Repeated liquid uptake

Abstract

The invention relates to repeated liquid aspiration. A method for aspirating a first liquid medium of two liquid media of different densities from a sample container comprises: lowering a pipette of a laboratory automation device into the sample container until a pipette tip of the pipette has passed a lowering distance from a surface of the first liquid medium in the sample container, wherein the lowering distance is chosen such that the pipette tip passes at least an aspiration volume in the sample container; aspirating liquid from the sample container during lowering the pipette by generating a negative pressure in the pipette, wherein the first liquid medium is aspirated and after the interface and the pipette tip have passed each other, a second liquid medium of the two liquid media is aspirated; measuring the pressure in the pipette during lowering the pipette and detecting the position of the interface when the slope of the pressure changes; when the lowering distance has been passed and the interface has not been detected, aspirating the aspiration volume from the first liquid medium and dispensing the aspiration volume of the first liquid medium into a further sample container.

Description

Technical Field

[0001] This invention relates to a method, a computer program, and a computer-readable medium for aspirating a first liquid medium from two liquid media of different densities from a sample container. Furthermore, this invention relates to a laboratory automation device. Background Technology

[0002] Laboratory automation equipment is used to automate tasks for laboratory assistants, such as testing for specific diseases in patients. Typically, this involves collecting and analyzing patient samples such as blood, urine, and stool through biochemical processes. These processes include various operations such as adding substances, incubation, separation, and quantitative or qualitative measurements of the amount or presence of substances indicating a specific disease.

[0003] In specific procedures, it is necessary to separate two different liquid media with different densities. These liquid media typically separate spontaneously within different layers of the sample container, for example, due to gravity or centrifugation. In this case, the interface between the two liquid media must be detected, and then laboratory automation equipment can use a pipette to aspirate one of the liquid media.

[0004] For example, in prenatal and cancer research, diagnostic methods aim to extract circulating tumor cells and cell-free DNA from plasma / serum of centrifuged blood samples. For this application, the extracted serum should not be contaminated with red blood cells (i.e., erythrocytes) and white blood cells.

[0005] To determine the interface between plasma and red blood cells, laser-based measurements are performed in the NIR (near-infrared) range of the spectrum; these measurements use light to determine the interface. However, opaque sample containers can cause problems. Other systems are based on imaging in the visible range of the spectrum. However, specific illumination is required, and sample containers with stickers may cause issues.

[0006] US 3,897,343 A relates to a blood collection and separator assembly of a type suitable for centrifugation to separate plasma from the cellular phase of blood. A piston with a density between plasma and red blood cells is used to separate plasma and red blood cells.

[0007] US 4 696 748 A uses a porous filter to separate blood.

[0008] CN 207 751 770 U and CN 107 917 835 A describe a method for separating plasma using a separation device.

[0009] US 2012 / 003731 A1 describes a method for determining the interface between plasma and blood cells. A pipette is lowered into a sample container, and the interface is detected based on pressure changes within the pipette.

[0010] CH 682 847 A5, US 5 463 895, US 5 512 247, and US 5 529 754 all relate to pipetting devices and pipetting methods performed therewith. They all describe that the pipette tip should be immersed only a short distance into the liquid medium to be aspirated, because the amount of sample carried on the outer surface of the pipette tip should be as small as possible. Summary of the Invention

[0011] When determining the interface between the first and second liquid media based on pressure analysis within the pipette, a potential problem is that the interface may only be detected after some of the second liquid media has already reached the inside of the pipette. This can be exacerbated by laminar flow of the second liquid media within the first liquid media at the pipette tip. However, sometimes it is desirable to completely avoid aspirating the second liquid media.

[0012] The purpose of this invention is to provide a simple, reliable, and accurate method for aspirating a first liquid medium from two liquid media of different densities from a sample container, wherein the interface between the two liquid media is detected by pressure measurement in a pipette. This method can be applied to more than two different liquid media.

[0013] This objective is achieved through the subject matter of the independent claims. Further illustrative embodiments will become apparent from the dependent claims and the following description.

[0014] The first aspect of the present invention relates to a method for aspirating a first liquid medium from two liquid media of different densities from a sample container.

[0015] This method can be executed automatically by laboratory automation equipment, particularly the controller of such equipment. A controller for laboratory automation equipment suitable for executing this method is also a further aspect of the invention.

[0016] The liquid medium can be a nearly incompressible fluid. The liquid medium can be a liquid and / or a paste-like substance. At a minimum, the liquid medium is suitable for aspiration by pipettes in laboratory automation equipment. Examples of liquid media are aqueous solutions, organic solvents, oil solutions, plasma, separated blood cells, precipitated proteins, gels, granular slurries, etc. In particular, this method can be used to separate plasma from centrifuged blood cells and for liquid-liquid extraction.

[0017] According to one embodiment of the present invention, the method includes: lowering a pipette from a laboratory automation device into the sample container until the pipette tip has passed a descent distance from the surface of a first liquid medium in the sample container, wherein the descent distance is selected such that the pipette tip passes at least through an aspirate volume in the sample container; drawing liquid from the sample container during the descent of the pipette by generating a negative pressure in the pipette. The first liquid medium is drawn before the interface and the pipette tip pass each other. A second liquid medium is drawn after the interface and the pipette tip pass each other. The sample container may be a sample tube or, for example, a hole in a microplate according to ANSI / SLAS Microplate Standards 1 to 4-2004.

[0018] The interface between two liquid media can be a phase boundary and / or a layer between the two liquid media. Geometrically, the interface can be a fundamental planar region between two volumes of two liquid media.

[0019] Aspiration can be performed by applying negative pressure inside the pipette. For this purpose, a pump, such as a plunger or a general negative pressure source, can be used connected to the pipette.

[0020] The aspirate volume can be the maximum volume that the pipette can aspirate, i.e., until the pipette is completely filled. The aspirate volume of a pipette can also be the volume that the pipette is designed to fill. The aspirate volume can also be a volume smaller than the maximum volume of the pipette. For example, as described below, when the first liquid medium is repeatedly removed from the sample container, the amount of the first liquid medium that must be discarded can be reduced by decreasing the aspirate volume, as it may be purified by the second liquid medium due to interface detection.

[0021] The descent distance from the surface of the first liquid medium, corresponding at least to the aspirated volume, can be calculated from the known horizontal area of ​​the sample container. The descent distance can be the aspirated volume divided by the horizontal area, and an offset can be added to the descent distance such that the volume of the first liquid medium remains above the pipette tip when the aspirated volume has been drawn from the sample container. The method further includes measuring the pressure in the pipette during the descent and detecting the position of the interface when the slope of the pressure changes.

[0022] As the pipette tip passes through an interface, the type of liquid medium changes, and therefore its viscosity changes. This causes a change in the pressure slope of the air cushion pressure inside the pipette and / or the tubing. Specifically, when a pump or similar device pumps at a constant volumetric rate—that is, when the amount of fluid removed or added from the volume connected to the pipette is removed or added at a constant rate—the pressure has a constant slope when the viscosity of the aspirated liquid medium is the same. When the viscosity changes, the pressure slope also changes, which can be measured by a pressure sensor connected to the volume of the pipette and the pressure device. The same applies when liquid from the pipette tip is dispensed into two liquids of different viscosities. Again, in this case, the pressure slope in the pipette changes.

[0023] Based on the pressure signal over time, the slope of the pressure signal can be determined, and thus the point where the slope changes can be identified. The location of the interface can then be determined as the position of the pipette tip where the slope has changed.

[0024] The method further includes: drawing the aspirated volume from the first liquid medium when the descent distance has been completed and no interface has been detected, and dispensing the aspirated volume of the first liquid medium to a further sample container. The pipette tip may stop when the descent distance has been completed. The pipette tip is moved to the following liquid level and / or position in the sample container, where the aspirated volume can be drawn from the sample container without further descent of the pipette into the sample container. This differs from a typical aspiration procedure, in which aspiration is performed by continuously and slowly descent of the pipette (compared to the liquid level velocity) into the sample container. It is possible that the pipette is completely filled with the first liquid medium before an interface has been detected. In this case, the first liquid medium in the pipette can be dispensed into a further sample container.

[0025] Since the interface measurement is performed by aspirating the first liquid medium during the pipette descent, only the remaining aspirated volume may be aspirated once the descent distance has been traveled and / or when the pipette movement stops. The amount of the first liquid medium aspirated during the descent can also be considered within the descent distance corresponding to the aspirated volume.

[0026] Typically, the aspirated volume can be drawn during the descent of the pipette tip, wherein a first portion of the aspirated volume is drawn, and a second remaining portion of the aspirated volume is drawn once the descent distance has been completed.

[0027] According to one embodiment of the invention, the method further includes: when the aspirated volume of the first liquid medium has been disposed into the further sample container, repeating the following steps: optionally returning to the source container, lowering the pipette by the descent distance corresponding to the aspirated volume, detecting the interface, and, if the interface is not detected, aspirating the aspirated volume (and particularly the remaining portion of the aspirated volume). When contamination due to interface detection is not yet possible, the process of aspirating the aspirated volume and dispensing it into the further sample container can be repeated until an interface is detected.

[0028] Because this method can be used to separate a first liquid medium from a second liquid medium, it can be repeated and / or repeated several times, and most of the first liquid medium can be transferred from the first sample container to the second sample container. If no interface is detected during filling, the following steps can be repeated several times: filling the pipette to the aspirate volume; dispensing the first liquid medium; and returning the pipette tip to the sample container. The aspirate volume for each repetition does not need to be constant. For example, it may be constant for a fixed number of repetitions (such as three repetitions) and then reduced to half.

[0029] On the other hand, when the interface is detected, the contents of the pipette can be discarded. This could be the entire contents of the pipette or just a small portion of it.

[0030] According to one embodiment of the present invention, the method further includes: when the descent distance has been reached and the interface has not been detected, retracting the pipette tip a safe distance before aspirating the remaining volume. The pipette tip can be retracted a safe distance when aspiration is completed without detecting an interface to ensure that no second liquid medium is aspirated. Aspiration can be stopped or continued during the above-described pipette tip retraction step.

[0031] According to one embodiment of the present invention, the method further includes: stopping the pipette before aspirating the remaining aspirate volume after the descent distance has been passed and the interface has not been detected. The remaining aspirate volume can be aspirated without moving the pipette.

[0032] According to one embodiment of the invention, a first aspiration rate during the descent of the pipette over the specified distance is lower than a second aspiration rate after the descent distance has been completed. The first aspiration rate can be adjusted for interface detection. This can be accomplished at a lower aspiration rate during pure aspiration without pipette movement and / or interface detection. The amount of liquid medium aspirated during the measurement process can be (particularly far) less than the amount aspirated when the descent distance has been completed and / or when the pipette tip stops.

[0033] As described above, the pipette tip descends into the sample container along a path intersecting the interface. For example, the pipette can move with the pipette arm of a laboratory automation device, which may include one or more motors for moving the pipette in several dimensions, particularly the tip. The pipette can move in the z-direction, i.e., vertically, and optionally in the x and y directions. The path intersecting the interface can be defined by moving the pipette tip along, for example, a straight line through the sample container, where the interface is intended. For example, the path may begin at the top of the sample container and / or at the gas-liquid interface within the container and may extend towards the bottom of the container.

[0034] According to one embodiment of the invention, the pipette tip moves in a vertical direction. The vertical direction can be defined as a direction orthogonal to the interface plane. This plane can be adjusted to be orthogonal to the direction of gravity. The movement of the pipette tip can have a component different from the vertical direction. For example, the pipette tip can move along a path inclined relative to the vertical direction.

[0035] As described above, the movement can be from the top to the bottom of the sample container. When the gas-liquid interface is detected, aspiration and / or descent can begin. This can be accomplished, for example, by measuring the capacitance change between air and liquid.

[0036] This can be either: the pipette tip moves while the container remains in the same position; or the container moves while the pipette tip remains in the same position. When the container moves toward the pipette, this can also be considered a descending pipette.

[0037] According to one embodiment of the invention, the aspiration rate of the liquid medium is adjusted such that the movement speed of the liquid level in the sample container is slower than the movement speed of the pipette tip. This prevents more liquid medium from being aspirated because it is present above the pipette tip in the sample container. The aspiration rate can be adjusted by adjusting the volumetric rate of the pressure device. The movement speed of the pipette tip can be the vertical movement speed of the pipette tip.

[0038] According to one embodiment of the invention, the pipette moves at a speed at least 10% faster than the liquid level, particularly 2 to 10 times, 2 to 8 times, and 2 to 5 times (or faster). A specific pipette movement speed can be 6 times faster than the liquid level movement speed. During interface detection, when the pipette movement speed is high, i.e., before the pipette stops, it is possible to prevent a large amount of a second liquid medium from entering the pipette and failing to be detected, under certain conditions (such as laminar flow in the pipette tip).

[0039] According to one embodiment of the invention, the pipette tip is moved only when the pressure device generating negative or overpressure is operating at a constant volumetric rate. As described above, this results in a constant pressure slope when the viscosity of the aspirated liquid medium is the same. For example, the pressure device can be started first, and the movement of the pipette can begin after a certain time interval, such as when the system has stabilized. Similarly, the movement of the pipette can be stopped first, and the pressure device can be stopped after a certain time interval.

[0040] According to one embodiment of the invention, the pressure device includes a plunger, and the pipette is moved only when the plunger moves at a constant speed. With such a pump, a constant volumetric rate can be achieved in a fairly simple manner.

[0041] According to one embodiment of the invention, a liquid medium is drawn in by opening a valve between a hose and a vacuum and / or accumulator for a predetermined opening time. The pressure device may include a valve and an accumulator containing overpressure and / or negative pressure. With this drawing technique, a constant volumetric rate can be achieved in a fairly simple manner if the vacuum source provides a constant vacuum during the drawing time.

[0042] According to one embodiment of the invention, the method further includes: after detecting the interface, withdrawing the pipette from the sample container and discarding a certain amount of liquid from the pipette, thereby discarding the aspirated volume of the second liquid medium. Alternatively, a further small amount of the first liquid medium may be discarded subsequently. Since the interface is detected after the second liquid medium enters the pipette, it may be necessary to remove the second liquid medium from the pipette. The aspirated volume of the second liquid medium can be dispensed into a waste container, or the entire aspirated volume containing the first and second liquids can be discharged into the waste container. Dispensing is typically performed by creating overpressure in the pipette. The first liquid medium remaining in the pipette can be dispensed into a further sample container.

[0043] According to one embodiment of the invention, the method further includes: after detecting the interface, stopping the pipette tip at the detected position and generating overpressure to dispense an amount of the second liquid medium aspirated from the pipette, and optionally dispense an amount of the first liquid medium aspirated from the pipette. In this way, the amount of the second aspirated liquid medium can be returned to the volume of the second liquid medium inside the sample container. Furthermore, when an additional amount of the first aspirated liquid medium is dispensed, it cannot be reliably guaranteed that the pipette contains only the first aspirated liquid medium. The pipette can then be removed from the sample container containing only the first liquid medium. The first liquid medium remaining in the pipette can be dispensed into a further sample container.

[0044] Alternatively, after detecting the interface, the pipette tip can be moved upward a specific (especially small) distance, and then aspirated (especially a small) amount of liquid medium to ensure that no second liquid medium leaks when the pipette is retracted from the sample container.

[0045] According to one embodiment of the invention, the method further includes: lowering the pipette tip in the sample container to the liquid level in the container where the pipette was at the end of a previous detection move in which interface detection was performed; and continuing to detect the interface at that position. In this way, the pipette tip can return to the liquid level in the container, i.e., the liquid level where the first liquid medium is actually located. In this context, the term "liquid level" can refer to a position in the sample container at the same height as the pipette along the path through the sample container. As in the first cycle, the gas-liquid interface can also be detected, for example, by a capacitive method, when the pipette is lowered into the sample container. Detecting the gas-liquid interface in each cycle allows for a lower tolerance in finding the liquid level in the container where the pipette was at the end of a previous detection move in the previous cycle.

[0046] According to one embodiment of the invention, the method further includes: after the first liquid medium of the aspirated volume has been dispensed into the further sample container, lowering the pipette tip in the sample container to the liquid level at the end of a previous detection move in which interface detection has been performed, and continuing to lower the pipette into the sample container from that liquid level until the pipette tip has traveled a distance from the surface of the first liquid medium corresponding to the aspirated volume at the location where the interface was detected. In this way, it is not necessary to perform interface detection in the area already performed. The liquid level at the end of the previous detection move can be stored in a controller and / or device that automatically performs the method.

[0047] According to one embodiment of the invention, the pipette tip is lowered to a liquid level a safe distance from the liquid level at which the detection movement stops. The safe distance can be selected such that the pipette tip is within the volume of the first liquid medium. When the path of the pipette tip is from the top to the bottom of the container, the pipette tip may return to a point above the liquid level at which the interface detection stops.

[0048] According to one embodiment of the invention, a safety distance is selected such that the pressure in the pipette stabilizes at a constant slope during aspiration until the previously detected level where the movement stopped has been reached. In this way, accurate slope detection is ensured, even when the level in the sample container has almost reached the interface. The pressure with a constant slope can be a pressure that exhibits a pressure curve over time that does not deviate substantially from a straight line. For example, noise may be negligible.

[0049] According to one embodiment of the invention, the pipette has an elongated nozzle with a diameter along its extension, the extension varying by at least 10% from the average diameter of the nozzle. The nozzle can be formed in a tubular shape. The nozzle diameter can be substantially constant or the nozzle can have a constant diameter. Alternatively, the nozzle diameter can increase from the tip of the pipette along the nozzle. Using such a nozzle, the viscous friction between the pipette and the liquid medium can be adjusted to a value suitable for performing the method and, in particular, interface detection. Furthermore, the discarding of a second liquid medium can be simplified because laminar flow is generated in the substantially tubular nozzle.

[0050] A further aspect of the invention relates to a computer program for drawing a first liquid medium from two liquid media, the computer program being adapted, when executed by a processor, to perform the steps of the methods described above and below. The computer program can be executed in a computing device, such as a controller and / or a PC of a laboratory automation device, which can be communicatively interconnected with the laboratory automation device. The method can also be executed by an embedded microcontroller of the laboratory automation device.

[0051] A further aspect of the invention relates to a computer-readable medium in which such a computer program is stored. The computer-readable medium may be a floppy disk, hard disk, USB (Universal Serial Bus) storage device, RAM (Random Access Memory), ROM (Read-Only Memory), EPROM (Erasable Programmable Read-Only Memory), or flash memory. The computer-readable medium may also be a data communication network, such as the Internet and / or cloud storage devices, which allow the download of program code. Typically, the computer-readable medium may be a non-transitory or transient medium.

[0052] A further aspect of the invention relates to a laboratory automation device.

[0053] According to one embodiment of the present invention, a laboratory automation device includes: a pipette arm for carrying a pipette; a device for changing the pressure in a volume connected to the pipette for aspirating and dispensing a liquid medium in the pipette; a pressure sensor for measuring pressure in the volume connected to the pipette; and a control device for controlling the device and the pipette arm, and for receiving a pressure signal from the pressure sensor, wherein the control device is adapted to perform the methods described above and below.

[0054] It should be understood that the features described above and below can be the features of the control device, computer program, and computer-readable medium described above and below, and vice versa.

[0055] These and other aspects of the invention will become apparent and will be illustrated by referring to the embodiments described below. Attached Figure Description

[0056] Embodiments of the present invention will now be described in more detail with reference to the accompanying drawings.

[0057] Figure 1 A laboratory automation device according to an embodiment of the present invention is schematically shown.

[0058] Figure 2 A flowchart illustrating a method for aspirating a liquid medium according to an embodiment of the present invention is shown.

[0059] Figure 3 A sample container is shown, illustrating the repeated aspiration process used during embodiments of the invention.

[0060] Figure 4 A graph showing the pressure generated during the method according to an embodiment of the present invention as a function of time is shown.

[0061] Figures 5A to 5D The graph shows the changes in pipette tip position and liquid level over time during the method according to an embodiment of the present invention.

[0062] Figure 6 A graph showing the pipette tip and pump speed generated during a method according to an embodiment of the present invention is shown.

[0063] Figure 7 A pipette is shown that can be used in a method according to an embodiment of the present invention.

[0064] The reference numerals used in the accompanying drawings and their meanings are listed in summary form in the reference numeral list. In principle, the same parts have the same reference numerals in the accompanying drawings. Detailed Implementation

[0065] Figure 1 The diagram schematically illustrates a laboratory automation device 10, which includes an automatically movable pipette arm 12 attached to a pipette 14. Figure 1 As shown, the pipette tip 16 of the pipette 14 can be lowered into the sample container 18 via the movable pipette arm 12. For example, the container 18 may be a test tube containing a centrifuged blood sample.

[0066] The pipette arm 12 can move the pipette 14 and the pipette tip 16 in three dimensions, can lower the pipette tip 16 into the sample container 18, and can withdraw the pipette tip 16 from it.

[0067] Liquid media 20a and 20b, having different densities, are contained in sample container 18 and are separated from each other in the vertical direction, for example, by gravity or centrifugal force. The two liquid media 20a and 20b are separated by interface 22. Interface 22 can be considered as a layer or plane between liquid media 20a and 20b. It should be noted that a second interface 24 exists between liquid media 20a and ambient air.

[0068] For example, the first liquid medium 20a may be plasma, and the second liquid medium 20b may contain blood cells, such as red blood cells and white blood cells. Figure 1 As shown, red blood cells and white blood cells can also be separated from each other, and white blood cells can form a small layer 26 between the plasma and red blood cells. This layer 26 can be considered as part of the interface 22.

[0069] As a further example, the two liquid media 20a and 20b may be two immiscible liquids used for liquid-liquid extraction.

[0070] Furthermore, the laboratory automation equipment 10 includes a pump 28 connected to a pipette 14 via a hose 30. With the pump 28, pressure can be applied to the hose 30 and the pipette 14, causing the pipette 14 to draw or dispense liquid media 20a, 20b, or any other fluid. For example, the pump 28 includes a plunger 29 that is moved to generate negative and overpressure in the hose 30 and the pipette 14.

[0071] Pressure sensor 32, which can be attached to tubing 30 and / or pipette 14, is adapted to measure pressure in tubing 30 and / or pipette 14.

[0072] The control device 34 of the laboratory automation equipment 10 may be part of or connected to the laboratory automation equipment 10, and may control the pipette arm 12 and the pump 28, and may receive pressure signals from the pressure sensor 32.

[0073] Typically, liquid is drawn from sample container 18 using pipette 14 by creating a negative pressure within it. A first liquid medium 20a is drawn, and a second liquid medium 20b is drawn after the interface 22 and pipette tip 16 have passed each other. Pressure in pipette 14 is measured using pressure sensor 32, while the pipette tip 16 and interface 22 move relative to each other, and the position of interface 22 is detected as the pressure slope changes.

[0074] Figure 1 It is also shown that the first liquid medium 20a will be dispensed into a further sample container 36 therein, as described below.

[0075] Figure 2A flowchart is shown for aspirating the first liquid medium 20a and for transferring the first liquid medium 20a to a further sample container 36. This method can be performed by a laboratory automation device 10 controlled by a control device 34.

[0076] In step S10, the pipette tip 16 is inserted into and descends into the container 18 until it reaches the liquid level 24, which serves as the boundary between the first liquid medium 20a and the air. The liquid level 24 can be detected using a capacitive method. Afterward, the pipette tip 16 moves within the sample container 18 along at least a portion of a path 38 that intersects the interface 22 between the first liquid medium 20a and the second liquid medium 20b. As shown, the pipette tip 16 can move vertically; however, more complex paths 38 are possible, which may have a horizontal component and / or inclined segments.

[0077] In step S12, when the pipette tip is immersed in the first liquid 20a, liquid is drawn from the sample container 18 by the pipette 14 using a negative pressure generated in the pipette 14 by the pump 28. Then, when the pipette tip 16 is above the interface 22, the first liquid medium 20a is drawn, and after passing the interface 22, when the pipette tip 16 is below the interface 22, the pipette tip 16 draws the second liquid medium 20b.

[0078] However, this method allows for the attempt to draw in as little of the second liquid medium 20b as possible, for example, as described below.

[0079] In step S12, the amount of the first liquid medium 20a in pipette 14 is estimated, which can be determined from the volumetric rate of pump 28. When the aspirated volume of pipette 14 has been filled, the method continues in step S14, wherein the contents of the pipette are dispensed into the second sample container 36.

[0080] Also during step S12, while the pipette tip 16 is moved, the pressure sensor 32 measures the pressure signal 40 in the pipette 14 (see...). Figure 4 When the position of interface 22 is detected by evaluating the slope of pressure signal 40, the method continues in step S16.

[0081] Otherwise, the method continues in step S14, wherein the pipette 14 is withdrawn from the sample container 18 and the first liquid medium 20a in the pipette 14 is dispensed into a further sample container 36.

[0082] The method then continues in step S12, where the pipette tip 16 is lowered again into the sample container 18. For example, blood samples typically have a volume of 9 mL or 6 mL, with a plasma content of 53-59%, and the pipette 14 will typically be filled before reaching the interface 22.

[0083] Similarly, the gas-liquid interface, i.e., liquid level 24, can be detected, for example, using a capacitance method. After the initial filling of pipette 14, plasma is dispensed into container 36 and pipette 14 can draw more plasma. This can be repeated until the red blood cell interface 22 is reached.

[0084] In step S16, once interface 22 has been detected, the post-detection process can be executed. Specifically, after interface 22 is detected, pipette 14 can be withdrawn from sample container 18 and / or a certain amount of liquid from pipette 14 can be dispensed, thereby discarding the aspirated amount of second liquid medium 20b and, if necessary, discarding a small portion of first liquid medium 20a.

[0085] After step S16, the method can continue to dispense the first liquid medium 20a in pipette 14 into a further sample container 36 and can then stop. Alternatively, pipette 14 can be returned to the first liquid container 18 and the remaining amount of the first liquid medium 20a can be aspirated based on interface detection. This is feasible because the location 22 of the interface position is now known.

[0086] exist Figure 3 The diagram shows the movement path 38 of the pipette tip 16 during step S12 when aspirating liquid. On the first pass, the pipette tip 16 moves a descending distance l from liquid level z0 (where the liquid level is at the start time) to liquid level z2, where the pipette 14 has been filled or is filled to its aspirate volume. The movement from z0 to z2 can be completed relatively quickly and / or at a certain speed, such that the aspirated liquid media 20a, 20b are sufficient for interface detection. At z2, when no interface 22 is detected, the movement can stop and the first liquid media 20a can be aspirated until the pipette 14 is filled to its aspirate volume.

[0087] Alternatively, before the pipette 14 is filled to the aspiration volume, the tip of the pipette can be withdrawn from the liquid level z2 to a safe distance d to the liquid level z1, and at the liquid level z1, the pipette 14 can be filled to the aspiration volume.

[0088] During the second pass and optionally all subsequent passes, the pipette tip 16 in sample container 18 may be lowered to the level where the pipette tip 16 was at the end of the previous detection move. The pipette tip 16 may also be lowered to level z1, which is a safe distance d higher than the level z2 at the end of the previous detection move. The benefits of the safe distance d will be described with reference to the following figures. The withdrawal safe distance after reaching distance l may differ from the descent safe distance, which is used when lowering the pipette back into the container.

[0089] Figure 4 A graph showing a pressure signal or pressure curve 40 is displayed, which is recorded during the movement of the pipette tip 16 through interface 22. For example, Figure 4 The pressure 40 shown can be the pressure of the final detection movement performed in step S12 when interface 22 is finally detected.

[0090] Figure 4 This is an example of a pressure signal 40 recorded during plasma aspiration as it enters the red blood cells. The slope of pressure 40 changes as the pipette tip 16 passes through interface 22.

[0091] Specifically, the graph shows time t on the horizontal axis and pressure p on the vertical axis. At time t=0, a negative pressure is generated, and in the first time interval 42, the first liquid medium 20a begins to enter the pipette tip 16. During this time interval 42, the pressure 40 varies because the flow in the pipette tip 16 is not yet stable. After a stabilization phase in time interval 43, the slope of the pressure 40 is constant. The volumetric rate of the pump 28 has been adjusted to accommodate the amount of the first liquid medium 20a flowing into the pipette 14. At point 44, the pipette tip 16 enters the second liquid medium 20b, which is more viscous than the first liquid medium 20a, and in the following time interval 46, the slope of the pressure 40 changes (the slope decreases). This change in slope is detected by the controller 34, which then stops the movement of the pump 28 and the pipette tip 16. After the pump 28 stops, the pressure begins to rise again.

[0092] Since the first time interval is 42, it may be beneficial to introduce a waiting time t after starting pump 28 and before interface detection 22 becomes active. w This allows us to ignore the pressure change when the first liquid medium 20 begins to enter the empty pipette tip 16. Furthermore, it may be advantageous to stop the z-movement of the pipette tip 16 and the pump 28 upon detection of the interface to avoid drawing in large amounts of the second liquid medium 20b and to minimize the risk of contamination.

[0093] To ensure that the waiting time t is... wThe limited blind phase occurs within a safe distance d, therefore v z t w < d, where v z This is the z-axis velocity of the pipette tip 16. Additionally, the velocity v of the liquid level... l The velocity v should be less than 16 rpm at the tip of the pipette. z It should be noted that v z It may only be the z-component of the velocity of the pipette tip 16, and the velocity of the pipette tip 16 may also have a component in another direction.

[0094] This causes a velocity v at the tip of the pipette 16 during detection movement. z upper and lower boundaries

[0095] v l < v z < d / t w

[0096] Figures 5A to 5D The graphs show the z-coordinates of the liquid level 50 at the tip of the pipette 16 and the liquid level 52 in the sample container 18, respectively, during the descent and retraction of the pipette 14 in the sample container 18 under different conditions.

[0097] These graphs all start from time t0, at which point it has been detected that the tip of the pipette 16 has contacted the liquid level 24 in the sample container 18 and the pump 28 has started.

[0098] exist Figure 5A and Figure 5B In the process, pipette 16 descends at a constant speed into sample container 18 to a distance l until time t1 is reached. Interface detection is performed during this movement. Figure 5A and Figure 5B No interface was detected.

[0099] Pipette 16 is retracted a safe distance d and stopped there. Then, the remaining aspirate volume is drawn up to time t3. After that, the pipette is completely removed from sample container 18 and dispensed into a further sample container 36.

[0100] exist Figure 5A In this process, liquid medium 20a is drawn at a constant rate between times t0 and t3. The liquid level 52 decreases at a constant rate between these two time points.

[0101] exist Figure 5B During the descent of pipette 14 between t0 and t2, liquid medium 20a is aspirated at a first aspiration rate. This first aspiration rate is lower than a second aspiration rate between t2 and t3, during which the remaining aspiration volume of liquid medium 20a is aspirated between t2 and t3.

[0102] Figure 5C and Figure 5D This shows that before pipette 14 has moved a distance l, in time... 1. Context 22 was detected.

[0103] exist Figure 5C Then, pipette 14 is stopped and withdrawn from sample container 18. The aspirated amount of second liquid medium 20b can then be discarded by aliquoting a small amount of the contents of pipette 14. The remaining contents can be aliquoted into a further sample container 36. Alternatively, the entire contents of pipette 14 can then be discarded.

[0104] exist Figure 5D In time After the interface detection at 1, the volume of contents of pipette 14 is dispensed until time. 2. This dispensing is done at the level where the interface has been detected. Afterward, the pipette 14 is withdrawn from the sample container 18 and can, for example, be dispensed into a further sample container 36.

[0105] exist Figures 5A to 5D During the detection movement, the aspiration speed of the liquid medium 20a being aspirated has been adjusted so that the movement speed of the liquid level 52 in the sample container 18 is slower than the movement speed of the pipette tip 16.

[0106] Figure 6 The speed / volume rate v of pump 28 is shown. p The graph shows that the speed / volume rate of the pump 28 can be correlated with the speed of the plunger 29 and the speed v of the pipette tip. z They are proportional. Both quantities are shown as a function of time t.

[0107] To improve interface detection, the pipette tip 16 is only used when the pump 28 has a constant speed and / or volumetric rate v. p The pump 28 moves at a constant volumetric rate v. If the pump 28 is in an acceleration or deceleration phase, interface detection may be more difficult and / or less accurate. This can be: only when the pump 28 generating negative pressure moves at a constant volumetric rate v. p During operation, the pipette tip 16 moves. When the pump 28 includes a plunger 29, the pipette 14 can move only when the plunger 29 moves at a constant speed v. p Move while moving.

[0108] Figure 7Two tip designs of the pipette 14 that can be used in the laboratory automation equipment 10 when performing the method are schematically shown. The upper pipette 14 has an elongated nozzle 54 with a diameter along its extension that varies by at least 10% from the average diameter of the nozzle 54. The average diameter of the nozzle 54 can be smaller than the average diameter of the rest of the pipette (i.e., the pipette body 56), for example, less than more than three times. The elongated nozzle 54 can be advantageous for flushing from the pipette tip 16 (see description of step S16) an unwanted second liquid medium 20b.

[0109] The lower pipette 14 has a tapered pipette tip with a small orifice 58, for example, the diameter of which may be less than 10% of the diameter of the pipette body 56. The smaller orifice 58 allows for higher sensitivity and triggers a faster stop when the pipette tip 16 enters the second liquid medium 20b.

[0110] The upper pipette 14 with nozzle 54 can also have such a small orifice 58.

[0111] After interface detection, a certain amount of the second liquid medium 20b is typically present in the pipette tip 16. The amount of the second liquid medium 20b can be calculated from the pressure-time curve given the aspiration rate. To better flush the second liquid medium 20b in, for example, the nozzle 54, an additional amount of the first liquid medium 20a can be dispensed.

[0112] After interface detection, a negative pressure may exist within pipette 14. Therefore, even though pump 28 has stopped, a further amount of the second liquid medium 20b may flow into the pipette tip. This can be minimized by directly reversing the operation of pump 28 after interface detection and / or switching to dispensing, rather than simply stopping the movement. After interface 22 is detected, pipette tip 16 can stop at the detected location, and an overpressure can be generated in pipette 14 to dispense a certain amount of the second drawn liquid medium 20b from pipette 14.

[0113] Although the invention has been shown and described in detail in the accompanying drawings and foregoing description, such illustrations and descriptions should be considered illustrative or exemplary rather than restrictive; the invention is not limited to the disclosed embodiments. Other variations to the disclosed embodiments can be understood and implemented by those skilled in the art and practice of the claimed invention through study of the drawings, disclosure, and claims. In the claims, the word "comprising..." does not exclude other elements or steps, and the indefinite articles "a" or "an" do not exclude a plural. A single processor or controller or other unit, such as an FPGA, may fulfill the functions of several items listed in the claims. The fact that certain measures are recited in mutually different dependent claims does not imply that combinations of these measures cannot be advantageously used. No reference numerals in the claims should be interpreted as limiting the scope.

[0114] List of reference numerals

[0115] 10. Laboratory automation equipment

[0116] 12 pipette arms

[0117] 14. Pipettes

[0118] 16. Pipette tip

[0119] 18 Sample containers

[0120] 20a First Liquid Medium

[0121] 20b Second Liquid Medium

[0122] 22 Interface

[0123] 24 Second interface, liquid level (between the first liquid medium and air)

[0124] 26th floor

[0125] 28. Pressure equipment, pumps

[0126] 29. Plunger

[0127] 30 Hose

[0128] 32 Pressure Sensors

[0129] 34 Control equipment

[0130] 36 Further sample containers

[0131] 38 Paths

[0132] 40 Pressure

[0133] z0 Liquid level in the sample container

[0134] z1 Liquid level in sample container

[0135] z2 Liquid level in the sample container

[0136] descent distance

[0137] d Safe distance

[0138] 42. Stable time interval

[0139] 43 Constant slope time interval

[0140] 44. Changes in pressure slope

[0141] 46. ​​Constant slope interval

[0142] 48. Pump Stoppage

[0143] t w Waiting time

[0144] 50. Liquid level at the tip of the pipette

[0145] 52 liquid level

[0146] t0 Detects the start of movement

[0147] t1 Movement detection ended, no interface detected.

[0148] t2 The safe distance for moving backward ends

[0149] t3 Removal of pipette begins

[0150] t4 End of pipette withdrawal

[0151] 1. Movement detection complete; interface detected.

[0152] 2. The second liquid medium distribution is completed.

[0153] v z The velocity of the pipette tip (and / or its z-component)

[0154] v p Volumetric rate / pump speed

[0155] 54 nozzles

[0156] 56. Pipette body

[0157] 58 orifices

Claims

1. A method for aspirating a first liquid medium (20a) from two liquid media (20a, 20b) of different densities from a sample container (18), the method comprising: Calculate the descent distance (l) from the surface of the first liquid medium (20a), the descent distance (l) corresponding at least to the aspirated volume; The pipette (14) of the laboratory automation equipment (10) is lowered into the sample container (18) until the pipette tip (16) of the pipette (14) has passed the descent distance (l) from the surface of the first liquid medium (20a) in the sample container (18), wherein the descent distance (l) is selected such that the pipette tip (16) passes at least the aspirated volume in the sample container (18); By generating a negative pressure in the pipette (14), liquid is drawn from the sample container (18) during the descent of the pipette (14), wherein the first liquid medium (20a) is drawn, and the second liquid medium (20b) of the two liquid media is drawn after the interface (22) and the pipette tip (16) pass each other. The pressure (40) in the pipette (14) is measured during the descent of the pipette (14), and the position of the interface (22) is detected when the slope of the pressure-time curve changes; When the descent distance (l) has been passed and the interface (22) has not been detected: stop the pipette (14) before taking the aspirated volume, take the aspirated volume from the first liquid medium (20a); and transfer the aspirated volume of the first liquid medium (20a) to a further sample container (36).

2. The method according to claim 1, Once the aspirated volume of the first liquid medium (20a) has been placed into the further sample container (36), the following steps are repeated: The pipette (14) is lowered by the descent distance (l) corresponding to the volume to be aspirated, the interface (22) is detected, and the volume to be aspirated is aspirated when the interface (22) is not detected.

3. The method according to claim 1 or 2, in, When the descent distance (l) has been passed and the interface (22) has not been detected, the pipette (14) is withdrawn a safe distance (d) before the aspirated volume is drawn.

4. The method according to claim 3, The first aspiration rate during the descent distance (l) of the pipette (14) is lower than the second aspiration rate after the descent distance (l) has been passed.

5. The method according to claim 3, The aspiration rate of the first liquid medium (20a) is adjusted such that the movement speed of the liquid level (52) in the sample container (18) is greater than the movement speed (v) of the pipette tip (16). z )slow; The moving speed (v) of the pipette tip (16) mentioned above z It moves at least 10% faster than the liquid level (52).

6. The method according to claim 3, The pipette tip (16) is moved only when the device (28) that generates negative or overpressure is operating at a constant volume rate.

7. The method according to claim 6, The device (28) that generates negative or overpressure includes a plunger (29), and the pipette (14) is moved only when the plunger (29) moves at a constant speed.

8. The method according to claim 3, After the interface (22) is detected, the pipette (14) is withdrawn from the sample container (18) and a certain amount of liquid is discarded from the pipette (14), thereby discarding the second liquid medium (20b) of the aspirated amount.

9. The method according to claim 3, After the interface (22) is detected, the pipette tip (16) is stopped at the detected position and an overpressure is generated to dispense the amount of the second liquid medium (20b) and / or the amount of the first liquid medium (20a) drawn from the pipette (14).

10. The method of claim 3, further comprising: After the first liquid medium (20a) of the aspirated volume has been dispensed into the further sample container (36), the pipette tip (16) in the sample container (18) is lowered to the liquid level at the end of the previous movement where the interface detection has been performed, and the pipette (14) is lowered into the sample container (18) from that liquid level until the pipette tip (16) has passed the descent distance (l) corresponding to the aspirated volume from the surface of the first liquid medium (20a).

11. The method according to claim 10, The tip of the pipette (16) is lowered to a level (z1) at a safe distance (d) from the level (z2) where the previous movement stopped.

12. The method according to claim 5, The moving speed (v) of the pipette tip (16) mentioned above z It moves 2 to 10 times faster than the liquid level (52).

13. A computer program for drawing a first liquid medium (20a) from two liquid media (20a, 20b) of different densities from a sample container (18), wherein, when executed by a processor, the computer program is adapted to perform the steps of the method according to any one of claims 1 to 12.

14. A computer-readable medium storing a computer program according to claim 13 in the computer-readable medium.

15. A laboratory automation device (10), comprising: A pipette arm (12) is used to carry a pipette (14); Pressure device (28) for changing the pressure in the volume (30) connected to the pipette (14) for drawing and dispensing liquid media (20a, 20b) in the pipette (14); Pressure sensor (32) for measuring pressure in the volume (30) connected to the pipette (14); A control device (34) is used to control the pressure device (28) and the pipette arm (12), and to receive a pressure signal (40) from the pressure sensor (32); The control device (34) is adapted to perform the method according to any one of claims 1 to 12.

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