JPH06504369A - Transfer of biological samples - 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] This invention enables the transfer of Vetribrate to individual wells of microtiter plates. Concerning the movement of biological samples, for example the movement of cell colonies.

Background of the invention This invention is suitable for automatic colony picker equipment and for use in scientific research. was devised to provide a method for

The process that has to be carried out is that of a 90 mm diameter veterinary plate. Aliquot each colony on a microtiter plate into individual wells on a microtiter plate. It is the movement of cells. The maximum size of the colony is 0.5 mm at the bottom and 3 mm at the top. can be randomly grown with 50 to 100,0 colonies per vetri dish. obtain. The wells on the microtiter plate are spaced 12 x 8 grids apart, 9 mm from the center. It is composed of lids. Transferring colonies from vetri dishes to microtiter plates This procedure is currently performed manually.

When collecting colonies, avoid extreme concentration because they are small and can be difficult to see. or required.

The microtiter plate is first unpacked from the sterile plastic packaging. It will be done. This may be done in a non-sterile environment as long as it remains on the lid or plate. stomach. Each plate has a separate base and Written and labeled using indelible marker on both lids. each my A written record was also kept on the black titan well plate and each well was removed from the petri dish. It shows in detail where the colonies came from. The plate is then cross-contaminated or The plates are moved to a flow foot where they can be opened without fear of external contamination. microphone The rotaita well is then filled with a filling means that dispenses a precise amount of liquid into each well. Fill with growth medium.

Cells from each colony in a Petri dish are collected on a basis of 1 colony per well. Must be transferred to a microtiter plate. In addition, cross-contamination loading cells from more than one colony into a well) must be avoided. stomach. This can be done using a sterile toothpick or needle. Therefore, a new needle is used for each colony. repeated Actions that should be taken include: l) Take out a new sterile needle.

2) Push the needle into the colony on the Petri dish to collect some cells.

3) Transfer the cells to the well and use the needle 1 of the microtiter well to transfer the cells to the well. Turning the.

4) Discard the needle.

5) Repeat from step l.

This work causes mental fatigue, with a maximum of approximately 1,500 collections per day.

This operation is carried out in a flow hood to ensure that the wells are not contaminated from the environment. Must be executed below.

Known collection procedures impose significant bottlenecks on the DNA sequencing and mapping process. Introducing Difficult to recruit staff to perform this tedious work Scientists who must collect materials must do so themselves. cause a situation where This process is directed at isolating specific genes and This means that if more samples are collected, the required genes will be isolated. produces a greater probability than is expected. A typical experiment requires as many as 50,000 samples. and a considerable amount of time must be spent on collection. I want to do it, but it's not a scientific book. faces a situation in which it is not possible to perform this collection or to perform any experiments at all.

In other words, the collection process considerably inhibits performing many experiments. It acts as.

Lack of suitable machines and obvious access to machines to perform this collection process Demand drives the development of the invention According to one aspect of this invention, biological samples are The device for transferring from one container to a second container includes a head carrying a plurality of protruding needles. a support means for supporting the container; and a first hand for moving the head between a plurality of positions. a step, each position being given to a corresponding one or operative position of the needle; the head and the support means, and the head or the first container and the operative A collection position that is placed in relationship or a deposit that is placed in operative relationship with the head or second container. a) a second means for bringing the head into one of the positions; to provide relative movement between the head and the support means when the operating needle or the first container and then withdrawn from the first container, thereby causing the needle to be removed from the first container. and b) the head and support hand when the deposit is in position. resulting in relative movement between the stages, the operating needle enters the second container, and then there. causing the needle to withdraw from the needle, thereby depositing the sample into the second container. a third means for; The first and third means operate alternately in order to load each sample onto the needle. the second means is then operated, and the first and third means are then operated. The second container is operated to alternately deposit the loaded samples in sequence.

Preferably the first means provides for a rotational indexing movement of the head, and the needle is indexed rotationally of the head. Projects radially around the axis of rotation. In this case, each needle is in a downwardly protruding operative position. The head may be positioned on the support means, and the third means supports vertical movement of the head. Brings back action.

In a preferred embodiment, the second means are perpendicular to the direction of reciprocating movement of the head. providing sliding movement of both the head and the support means in mutually perpendicular directions to obtain .

A sterilization station is preferably provided to sterilize the needles prior to sample collection. The sterilization station is conveniently a bath of organic compounds such as ethanol for needle polishing. together with particles or beads that provide a purifying action, the latter being recycled in a sterilization tank. be transferred. Alternatively or in addition, a sterilization station cleans and sterilizes the needles. For this purpose, an ultrasonic generator may be provided.

Transferring a biological sample from a first container to a second container according to another aspect of the invention A method is provided, which is used sequentially to collect samples from a first container. using a head having a plurality of protruding needles, the head and the container steps resulting in relative movement between the steps and then sequentially starting from each needle. depositing individual samples into two containers.

The device according to the invention will be described by way of example with reference to the accompanying drawings, in which: FIG. 1 is a front perspective view of the device; FIG. 2 is a plan view of the device of FIG. 1; FIG. 3 is a partial perspective view of parts of the apparatus of FIGS. I and 2, and FIGS. 4a to 4d. Figure 4 illustrates how the device removes plates from the carousel. A series of diagrams, Figure 5 shows how the plates are placed accurately, and Figure 6 shows how each plate is placed. Figure 7 shows how the device can be identified and disassembled by the autopsy system. diagonal direction indicating how to divide the area of the veterinary plate into 6 frames to FIG.

Referring to FIG. 1, the device includes an external case IO that has a hinged door 12 that can be closed. surrounded by Sterilizing airflow up through case IO in the general direction of arrow 14. This flow is filtered to remove toxic components.

The apparatus consists of two plate handling devices, generally designated 20, between which are located Includes carousels 16 and 18. The circular conveyor 16 is an input circular conveyor. The carousel 18 is a micro-tight well. Serves as an output carousel for receiving plates. Plate handling equipment Input the selected veterinary rate from the carousel 16 and the selected microphone. It is possible to lower the rotator plate from the output carousel and select Assemble the selected veterinary plates and microtiter plates into the picker head. It is possible to move it to a position below the flashlight 22 and CCD camera 24. Details below The picker head assembly removes the microtiter from the Petri dish as described in It is possible to transfer colonies to individual wells of plates.

The case 10 allows the passage of airflow and includes two spaced carousels 16.1. The intermediate panel 26 is formed with a large gap for accommodating the 8. Enter The power carousel 16 consists of two removable ramps angularly positioned at 180°. 28, each having space to maintain 15 vet plates 30. do. This carousel 16 is connected to a panel 26 and a plate handling device 20. by a stepper motor 34 for precise adjustment of the vertical position of the carousel 16. It is attached to a driven central lead screw 32. Another motor 36 can be selected rack 28 can be made to overlap well with the plate handling equipment a20. The circular conveyor 16 is rotatably driven to achieve this.

Similarly, the output carousel 18 has four dismounts angularly positioned at 90°. accommodating possible racks 38, each holding 15 microtiter plates 40 have the space to do so. The carousel 18 has an output carousel with respect to the panel 26. Driven by another stepper motor 44 for precise adjustment of the vertical position of the bear. is attached to another central lead screw 42. Another motor 46 is selected Enables the rack 38 to overlap well with the plate handling device 20. The output carousel 18 is rotatably driven in order to

With particular reference to FIGS. 1 and 2, the plate handling device 20 is shown vertically in the guide 50. Includes a transverse plate 48 mounted for direct transfer operation. There are two plates 48. horizontal slide 52.54 from which the suction plate 60.62 is attached. The respective arms 56 and 58 protrude. The device 20 is one of its components or 67 Under the control of the linear motor shown, the motion can be shifted back and forth on the horizontal slide 66. It also includes a plate holder 64 that allows for. Its anterior plate access position (FIG. 2), the holder 64 has a circular shape for plate removal and replacement. At the colony transfer position adjacent to and behind conveyors 16 and 18 (Figure 3) , the holder 64 holds the selected veterinary plate and the selected microtiter plate. The picker bed and camera are supported under the picker bed and camera as illustrated in FIG.

Picker head assembly 22 also supports camera 24 for sliding movement. Horizontal movement adjustment is possible on the horizontally fixed slide 68 which has a horizontal fixed slide 68. picker assembly The camera 22 and the camera 24 are under the control of a linear motor whose components are shown at 69. can move together along the slide 68. picker head assembly The picker head 70 carries six equally angularly spaced needles 72 and each Rotation of the head 70 about a horizontal axis to continuously bring the needle 72 into an operative position. and a stepper motor provided to move the needle vertically downwards. Stick out. A further motor 74 drives a disc crank 76 and a connecting loft 78. and they move the picker head 22 and camera 24 along the slide 68. Direction of sliding movement and direction of sliding movement of holder 64 along slide 66 This has the effect of reciprocating the picker head 70 in a direction perpendicular to both directions. do.

Figure 4 shows how the plate handling device inputs the selected veterinary plate 30 into the circular controller. Indicates whether to remove it from the bear 16.

In Figure 4a the suction plate 60 is shown in its retracted position. a The arm 56 is then driven along the slide 52 and enters the suction plate 60. force into the selected recess in the carousel. Suction to plate 60 The suction plate 60 then closes the lid of the selected veterinary plate 30. Grasping (Fig. 4b), and therefore the withdrawal of the arm along the slide 52, is selected. removed from the carousel and placed in its forward position (Fig. 4c). This causes the holder 64 to be pulled. Plate 48 in guide 50 The upward movement of then causes the lid of the veterinary plate to be removed. FIG. 4d), the latter being prepared to be transferred by movement of the holder 64 to the transfer station. Can be done. The suction plate 62 is connected to the selected micrometer from the output carousel. operate in a corresponding manner to remove and replace iterates. It will be understood.

The exact location of the selected veterinary plate (or microtiter plate) Rotate the plate against the spring to the correct position to close the microswitch By providing holder 64 with locating means in the form of pivot bar 80 (FIG. 5). The second microswitch is located in the plate receiving recess in the holder 64. provided on the other side of the

A microswitch detects the presence or absence of a plate for error recovery purposes. Ru. A stop 82 on holder 64 limits insertion of the plate. This configuration is the placed plates are positioned at the same location in the plate holder 64; ensure that

The location of colonies on the vetrib plates is identified by camera 24. Resolution In order to increase the degree of The camera continuously examines each frame separately. Ru. This results in a controlled shift of holder 64 along guide 66 and guide 68. This is done by shifting the camera 24 along. The image in the frame is of the colony. Processed by a control computer that calculates the coordinates. This information (Guide 66 holder 64 (along guide 68) and picker head assembly (along guide 68) To drive the picker head assembly or after being precisely positioned over the colony It is used until it is ready to be collected by the operator or the operating needle of the picker head.

The veterinary plate is positioned under the holder with a light source 84 ( 1 and 2). The holder 64 allows the light to It has a rectangular cutout that allows you to reach. As a result, the illumination light It is directed diagonally upward toward the tributate 30. The light hitting the colony is The camera 24 is therefore reflected from the colony as a light field against a dark background. I see. The diagonal direction of the illumination light is suitable for certain types of plates, e.g. phage plates. Improve contrast. Another type of plate, e.g. Ecoli, yeast In this regard, illumination is provided by a light source located on the holder. these two lights The sources are independently controlled.

A sterilization tank 86 (FIGS. 1 and 2) is provided to sterilize the picker needle 72. It will be done. The bath contains 70% ethanol and 0.5 mm diameter glass beads. 6 Place one colony into individual wells of a microtiter plate and then transfer to a picker. The needle is operated to immerse the needle in the sterilization bath 86. The head 70 then moves the needle 7 The beads are rotated to purify 2, and the beads provide an abrasive 1 action. picker head 7 0 is then ready to collect 6 more colonies from the veterinary plate. ing.

FIG. 6 shows how the identification code 88 on the side of the veterinary plate is detected by the camera with the help of the mirror 90. The identification of the selected veterinary brate is examined by the control computer. data that can be supplied to the data software. A similar array can be added to the software using micro Can be used to enter the titer plate's identification.

The device functions in the following manner. The input circular conhair 16 is loaded with a veterinary plate. each having a colony suspended in an agar gel. These colonies are 0.5 It can be as small as mm. By using the device, colonies are grown on an agar gel. From a random order to a defined grid in the wells of a microtiter plate. The colony can then be grown and further analysis performed. Ma The microtiter plate (empty at this stage) is placed on the output carousel 18. Ru.

The rotary drive and axial shifting motion that can be imparted to each carousel is The selected veterinary plate and the selected microtiter plate are The handling device 20 is made to overlap well, so that the selected plate can be moved as required. This means that it can be removed from the carousel means.

The selected veterinary plate is connected to the input carousel as described with reference to Figure 4. 16 and the selected microtiter plate is similarly output circular controller. removed from bear 18. Vetribrate and Matrix (as in Figure 4d) When the lid of the microtiter plate is removed, the holder 64 holds the selected microtiter plate. For transporting the plates and microtiter plates to the collection station, the It is moved backwards so that the At this stage, the individual microtiter plates The wells are filled with a suitable medium by the filling means 92 shown in FIGS. 1 and 2. It will be done. The filling means has an 8-way manifold, which allows 8 microtiter weights to be filled at one time. Distribute liquid culture medium to the staff. The amount of liquid dispensed is controlled by a pinch valve that controls the flow of pressurized liquid. controlled by sensor, pressure drops or fluid level is low. Indicates when it is too much.

The light source 84 is energized to illuminate the veterinary plate with obliquely directed light. The six divisions or frames of the tribrate are sequentially scanned by the camera 24. As a result of the controlled transfer of the radar 64 and the camera 24, as described with reference to FIG. An autopsy will be conducted as if it were done. The position of the colony on the agar medium is thus determined. Ru. These locations are typically in the form of xy coordinates, and this information is provided on slide 68. along the picker head assembly 22 and along the slide 66 the holder 6. used by the control computer to move the picker head to the collection position. Push the door assembly downwards just above the location of the first colony to be harvested. A needle 72 to be taken out is provided and positioned. The stepper motor is then energized by the picker This causes the head 70 to perform a vertical reciprocating motion. This is the needle tip Bring it downwards to make contact with the positioned colony that will adhere to it. It is pulled back from the agar by the upward movement of the needle. Next to the operative downward facing position The stepper motor is energized to rotate the helad 70 60 degrees to bring the needle. Ru. Movement of picker head assembly 22 and holder 64 as required A new needle 72 is brought over the next colony to be harvested. The motor is energized by This process causes new needles to be collected from established colonies. The steps continue until all six needles are loaded with their respective colonies.

Colonies are then deposited into individual wells of microtiter plates 40.

This may be one of the mounted needles or the one on the first well of the microtiter plate. transfer the picker head assembly 22 and holder 64 (if necessary) in the Deposit is carried out by bringing the picker head into position. The motor energization is Place in microtiter wells to deposit colonies into medium in wells. lower the needle, then the needle is raised. To give a newly posted needle rotation of the picker head 70 and the picker head assembly 22 and picker head assembly 22 as required. A transfer of Ruda 64 is then performed to deposit a second colony, and this Seed all six colonies or six colonies in each of the microtiter plates. This continues until he is entrusted to El.

The picker head assembly is then transferred to a sterilization bath 86. In the needle 72 tank Rotation of the head during immersion in the ethanol solution cleans the needle 72 and creates six new A set of colonies can be prepared for collection from the veterinary plates 30.

Once all colonies or veterinary plates have been removed (or microtitered) When the plate well is fully filled with colonies) the holder 64 is placed in the forward position. Shift the lid back to the veterinary plate (or microtiter plate), The vetribrate (or fully filled microtitre plate) used is The corresponding carousel is replaced by a new veterinary plate (or micro titer plate) is removed from the carousel.

The control computer controls the application of suction to the suction plate 60.62 and the portion controls all operations. The imaging system of which camera 24 is a part Colonies can be identified and three different colony types viz. Yeast colonies that are either blue or white in color or bacterial colonies that are either blue or white It is possible to distinguish between phage plaques that are either transparent or transparent. one piece The color may only appear on one veterinary plate. The device described allows colonies to It is possible to collect and move reliably, automatically and quickly.

As a variant, the head carries multiple vertical needles, which are horizontally spaced and each has its own solenoid, which moves a particular needle to its extended position It can be energized. Instead of relying on suction, plate handling means move the latter. A simple mechanical hook that engages the back of the plate may be used to

Vetribrate and microtiter instead of the pair of carousels described It is also possible to have a single circular conhair that accommodates both plates.

international search report Continuation of front page (72) Inventor Jones, Peter UK, CBI 24 H. Earl Cambridge, Sawston, Cattley's Walk, The Old Stables (no address) (72) Inventor: Watson, □Andrew Robert, UK, Niss R. ・68 A.X. Tyne and Wear, Sunderland, South ・Benz, 115 Weardill Avenue (72) Inventor Mallett, Frank UK, 4 varnished Cambridge, Kinnert Way, (72) Inventor Bond, Christophe Charles, England, 3 N.V. with C.V. A Cambridge, Collage Road,

Claims (13)

[Claims] 1. A device for transferring a biological sample from a first container to a second container, the device comprising: a head carrying a protruding needle, a support means for supporting the container, and a plurality of positions. and a first means for moving the head between, at each position, a corresponding one of the needles. is placed in an operative position, and further a head is placed in operative relationship with the first container. either a collection position or a deposit position in which the head is placed in operative relationship with the second container; bringing about relative movement between the head and the support means to bring the head to a) relative movement between the head and the support means when in the collection position; the operating needle enters the first container and then withdraws from the first container. and causing the needle to collect the sample from the first container. and b) providing for relative movement between the head and the support means when in the deposited position; causing the operating needle to enter the container and then withdraw from it, thereby a third means for depositing the sample into the second container; 3 means are operated alternately in order to load each sample onto the needle; means are then operated, and the first and third means are then loaded onto the second container. A device that is operated in alternating order to deposit a given sample. 2. The first means results in a rotational indexing movement of the head, with the needle on the axis of rotation of the head. 2. The device of claim 1, wherein the device projects radially. 3. The head is positioned on the support means and each needle projects downwardly when in the operative position. 3. The apparatus of claim 2. 4. The second means includes sliding movement of both the head and the support means at right angles to each other. 4. The apparatus of claim 3, wherein the apparatus provides: 5. Claim 3 or Claim 4, wherein the third means provides vertical reciprocating movement of the head. The device described. 6. The head is aligned both in the sliding direction of the support means and in the direction of reciprocating movement of the head. 6. The device according to claim 4 or claim 5, wherein the device slides in the direction of travel. 7. Prior to sample collection, including a sterilization station to sterilize needles A device according to any one of the claims. 8. Sterilization stations are equipped with particles or beads that provide an abrasive cleaning action on the needles. 8. The apparatus of claim 7, comprising a bath of organic compound. 9. The support means can be shifted into the plate access position where the plate handling means are placed Mounted for controlled rotation with a plate holder that can be shifted parallel to the axis of rotation , including a carousel so that it overlaps well with the plate access position. brings the stored selected containers to a carousel where they are placed on a plate 9. The material according to claim 8, wherein the material is introduced into and removed from the carousel by means of handling means. equipment. 10. A camera for inspecting the first container and a sample seat in the first container. An image processing means connected to the camera for extracting the target, and and illumination means for directing light onto the first container at an oblique angle. The device described in any of the above. 11. The preceding claims include means for passing a sterilizing air stream through the components of the device. The device described in any of the above. 12. A method for transferring a biological sample from a first container to a second container, the method comprising: having a plurality of protruding needles that are used sequentially to collect samples from one container the step of using the head and the relative movement between the head and the container. step and then sequentially remove individual samples from each needle into a second container. and depositing the method. 13. The first container is a Petri plate and the second container is a microtiter plate. 13. wherein the individual samples are deposited in individual wells of the plate. Method described.