JPS59147268A - Method and device for automatically moving liquid sample among plurality of vessel - Google Patents

Abstract

(57) [Summary] This bulletin contains application data before electronic filing, so abstract data is not recorded.

Description

DETAILED DESCRIPTION OF THE INVENTION Field of the Invention The present invention relates to a method for automatically transferring a liquid sample between a plurality of containers. More specifically, a separate plurality of microtitration trays, each containing between 1/10 and 1,1 liters of liquid, requires an initial filling and successive dilutions of the liquid sample. This invention relates to a system for filling or transferring a liquid sample between liquid containers. This serial dilution system essentially involves mixing the sample in separate containers with successively increasing proportions of diluent, whereby the concentration decreases continuously or serially. A series of samples are obtained. Sansol can therefore be tested at various concentrations to determine specific properties. For example, the sample may be serum, and tests may be performed to determine what concentration of serum will give the best results when acting on a particular substance.

PRIOR ART Initially, serial dilution of the sample was carried out manually, with different proportions of the sample being contained in different test tubes with the help of e.g. syringes or pipettes. mixed with. This operation can be quite time consuming when a large number of different concentrations are required; automatic or semi-automatic serial dilution methods have therefore been developed. An example of a machine for carrying out this type of process is disclosed in U.S. Pat. No. 3,188,181. The serial diluter disclosed in this patent has a horizontally movable carriage that accommodates tube hooks for multiple test tubes. A vertically movable syringe holder housing a plurality of shelfless syringes, each coupled to a plurality of pipettes, includes a cam for transporting each pipette into a liquid volume within a row of test tubes within said tube engagement. It is moved up and down using. A cam-actuated pumping head vibrates the syringe to mix the fluid in the pipette with the fluid in the test tube. After mixing, the fluid is removed from the test tubes, the syringe holder is lifted up and the carriage is moved to another row of test tubes to repeat the operation. Once the successive dilutions are complete,
The first row of test tubes in the tube stack will contain the undiluted concentration of the sample, and depending on the amount of liquid delivered by the pipette, the second row of test tubes will contain the undiluted concentration of the sample. % a degree of liquid, the third column would contain liquid at 25% concentration of the sample, and so on.

<Objective of the Invention> The object of the present invention is to effectively transfer liquid between a plurality of containers without cross contamination and with no error in the amount of liquid transferred, and which is possible with the conventionally known machine of the above patent. Another object of the present invention is to provide a new system that is particularly useful for performing continuous or serial dilutions of small volume liquid samples with substantially improved performance. More specifically, when dealing with certain types of solutions, it is desirable to completely prevent cross-contamination between various samples having different concentrations. Typically, this is done in a manual manner by using a disposable tip on a pipette to remove the liquid from one container and mix this liquid with the diluent in the other container. After each mixing operation, the tip is removed from the pipette and replaced with a clean tip. However, hitherto known continuous diluters do not provide the ability to replace each chiller Q between each cycle in successive dilution steps. As a result, when it is desired to prevent cross-contamination, the machine must either incorporate multiple stubbed cleaning steps for multiple pipettes, or provide seven lashing steps, i.e., rapid cleaning steps, between each cycle of the process. No. The washing process substantially increases the time required to perform serial dilutions since multiple steps are added to each cycle of the process. On the other hand, the flushing process generates large amounts of contaminated liquid that must be disposed of.

It is therefore a further particular object of the present invention to provide a method for controlling the Stellar ff:
It is an object of the present invention to provide a novel automatic diluter that can quickly and effectively automatically replace a disposable chia ff via the method.

Arrangements of the Invention The above objects of the invention are achieved by the invention by providing at least two working stations on a horizontally moving table below a plurality of pipettes for withdrawing and injecting liquid samples. Ru. In a preferred embodiment, at least one work station has a liquid receiving tray or similar structure having multiple rows of containers containing liquid samples and diluents, and at least a second work station The pipette has a hook section that accommodates multiple rows of disposable pipette tips. The pipette itself has a ring that can be inserted into the tip (C) and frictionally engaged when the head part on which the pipette rests is moved downwards to hold the tip together. - has an end provided with a groove. Once the chips have been picked up, the table is moved to carry the selected reservoir row in the liquid receiving tray below the chips, and then picks up the chips. Lowered for insertion into the reservoir.

A portion of the sample in the plurality of reservoirs is drawn into the plurality of pipettes by actuation of a plunger in each pipette. After lifting the head, the table is then given one or more steps to bring the generally next successive row of reservoirs into the active area of the chip. The head is then lowered to insert the tip into the diluent in these reservoirs and the sylanger is vibrated to mix the sample with the diluent. After all the liquid has been drained from the tip, the head is raised so that the bitted tip is just above the meniscus of liquid in the reservoir. The sylanger is then actuated to expel all the liquid with the aid of some excess air from the pipette and surface tension on the liquid between the tip and the reservoir. The table is then moved to carry the latch containing a plurality of tips down the pipette, and a solenoid-controlled tip ejection means returns the tips from the tapered end of the pipette to the latch. It is activated to push. Alternatively, chips may be released via slots in the table (other stations on one ZH table) into a trough or collection container. The table is then added one step to bring a new set of tips into the working area of the pipette and the cycle is repeated. In a preferred embodiment, the volume of each tip is larger than the volume of liquid in the plurality of containers in the tray, so that all the liquid is contained in the containers without substantial contact of the liquid with the grunges of the pipette. When you put it in, you take it out.

According to another feature of the invention, by determining when a plurality of pipettes fails to pick up all pipettes in a row, thereby failing to draw sample into the pipettes due to loss of tips; A detector can be provided to prevent a situation in which the sample in one reservoir is not serially diluted into all the various concentrations.

Similarly, a detector may be provided to determine that all tips have been ejected from their respective pipettes prior to picking up the next row of tips f.

All movements of the various elements of the machine can be detected and controlled as desired by the computer, thereby providing continuous sensing of the process as well as filling and transporting liquid for continuous dilution processes. It can give you flexibility in what you do.

DESCRIPTION OF THE PREFERRED EMBODIMENTS The present invention will be described in detail below with reference to the accompanying drawings showing preferred embodiments of the present invention.

As shown in FIGS. 1 and 2, an automatic continuous diluter suitable for carrying out the method of the invention has two movable main parts: a horizontally movable table 10 and a vertically movable rad section 12. Contains. As clearly shown in FIG. 2, the table 10 is mounted on sliding bearings 1 for horizontal movement.
6 and placed on a plurality of sturdy guide rods 14.

Movement of the table is provided by a stepper motor 18 via a pinion 20 connected to the motor and a rack 22 mounted on the underside of the table.

Similarly, the head portion 12 is mounted on sliding bearings 26 for vertical movement.
It is attached to a plurality of guide rods 24 via. Movement of the head section is provided by step motor 28 via pinion 30 and rack 32. FIG. 10 shows a plan view of the rack and binion mechanism for moving the head section.

Head portion 12 supports pipette plunger assembly 34. The assembly 34 has a series of pipettes 36t arranged in rows transverse to the axis of movement of the table 10. These plurality of pipettes 36 are attached to a block 37.
It is removably attached to the head using a holder and moves with the head. A granger mechanism 38 is attached to the bottom section for vertical movement of the plurality of beds 36.

Plunger mechanism 38 includes a series of plunger rods 40, each one of which is disposed inside a corresponding pipette 36. All of the syringers are mounted on a common actuation bar 42 for coordinated vertical movement. The operating par 42 is moved along the guide rod 44 by a stepper motor 46 and a ball bearing three screw drive mechanism 48.

As best seen in the detailed cross-sectional view of FIG. 3, movement of the plunger rod 40 relative to the pipette 36 changes the internal volume of the pipette causing fluid to be drawn into the pipette and thus ejected from the pipette. An airtight seal is provided between each rod and the top of its corresponding pipette by using a grommet 47 and a Q-ring 49 held by a weak spring 45. Each pipette 36 is attached to the mounting block 3
It has a piston portion 39 mounted so as to be able to reciprocate within a cylinder 35 formed at 7. Pipette 36 is thereby restrained vertically by spring 45 and
As a result, during steps where the tip is under force, the pipette can slide vertically within the block 37 against the spring 45. This ensures that all pipettes pick up slightly different sized tips or mouthpieces 62 and that the open ends of the plurality of tips 62 are at the same height relative to table 10 and liquid receiving tray 54. enable.

Table 10 is equipped with two work stations 50 and 52, each accommodating two trays. One of the trays contains the liquid sample and diluent.
It can be a conventional liquid receiving tray 54 having a reservoir arranged in a J-sock shape. Rear work stage old 5
The other tray b6 in 2 can be a tip tray having similarly arranged containers containing disposable pipette tips. A typical liquid receiving tray has 96 reservoirs arranged in a 12×8 matrix. As shown in FIG.
Provided to perform 2x7 serial dilutions. In that case, the first row of the plurality of reservoirs is filled with a predetermined amount of Sunnol to be diluted, and the remaining p reservoirs are filled with compensating fluid. In this case, the tep tray 56 is oriented to provide twelve containers in rows across the width of the tray.

Separately, as shown in FIG.
It can also be oriented in the longitudinal direction.

As shown in the detailed side view of FIG. 3, the bottom end of each pipette 36 is adapted to receive and abutmentally engage the inner surface of a disposable pipette tip 62 constructed in accordance with the present invention. , its outer surface is tapered. For example, tip 62 may be made from a non-wettable polypropylene material. The tips 62 in a row of containers 36 in the tip tray 56 are removed from the pipettes when the head 12 is lowered by the stepper motor 28 after the table 10 has brought the row of tips 62 to the working position of the pipettes. 36 is inserted and engaged at each end. As already mentioned, each terra f62
is a substantial portion of the total volume of the cylinder formed by the body of the bed 36 plus the interior volume of the chip. As best seen in FIGS. 4 and 5, support for each chip 62 within the container 63 of the tray 56 is provided by end supports as shown in FIG. 4 or formed on the outside of the chip 62. Either by the edge of the bottom flute section 65 that has been cut. The wall of container 63 is Chiso F62
The tip 62 is centered to engage the beveled end 60 of the tip.

The operation of successively removing a plurality of tips 62 from a plurality of pipettes is performed by a tip removal means.

This tip removal means is illustrated in detail in FIG.
It has a comb-like grate 64. This comb-shaped plate 64
has a number of recesses for accommodating a plurality of pipettes, such that the toothed portions of the recesses surround a substantial portion of the outer periphery of each pipette body, e.g. . The comb plate 64 is connected to and supported by a pair of vertically movable rods 66 on the head portion 12 . These rods are moved using a pair of solenoids 68 mounted on the top of the throat. When solenoid 58 is not actuated, removal plate 64 remains in the upper position shown in FIG. When the solenoid is actuated, the removal grate 64 moves vertically downwardly and forces the plurality of tips 62 downwardly, releasing the tips 62 from frictional engagement with the end of the respective pipette 36. FIG. 9 shows the solenoid 6 for the stem motor 46 and the head section 12.
80 shows a plan view of the inventive device showing position 80'ilt;

Operation of each of step motors 18, 28 and 46 and solenoid 6B is controlled by a suitable microprocessor 70.
controlled by Essentially, the microprocessor 70 operates as a pulse generator that in turn controls the operation of each of the various elements described above, and thus operates as a pulse generator for the table 10, head 12, granger assembly 34, and removal grate 64. The interrelated movements result in successive dilutions of the sample located in the tray 54 in the forward working position 50. Since the plurality of stepper motors impart a predetermined amount of rotation in response to each starting pulse supplied to it, the precise positioning of each movable element depends on the appropriate number of starting pulses supplied by the microprocessor. can be obtained through control.

In addition to controlling these multiple movable elements, microprocessor 70 also monitors the movement of the multiple elements via appropriately positioned sensors. For example, a monitoring device for the table 10 detects when a blade member 72 disposed on and extending from the side surface of the table and the blade member 72, and therefore the table 10, pass a predetermined point during its movement. The halo effect sensor 74 is configured to include a halo effect sensor 74. Every time the table passes the specified point, no.
Is the effect sensor 74 a signal? Send to microprocessor 70
), microgrosset? 70 can update information regarding the location of the table. Thus, if the step motor 18 loses the start signal υ while the table is moving, or if the number of nozzles stored in the microprocessor 70 does not match the position of the table, an error occurs. In addition to the sensor 74, a pair of limit sensors 76 can be placed at each end of the table's travel trajectory. The signals sent by these sensors indicate that the table is nearing the end of its travel and instruct it to interrupt the power supply to the computer or take other similar appropriate actions. is given to the microprocessor 70. A similar sensor device is part 1.
2 and the plunger bar 42.

Additionally, other sensors may be provided on the device to detect whether all tips in a row of trays 56 have been picked up by the pipette assembly. As shown in FIG.
9) or a similar light emitting device on one side of the table and a power point electronics 90 on the opposite side of the table. The two elements are placed in the column of pipette 36 -@
Aligned to the line. When one or more steps are within the array of sensor-monitored reservoirs 630, the light flux 82 from the LED is blocked and does not reach the optoelectronic element 90.

However, if all chips in the column are picked up sequentially by the pipette assembly, the beam of light will extend across the pan and be detected by optoelectronic element 90. By arranging the tray 56, a possible pick-up of the tray 56 due to friction between the chips 7'62 in the tray 56 and the reservoirs can also be detected.

12, 13 and 14 show other embodiments of the movable table moving device according to the invention. In this embodiment, another microtitration tray 88 is provided between the liquid receiving tray 54 and the chip tray 56. Tray 88 contains either a supply of biological material or a reagent that initially fills the receiving tray container. Although shown as a plurality of individual reservoirs, tray 88 could be a common feed trough or steel member. For example, a first charge of sample material is poured into a first row of containers in tray 54. Then, after replacing a plurality of chips 62 from tray 56, the reservoir remaining in tray 56 is filled with diluent transferred from another portion of tray 88 or with another supply liquid from another tray. As previously mentioned, after use, the vivette chizzo may be discharged into an empty container within the chip tray 56. However, table 1
It is also conceivable that other parts of 0 allow collection of all used chips. The table 10 has slots (
(not shown) or by placing the collection bottle in another position on the table.

In operating the device according to the invention, the automatic continuous diluter operates basically as follows.

First pick up a sample of chips in the tip tray 56, insert them into a row of reservoirs in the liquid receiving tray 54, extract liquid samples from these reservoirs, and dilute the next successive row of reservoirs. Introduce the chip into the liquid, vibrate the plunger to mix the liquid, and move the chip to tray 56 to expel all liquid and return the chip. This operation will be explained in detail below with reference to the example below of a program that can be used by a microprocessor to effectively perform a serial dilution process.

002 Move the head downward and grasp the chip 003
Move the head upwards Pick up the chip 006 Move the head downwards Insert the chip into the reservoir 008 Move the head upwards Remove the chip from the reservoir 010 Move the head downwards Insert the chip into the reservoir 011 Vibrate the plunger Insert the chip into the reservoir Mix the diluted solution013 Lower the plunger Discharge the liquid from the pet017
Move the plunger to the initial position 019 Move the head downward Insert the tip into the container 02
0 Tip remover downward Remove the tip 021 Tip remover upward 023 Head upward 024 M=M+1. N=N+1 025 Table to position M 026 To step 002 The cycle is repeated a number of times equal to the number of dilutions performed. If tip replacement is not required,
Steps 001-004 and steps 018-022 are omitted during the cycle.

Prior to initiating a serial dilution operation, the amount of liquid to be transferred during each cycle of the process is programmed into the microprocessor 70. This amount determines the extent to which the plunger rod 40 is raised during step 007 of the program. In turn, this operation determines the concentration of sample in successive reservoirs of tray 54. For example, to obtain a dilution range such that the concentration of column 1') is within the range of the concentration of the preceding column, the first column of the reservoir is filled with 100 microliters of sample, and the other columns are filled with 100 microliters of sample. Each reservoir is preferably filled with 50 microliters of diluent.

The microprocessor is configured to transfer 50 microliters from one reservoir to an adjacent reservoir during each cycle.

In Stella f011, the plunger rod 40 can be oscillated up and down approximately five times to ensure proper mixing.

At the beginning of each cycle of successive dilution steps, plunger rod 40 is placed at a predetermined calibration point within the pipette. A Hall effect similar to the type described above with respect to Table 10 can be used to monitor and control the position of the rod. In step 014 of the program, after the sample and diluent have been mixed in step 011, the plunger tip is moved to the calibration point to eject the liquid from the pipette at the height of the liquid in the reservoir in step 012 and at stella 7°013. The plunger tip is raised so that it is just above the returned plunger,
The tip is raised to a point just above the meniscus of the liquid in the container. All liquid is then expelled from the pipette by extending the plunger downwardly past the calibration point. This action effectively blows the liquid out of the pipette by also forcing out some of the air trapped inside the pipette, and into the tip due to capillary action due to surface tension acting on the liquid. This allows any liquid remaining and extending between the tip and the container to be drawn out of the tip. This step is particularly effective when the chip is made from non-wettable plastic, which is the preferred material as mentioned above.

Although certain steps are shown separately, these steps can be performed simultaneously. For example, step 016 and step o17 are performed simultaneously.

It will be apparent to those skilled in the art of foiling that the invention may be embodied in other specific forms without departing from its spirit or essential characteristics. Accordingly, all embodiments disclosed in this specification are to be considered illustrative and not restrictive. For example, although stepper motors are described as the preferred embodiment of the motor drive means for the table, head, and syringer mechanism, other precision positioning means may be used, such as a DC servo motor. Therefore, the scope of the present invention is determined by the claims rather than the foregoing description, and all changes within the range equivalent to the claims are to be included within the scope of the present invention.

[Brief explanation of the drawing]

FIG. 1 is a perspective view of a continuous dilution device incorporating features of the present invention. FIG. 2 is a side cross-sectional view of the continuous diluter taken along line 2--2 of FIG. FIG. 3 is an enlarged side sectional view of one embodiment of the pipette tip as it resides within the tip tray reservoir. FIG. 4 is an enlarged side cross-sectional view of the preferred shape of the pipette tip supported within the tip tray well. FIG. 5 is an enlarged side sectional view of the granger and pipette assembly. FIG. 6 is a plan view of a table on which two trays are placed for performing 12×7 dilutions. FIG. 7 is a plan view of the Cheagle with two trays arranged for 8×11 dilutions. FIG. 8 is a front cross-sectional view of the automatic continuous diluter taken along line 8--8 of FIG. FIG. 9 is a partial plan view of the device of the invention. FIG. 10 is a cross-sectional plan view taken along line 10--10 of FIG. FIG. 11 is a cross-sectional plan view taken along line 11--11 in FIG. FIG. 12 is a perspective view of an alternative embodiment of a continuous diluter that includes a fluid transfer or supply tray between a chip tray and a microfluidic receiving tray. FIG. 13 is a plan view of the embodiment shown in FIG. 12. FIG. 14 is a side sectional view taken along line 14--14 of FIG. 13. 10...Table, 12...Head part, 36...
Pipette, 40... Plunger rod, 50.52.
...Working station, 54...Liquid receiving tray, 56.
...Chip tray, 62...Chip, 70...Microprocessor, 88...Microtitration tray. Patent applicant Situs Corporation Patent agent Akira Aoki Patent attorney Kazuyuki Nishidate Patent attorney Kyosuke Donakayama Patent attorney Akira Yamaguchi Patent attorney Masaya Nishiyama Go-Ko [Near-8- Continued priority claim from page 1 @ May 5, 1983■United States (US)■4
89866 0 Inventor Timothy Wenberg 541 Arch Street, San Francisco, California, United States 0 Inventor Joseph Widoyunas 3202 Clipper Street, San Francisco, California, United States

Claims (1)

[Claims] 1. To transfer liquid between a plurality of sample containers containing different amounts of liquid or different types of liquid,
A device for filling a small amount of liquid sample by repeatedly using the same pipette and preventing cross-contamination of small amounts of liquid sample or error in handling amount during transfer or mixing, and the device a plurality of syringe-like members having a body portion and a pipette tip member, an engaging means, a means for supporting the plurality of pipette tip members, a means for causing the respective body portions to fully reciprocate, and actuating the engaging means. a body portion having a cylinder formed therein; a plunger disposed within the cylinder; and means for reciprocating the plunger through the cylinder. the pipette tip member being affixable to the body portion for insertion into the sample container for removing liquid from or injecting liquid into the plurality of containers; , an elongated portion of the pipette tip member overlies the other end of the body portion to form a substantial portion of the fluid volume of the body portion, and the pipette tip member forms a fluid seal with the cylinder. has a portion frictionally engageable with a portion of a side wall surface of the body portion, the engagement means contacting the tip member to frictionally disengage the tip member from the side wall surface; a plurality of fingers slidably surrounding an exterior of the pipette tip member, the means for supporting the plurality of pipette tip members supporting the pipette tip member in a vertical position for selective frictional engagement with the sidewall surface; The means for reciprocating the respective body parts is means for frictionally engaging the tip f'x with the surface, and the means for fully operating the engagement means is for removing all the tip members from the cylinder. of the liquid in the sample container, thereby preventing cross-contamination or errors during subsequent dilutions or transfers of multiple samples due to residual fluid carried by the pipette tip member. when transferring liquid between multiple sample vessels containing different amounts of liquid or different types of liquid, all characterized by the fact that different pipette tip members can be selectively and automatically changed after equipment that prevents cross-contamination of small liquid samples or errors in handling volumes. 2. Each of the body portions of the syringe-like member is elastically configured to allow adaptability in mating between the body portion and the pipette tip member to allow full lifting of the tip member of slightly different dimensions. Apparatus according to claim 1. 3. The device according to claim 1, wherein the means for moving the sylanger comprises a step motor and a ball bearing screw 1 drive mechanism interconnecting the step motor and the sylanger. 4. A method consisting of the following multiple steps in which a liquid sample is automatically moved and diluted in a sterile manner between multiple rows of liquid containers; (a) each container is filled with a predetermined amount of liquid of a known concentration; the plurality of rows of liquid containers in a first working station are transferred for horizontal movement onto a support table having a respective row positionable below a row of pipettes; (a)--placing a row of heads on a head part above the support table and movable perpendicularly to the table; c) arranging at least one row of pipette tips 7' for horizontal movement on the support table at the second working station. 2) moving the support table to place the -row of pipette tips in desired positions on a plurality of pipettes on the head; e) engaging a row of pipette tips in the desired position and lowering the plurality of pipettes to raise the engaged pipette tips; f) moving the table to position a row of containers at the first work station below the row of pipettes; g) inserting a tip engaged with liquid in the arranged row of containers and withdrawing at least a portion of the liquid from the tip into the pipette; h) Lifting into the pipette'. b) moving the table to place another row of containers below the pipette; n) Lowering the pipette to drain the liquid in the pipette into a container in another row. ) Lifting the pipette. e) Selected position # at said second work station
b) disengaging the engaged tip from the pipette. 5. A patent claim in which steps (c) to Stella f (e) are repeated for the next row of chips and rows of containers at each work station until the desired number of transfers and dilutions are completed. The method described in item 4. 6. The step (n) is performed at least once after discharging the liquid into the container, taking up the liquid from the container into the pipette, and again discharging the liquid from the pipette to add the liquid in the container. A method according to claim 4, including a step 7'' for mixing with the liquid. 7. Evacuation of the liquid in the pipette in the steller 7' includes actuation of a plunger in the respective pipette; Claim 4, wherein the tip is located exactly at the meniscus of the liquid in the container in order to expel the liquid remaining in the tip by capillary attraction between the liquid in the tip and the liquid in the container. 8. The method according to claim 4, comprising the step of detecting whether all of the chips on a row are picked up consecutively during step (d). 9. The method as claimed in claim 4, wherein chips are released onto the second work station during step (e). 10. Detecting whether all chips have been released after step (wa). 5. The method of claim 4, further comprising the further step of: