23a again, and all the samples in the processing vessel 24a are dispensed. Suction into the chip 15 and discharge to the first processing container 24a are performed at least once. This mixes the nucleic acid-containing sample with the third reagent. Next, the reagent dispensing tip 94 is moved to the chip remover 27 b, and the used reagent dispensing tip 94 is removed.

Next, the nucleic acid is captured on the solid phase in the nucleic acid capture chip. Specifically, the nozzle holder 13 is operated to move the dispensing and stirring nozzle 36 onto the tip rack 30, and the first nucleic acid capturing chip is fitted to the dispensing and stirring nozzle 36. Thereafter, the dispensing nozzle 36 moves to the position of the first processing container 24a on the container rack 23a with the nucleic acid capturing chip 31 coupled thereto. The nucleic acid capture chip 31 is lowered, and the entire amount of the mixture of the sample and the first, second, and third reagents contained in the first processing container is aspirated by the syringe pump 10 into the chip. It is sucked into the nucleic acid capturing chip 31 by operation. to this The mixed solution comes into contact with the surface of the glass powder 44 as a solid phase in the chip 31. Next, the sucked mixture is discharged back into the first processing container 24, and the discharged mixture is again sucked into the same nucleic acid capturing chip 31. By repeating the discharge and inhalation of the mixed solution a plurality of times, the number of contacts between the solid phase surface and the mixed solution is increased, and the efficiency of nucleic acid adsorption by the solid phase is increased. After the predetermined number of times of suction and discharge, the entire amount of the mixed solution is finally sucked into the first nucleic acid capturing chip 31, and the chip 31 is moved to the washing unit 29.

Next, the nucleic acid capturing chip 31 is washed. Hereinafter, the cleaning method will be described with reference to FIG. 2 and FIG. In FIG. 2, a and b indicate cleaning inside the chip, c indicates cleaning outside the chip, d indicates cleaning at the tip of the chip, and e and f indicate liquid droplet removal. FIG. 9 is a flowchart showing a cleaning process. The inside of the chip is washed twice using the fourth reagent (first washing reagent, the main components of which are guanidine hydrochloride and diethylene glycol dimethyl ether) and the fifth reagent (second washing reagent, the main component of which is ethanol). The fourth reagent removes mainly proteins, and the fifth reagent removes impurities other than proteins that cannot be washed with the fourth reagent. In other words, after performing steps a and b in FIG. 2 twice by changing the cleaning solution, the steps after c are performed.

First, the sample in the nucleic acid capturing chip 31 is discharged, and then the nucleic acid capturing chip 31 is filled with the fourth reagent (a). Specifically, first, the nucleic acid capturing chip 31 is inserted into the washing tank 103. At this time, the distal end portion 48 of the nucleic acid capturing chip 31 is inserted into the waste liquid hole 104. Further, the nucleic acid capturing chip 31 is fixed in close contact with the bottom of the washing tank 103. As a result, the washing tank 103 and the waste liquid hole 104 are separated.

Next, the sample accumulated in the chip after the nucleic acid adsorption step is discharged into the washing section 29 by the discharging operation of the syringe pump 10. Next, the flow path switching valve 83 is driven to connect the washing liquid flow path to the nucleic acid capturing chip 31. At the same time, the air switching valve 82 is driven to physically connect and open the inside and outside of the chip. Thus, the pressure inside the nucleic acid capturing chip 31 is always equal to the pressure outside the chip.

Next, the syringe pump 80 is driven to inject the fourth reagent into the chip. Since the pressure in the chip does not increase, the washing solution discharged by the syringe pump 80 is not discharged from the opening at the tip of the nucleic acid capturing chip 31 and accumulates in the chip. At this time, the level of the cleaning liquid in the chip is higher than the position of the solid phase, and the solid phase is immersed in the cleaning liquid.

Then, the air switching valve 82 is driven to shut off the flow path connecting the inside of the chip and the outside of the chip.

Next, the fourth reagent held on the nucleic acid capture sample chip is discharged (b). Specifically, the following procedure is performed.

First, the air discharge pump 87 is driven to increase the pressure in the chip. The cleaning liquid in the chip flows in one direction from the head (upper) to the tip (lower), passes through the solid phase, and is discharged from the tip. At this time, the sample, cleaning liquid, and the like are scattered into the waste liquid hole 104 from the tip of the chip by air blow discharge. However, since the chip and the bottom of the washing tank 103 are in close contact with each other, droplets do not scatter above the washing tank 103, and no contamination occurs for other samples. As a result, the inner wall of the chip and the surface of the solid phase can be washed, and all the residual liquid can be discharged from the nucleic acid capturing chip 31.

The air blow discharge not only discharges the sample and the cleaning liquid, but also promotes the drying of the carrier for trapping the nucleic acid. Due to the synergistic effect with the liquid discharge by wind pressure, the liquid in the chip can be efficiently removed.

When the viscosity of the reagent is high, first activate the air switching valve 82 to capture nucleic acids. The syringe pump 10 is connected to the atmosphere via the tip 31. In this state, the syringe pump 10 is driven to suck air. Then, the air switching valve 82 is driven again to shut off the air. Then, the syringe pump 10 is driven to push out the remaining reagent in the chip. By this repetition, the residual liquid adhering to the inner wall of the chip, the solid phase, or the like can be reliably discarded.

The second washing step is performed in the same manner as the first washing, using the fifth reagent (second washing solution). Thereby, impurities other than the protein that cannot be washed with the fourth reagent can be removed.

Next, the outside of the nucleic acid capture sample chip 31 is washed (c). Since the fourth reagent remaining outside the chip affects the subsequent elution step, it must be sufficiently washed in this step. Specifically, the fifth reagent (second cleaning liquid) is injected into the water storage tank 102 from the cleaning liquid injection port 101 in a state where the chip and the bottom surface of the cleaning tank 103 are in close contact with each other. Since this cleaning solution is used only for cleaning the outside of the chip, it is not suitable for cleaning the inside of the chip, but inexpensive sterilized water or the like can be used.

The injected cleaning liquid flows into the cleaning tank 103 through the communication channel 106 connected to the cleaning tank 103. Since the communication channel 106 has a smaller cross-sectional area than the cleaning liquid inlet 101, the water storage tank 102 stores a fixed amount of cleaning liquid. The washing solution that has flowed into the washing tank 103 accumulates in the washing tank 103 because the waste liquid hole 104 is closed by the nucleic acid capturing chip 31. As a result, the outer wall of the chip excluding the tip end portion 48, that is, a portion with a low degree of contamination can be cleaned. At this stage, the tip end portion 48 with a high degree of contamination does not come into contact with the cleaning solution, so that the contamination at the tip end portion 48 does not diffuse.

The washing water accumulates up to the position of the bypass channel 105, and the washing water exceeding that position is discharged to the waste liquid hole 104 via the bypass channel 105. Pa If the position of the bypass path 105 is higher than the expected contamination site, contamination outside the chip can be reliably removed.

Next, the tip 48 of the nucleic acid capturing chip 31 is washed (d). Intensive cleaning of the parts not cleaned in the previous process. Specifically, the nucleic acid capturing chip 31 that is in contact with the washing tank 103 is moved upward to provide a predetermined gap. The upward movement amount is, for example, about 0.5 mm. The cleaning liquid used for external cleaning of the chip is discharged to the waste liquid hole 104 from this gap. Since the gap is small, the washing water jumps out of the chip and flows down to the waste liquid hole 104 along the shape of the chip. The manner in which the washing water jumps out can be adjusted depending on the amount of accumulated washing water and the shape of the gap. Thereby, the distal end portion 48 of the nucleic acid capturing chip 31 can be washed. In particular, it is possible to reliably clean a place where cleaning is difficult, such as a step portion having a substantially convex shape. Even if the cross section used in the present embodiment is other than the chip having a substantially convex shape, the cleaning can be surely performed by adjusting the way of the washing water popping out.

In addition, a part of the cleaning tank 103 in contact with the chip and the inside of the waste liquid hole 104 can be cleaned. As a result, droplets and the like that have been scattered in the waste liquid holes 104 by air blowing can also be removed. Therefore, it is possible to avoid contamination due to the splash of the air propellant outside the waste liquid hole.

Next, the liquid ball adhering to the tip end portion 48 is removed (e and f). Thereby, scattering at the time of tip movement can be prevented. Specifically, the cleaning liquid is injected again and the cleaning water is stored in the water tank 102. In order to maintain the water level in the water storage tank 102, the drive of the water injection pump is continued. Then, with the water level maintained, the nucleic acid capturing chip 31 is inserted into the water storage tank 102, and the tip end portion 48 is brought into contact with the washing liquid surface (e).

Next, the nucleic acid capturing chip 31 is moved upward (f). The moving speed is A relatively slow one is desirable. For example, the speed is about 1-2 mm per second. As a result, the liquid droplets attached to the tip of the nucleic acid capturing chip 31 are captured on the cleaning liquid surface by surface tension, so that the liquid droplets can be efficiently removed. In the nucleic acid extraction procedure, a trace amount of reagent is used. In particular, in the subsequent extraction step, nucleic acids are extracted using a very small amount of extract. For this reason, even if the washing liquid is contaminated, the volume of the reaction solution is disturbed and the extraction accuracy is deteriorated. In addition, the liquid droplets attached to the tip end portion 48 are scattered by the movement of the nucleic acid capturing chip 31 and cause contamination. However, the above-described contamination can be avoided by the liquid ball process. It should be noted that the same effect can be obtained by fixing the height of the nucleic acid capturing chip 31 and raising and lowering the liquid level by the cleaning liquid injection pump. That is, the nucleic acid capturing chip 31 and the washing liquid surface may be relatively moved and brought into contact.

Further, the cleaning liquid injected from the cleaning liquid inlet 101 is temporarily stored in the water tank 102, and then discharged to the waste liquid hole 104 through the cleaning tank 103. Thereby, the water storage tank 102, the cleaning tank 103, and the waste liquid hole 104 can be washed. Next, the nucleic acid captured on the solid phase is eluted, and the nucleic acid is collected in a storage container. Specifically, the washed nucleic acid capturing chip 31 is moved to the liquid receiving section 28 and waits. In the nucleic acid elution step, the reagent nozzle 39 moves to the position of the reaction container 24b. The syringe pump 32 is operated to aspirate the sixth reagent (eluent, the main component is Tris Buffer), which is a dispenser reagent, and a single amount of the eluent is discharged by the syringe pump 32 being pushed out. Discharge into the first processing container 24b. The reagent dispensing nozzle 39 holding the remaining amount of the eluent moves to the chip removing portion 27 b and stands by. Next, the nucleic acid capturing chip 31 is moved to the first processing container 24 b on the container rack 23 b, and the eluate in the first processing container 24 b is captured. Into the tip 31 for use. As a result, the eluent comes into contact with the solid phase, and the nucleic acid adsorbed on the solid phase surface is eluted into the eluent. After discharging the eluate sucked into the chip 31 into the original processing container 24b, the operation of sucking again into the same chip 31 is repeated a predetermined number of times. Then, the reagent dispensing nozzle 39 moves to the second processing container 24b, and discharges the next one eluent into the corresponding processing container. Subsequently, the nucleic acid capturing chip 31 is moved to the second processing container 24b, the eluate in the processing container 24b is sucked into the nucleic acid capturing chip 31, and the same nucleic acid as described above is used. Perform the elution procedure. The elution step is repeated to elute the nucleic acid adsorbed on the nucleic acid capturing carrier. After the above steps are completed, the nucleic acid capturing chip 31 is moved to the position of the chip remover 27a, and the chip is removed.

Next, the nozzle holder 17 is driven to move the dispensing and stirring nozzle 36 onto the tip rack 14 b, and the dispensing tip 15 is fitted to the dispensing and stirring nozzle 36. Then, it is moved to the first row of the reaction vessel 24b containing the eluate recovered in the previous step. Next, the eluate in the reaction vessel is aspirated into the dispensing tip 15. Next, it is moved to the position of the container 26 for the purified tip dispensing product. By the pushing operation of the syringe pump 32, the eluent contained in the dispensing tip 15 is discharged into the first purified product container 26. As a result, the eluate containing the nucleic acid eluted from the solid phase is collected in the purified product container 26. Subsequently, the dispensing tip 15 is moved to the second processing vessel 24b, the eluate in the processing vessel 24b is sucked into the dispensing tip 15 and the same as described above. After performing the nucleic acid elution operation, the eluate containing the nucleic acid is collected in the first purified product container 26. After dispensing the eluent, the dispensing tip 15 a is moved to the chip remover 27 a, and the used dispensing tip 15 is removed from the dispensing nozzle 36. The dispensing nozzle 36 with the dispensing tip removed is 29 Move to 9 and discharge water from the nozzle tip, then inhale a small amount of air into the nozzle tip and wait at that position. This completes the procedure for purifying the nucleic acid for the first sample. Thereafter, the purifying apparatus 100 continues the nucleic acid purification operation on the second and subsequent samples, and the operation is a repetition of the above-described example.

According to the present embodiment, the up-and-down movement of the nucleic acid capturing chip 31 and the control of the injection of the washing solution can prevent the sample and the reagent from scattering, and can reliably wash the chip. As a result, contamination can be prevented, and an apparatus for purifying nucleic acid with high washing efficiency can be provided. In addition, the device configuration is simple, and cost reduction can be achieved.

Although the present embodiment relates to a nucleic acid purifying apparatus, the present invention is not limited to this. For example, it can be applied to an automatic analyzer using a chip, such as an automatic biochemical analyzer, an automatic immune analyzer, and an automatic urine analyzer. Industrial applicability

It is possible to provide an automatic analyzer and a washing method capable of preventing the occurrence of contamination and efficiently washing a sample container.

Claims

The scope of the claims

1. An automatic nucleic acid extraction apparatus,

A nucleic acid-capturing chip that contains a nucleic acid-capturing substance and aspirates and z- or discharges a solution containing nucleic acids

A moving mechanism for moving the nucleic acid capturing chip,

A washing container,

A washing solution injection mechanism for injecting a chip washing solution for washing the nucleic acid capture chip into the washing container,

The cleaning container has a waste liquid hole for discharging the chip cleaning liquid,

The waste liquid hole is closed by the nucleic acid capturing chip.

2. The automatic nucleic acid analyzer according to claim 1, wherein

The chip washing solution is stored in a washing container in which the waste liquid hole is closed by the nucleic acid capturing chip, and the nucleic acid-containing solution is discharged from the nucleic acid capturing chip.

3. The automatic nucleic acid analyzer according to claim 1, wherein

The chip washing liquid is collected in the washing container in which the waste liquid hole is closed by the nucleic acid capturing chip, and thereafter, the nucleic acid capturing chip is separated from the waste liquid hole, and the collected chip cleaning liquid is flown into the waste liquid hole to wash the nucleic acid capturing chip.

4. The automatic nucleic acid analyzer according to claim 3, wherein

After washing the nucleic acid capture chip, bring the tip of the nucleic acid capture chip into contact with the cleaning solution, and then release.

5. An automatic analyzer including the following components;

A cleaning liquid supply mechanism for supplying a cleaning liquid for cleaning a sample container having an opening for sucking and / or discharging a sample;

A cleaning container having a discharge hole for discharging a cleaning liquid;

Move the sample container and the or the washing container, The sample container and the washing container are brought into contact with each other to form a liquid storage space that can hold the washing solution.

A moving mechanism that provides a predetermined gap between the sample container and the cleaning container and allows a predetermined amount of cleaning liquid held in the liquid storage space to flow to the discharge hole.

6. The automatic analyzer according to claim 5, wherein

The opening has a convex shape with an opening at the tip.

7. The automatic analyzer according to claim 6, wherein

The sample container is a nucleic acid capturing chip containing a nucleic acid capturing substance.

8. The automatic analyzer according to claim 6, wherein

The cleaning container has a liquid level positioning hole for discharging the cleaning liquid injected into the liquid storage space.

9. The automatic analyzer according to claim 5, wherein

Having a pump for controlling the pressure in the sample container,

The pump discharges the sample from the opening when the sample container and the cleaning container are in contact with each other.

10. The automatic analyzer according to claim 5, wherein

The moving mechanism causes a predetermined amount of the cleaning liquid held in the liquid storage space to flow to the discharge hole, and then brings the opening of the sample container into contact with the cleaning liquid.

1 1. Automatic analyzer including the following components;

A cleaning liquid supply mechanism for supplying a cleaning liquid for cleaning a sample container having an opening for discharging a sample;

A cleaning container having a discharge hole for discharging a cleaning liquid;

Move the sample container and / or wash container,

The sample container and the washing container are brought into contact with each other to form a liquid storage space that can hold the washing solution. A moving mechanism that separates the sample container from the washing container and allows the washing liquid held in the liquid storage space to flow to the discharge hole;

A pump that controls the pressure in the sample container and discharges the sample from the opening to the discharge hole when the sample container is in contact with the washing container.

1 2. The automatic analyzer according to claim 1, wherein

A sample container is provided with a fluid resistance that prevents sample discharge.

13. The automatic analyzer according to claim 11, wherein

The sample container includes a nucleic acid-adsorbing substance.

14. The automatic analyzer according to claim 11, wherein

The sample container comprises quartz wool.

15. The automatic analyzer according to claim 11, wherein

The discharge hole has a hole shape into which the opening can be inserted.

16. The automatic analyzer according to claim 11, wherein

17. The automatic analyzer according to claim 11, wherein

The moving mechanism causes the opening of the sample container to come into contact with the cleaning liquid after flowing the cleaning liquid held in the liquid storage space to the discharge hole.

18 8. A method for cleaning a sample container including an opening for aspirating and / or discharging a sample, including the following steps;

Close the discharge hole of the washing container with the sample container, inject the cleaning solution into the washing container, wash the surface of the sample container not including the opening, separate the sample container from the discharge hole, and remove a predetermined amount of Rinse the washing solution into the discharge hole to wash the surface of the sample container including the opening.

1 9. The cleaning method according to claim 18, wherein A cleaning method in which the opening has a convex shape having an opening at the tip.

20. The cleaning method according to claim 18, wherein

21. The cleaning method according to claim 18, wherein

With the discharge hole of the washing container closed with the sample container, the sample in the sample container is transferred from the opening to the discharge hole.

22. The cleaning method according to claim 18, wherein the method includes the steps of: contacting the opening with a cleaning solution, separating the opening, and then removing the cleaning solution attached to the vicinity of the opening.

23. A method for cleaning a sample container including an opening for discharging a sample, including the following steps;

Close the discharge hole of the washing container with the sample container, transfer the sample in the sample container into the discharge hole, inject the cleaning liquid into the cleaning container, separate the sample container and the discharge hole, and discharge the cleaning liquid from the provided gap. Pour into holes.

24. The cleaning method according to claim 23, wherein

25. The method according to claim 23, wherein

The sample container includes a nucleic acid-adsorbing substance.

26. The method of claim 23, wherein

The sample container comprises quartz wool.

27. The method of claim 23, wherein

The discharge hole has a hole shape into which the opening can be inserted.

28. The method of claim 23, comprising the following steps:

The opening is brought into contact with the cleaning liquid and then released, reducing the amount of cleaning liquid attached near the opening.