A liquid handling device and method for detecting fluorescence
Background
Liquid handling covers a broad range of products and processes where handling liquid samples, such as blood samples and saliva samples, is critical and minor errors and mistakes may render a test batch useless or if an error is undetected lead to maltreatment of a patient.
Many different devices can be acquired for liquid handling. It can be done by hand or dedicated devices can be obtained that perform dedicated routines and applications within a narrow area of liquid handling. Also, large batch machines are available that may provide a large throughput of liquid samples.
However, for some applications there is a need for automated liquid handling devices which are designed to help automate routine liquid handling tasks to free up time where the manual liquid handling is too cumbersome and where the dedicated liquid handling machines do not provide sufficient flexibility and where the high-volume batch machines are too expensive and bulky.
For this segment there are provided desktop automated liquid handling devices, such as the epMotion range of liquid handling devices by Eppendorf SE.
Such automated liquid handling devices, also referred to as liquid handling devices herein, enable a large range of liquid handling applications. One common feature is that these automated liquid handling devices comprise a worktable on which different types of labware can be placed depending on the specific need.
Automated liquid handling devices or automated liquid handling systems are used, in particular, in biological, biochemical, medical, forensic or chemical laboratories for handling mostly liquid laboratory samples. The handling operations in particular include metering, mixing, dividing, diluting, heating/cooling, biochemically or chemically altering, or analyzing the samples. In the process, the samples may be altered with regard to their quantity, composition, or physically, biochemically, chemically, or in another manner. Automated liquid handling devices are used, in particular, for producing dilution series, distributing reagents, transferring samples from vessels to plates, normalizing samples, PCR setups, real-time PCBs, purifying nucleic acids using magnetic bead methods, preparing samples for next-generation sequencing, and processing cell essays or routine pipetting procedures. Handling samples automatically is faster, more accurate and more reliable than doing so manually. An automated liquid handling device and a method for handling laboratory samples are for example described in EP 2 713 166 A1 .
The automated liquid handling devices like ep Motion(R) 5070, 5073, 5075 by Eppendorf AG have a worktable with workstations for sample containers and storage areas for gripper and dosing tools. Samples can be stored and handled in the sample containers. The sample containers include, for example, reagent vessels, reaction vessels, reservoirs, microtiter plates (microplates), pipetting containers like pipette tips or syringes, and waste containers. As disclosed herein sample containers are also generally referred to as well plates, but can be of any other type understood by the person skilled in the art as sample containers as discussed herein.
Sample containers are commonly placed directly on the worktable of the liquid handling device or stored in storage devices on the on the worktable of the liquid handling device. In some cases the sample containers or storage devices may be placed outside of the automated liquid handling device. Said storage devices include, for example, pipette tip carriers, holders for pipette tip carriers, racks for reservoirs or vessels, and adapters for adjusting the height of microtiter plates. A carrier for pipette tips comprising a pipette tip holder and a pipette tip carrier is described in DE 10 2.009 006 51 1 B4. A modular storage system for laboratory liquids is described in EP 2 168 684 B1 .
Furthermore, the automated liquid handling devices comprise an XYZ robot arm having a tool holder to which a gripper tool and/or a dosing tool can optionally be connected. A gripper tool, a pipetting tool and a tool holder for an automated liquid handling device are described in EP 1 407 861 B1 . The robot arm can be moved in a program- controlled manner in order to pick up a gripper or dosing tool from a storage place or set said tool down thereon, to relocate sample containers in the work area using the gripper tool, to pick up and release pipette containers like pipetting tips or syringes using the pipetting tool, and to take up and dispense liquids using the pipette tips or syringes.
The dosing tools used with pipetting tips have at least one neck onto which a pipette tip can be clamped in a sealing manner. By means of a displacement apparatus arranged in the dosing tool and connected to a hole in the neck via a line, air can be displaced through an upper opening in the pipette tip in order to aspirate or expel liquid through a lower opening of the pipette tip. In the case of a multiple pipetting tool, the necks for clamping on pipette tips are arranged in one or more rows corresponding at least to some of the wells in a standardized microtiter plate. Standardized microtiter plates having 96 or 384 wells and multiple dosing tools adapted to the spacing of the wells are used, in particular. The pipette tips made of plastics material can be separated from the pipetting tool and discarded in a waste container after one single use in order to prevent cross-contamination of different laboratory samples during metering.
The workstations have a rectangular format adapted to the footprint of a standardized microtiter plate (SBS/ANSI format) in order to store thereon standardized microtiter plates, pipette tip carriers or other sample containers having a footprint that corresponds to the footprint of a standardized microtiter plate. The footprints of the workstations are limited by alignment means, which ensure precise alignment of the sample containers.
However, some applications and processes are not provided on the worktable and have to be performed in another location. This requires that the samples are moved manually off the worktable and out of the automatic workflow which is both time consuming and introduces many other risks for errors such as risk of contamination and spillage.
Such is the case when the luminescence, typically fluorescence, examination step is introduced. Fluorescence detection is used in many liquid handling operations. A fluorescent dye is added to the sample which binds to a specific component in the sample, and by exciting light with a specific wavelength on the sample the emitted radiation from the fluorescent dye is detected and can be used to derive the amount of the specific component the dye binds to is present in the sample, which may be referred to as the sample concentration. There are a vast number of such luminescent dyes available on the market, each with specific application focus and using different wavelengths, but the general principles remain the same.
The automated liquid handling devices are used to add the fluorescent dye to the sample, however, for excitation and detection the samples need to be moved off the worktable and to a separate device for fluorescent registration. As mentioned this is time consuming and introduces a step in the process which is susceptible of errors. Thus, there exists a need for providing luminescent detection, and especially fluorescent detection on the worktable of automated liquid handling devices.
Summary
In one aspect there is disclosed a liquid handling device for transferring liquid and handling liquids. The liquid handling device comprises an operating chamber arranged within a housing, and a worktable area arranged in the operating chamber for receiving/arranging labware. The labware can for example be at least one well plate, at least one pipette tip container containing at least one pipette tip and/or at least one dosing tool.
A carrier arm for moving tools between labware arranged on the worktable area is furthermore provided in the operating chamber.
A luminescence tool is furthermore provided in the liquid handling device, where the liquid handling device comprises at least one probe head with at least one light emitter for emitting a luminescent signal and at least one light receiver for detecting a luminescent signal, wherein the luminescence tool is configured to be arranged on the carrier arm.
By providing the luminescence tool on the carrier arm it is possible to perform luminescent registration on the worktable. For example, it has been shown that luminescent registration in well plates can be performed directly using a luminescence tool as disclosed herein. Generally, as used herein the term ‘luminescent registration’ should be understood as the action of both emitting the luminescent signal to a sample and subsequently detecting the luminescent signal returned from the sample. It is well understood within luminescent excitation that the signal emitted to the sample by a probe unit may have a different wavelength than the signal subsequently received by the probe unit from the sample.
Accordingly, moving the samples in the well plate or the whole well plate into a separate luminescent registration device can be avoided. Thus, separate handling of the sample is avoided and risk of spillage, contamination and other factors that may compromise that sample is reduced or eliminated.
Even further, luminescent registration, such as fluorescent registration is in many cases used in a liquid handling workflow where liquid handling is performed before and after the luminescent registration. This can for example be the case when the samples are normalized. Thus, having an intermediate luminescent registration step will typically require the user’s attention, either in order to handle the samples or for example in order to export the luminescent measurement data from a luminescent registration device and importing it to a liquid handling device and programming the liquid handling device to perform the proper normalization based on the luminescent measurement data. In other words, a liquid handling device as the one disclosed herein will allow the user to walk away from the process, increasing the users walk away time (time available for completing other tasks in the laboratory while the liquid handling device performs its applications), and allowing the user to focus on other tasks.
Moreover, luminescence is a general term covering a plurality of different principles involving the ability of a molecule to emit light. For example, such light emission can be triggered chemically, electrically or by radiation. One type of such luminescence is photoluminescence, where the molecule is triggered to emit light with one wavelength by exposing it to light at another wavelength. Typically, well known photoluminescence principles are fluorescence and phosphorescence where in particular fluorescence is generally used within the field of liquid handling.
Within fluorescence the time between absorption and emittance of light from the fluorescent molecule is very short, in some case it can appear almost instantaneously, whereas, in phosphorescence the time is longer between absorption and emittance, for example minutes or hours.
Thus, as used herein terms such as luminescence, luminescent and luminescent registration/measurement are used to generally refer to the principles applied herein and although this would typically be applied using fluorescence within the current field the current disclosure also covers that in particularly within other photoluminescence areas, such as phosphorescence the teachings herein can be applied.
In general terms such as luminescence and fluorescence are used herein when referring to the principle thereof or to tools and objects that are used to take advantage of the principle. Luminescent and fluorescent are used when referring to a device or sample that itself exhibit such properties, e.g. the luminescent or fluorescent dye itself, the sample wherein the dye is mixed or any other object or device that specifically exhibits such properties. Luminescent and fluorescent measurement/registration as used herein refers to the action of detecting the luminescent or fluorescent properties of an object, such as a sample.
Furthermore, when referring to a luminescent or fluorescent signal or wavelength it should be understood that such signal or wavelength is optical and applied with the intent to excite a luminescent or fluorescent dye when such signal or wavelength is emitted from e.g. the luminescence tool, or measured/d elected with the intent to meaure an optical wavelength emitted from the fluorescent dye (after it has been excited).
Typically, luminescent and fluorescent dyes are indicated to have a certain excitation and emission value, however, in many cases such dyes operate within a wavelength range where the indicated value is the optimal value. E.g. a fluorescent dye may be indicated to have an excitation wavelength of 560nm, however, it will still be excitated when exposed to light with wavelengths between 500 and 580 nm. It will, however, be most efficient at 560 nm.
Furthermore, a fluorescent dye indicated to have an emission wavelength of 570nm when excited will still excite a range of wavelength, for example from 550 to 680 nm however the energy content (intensity) will peak at 570nm.
For the purpose of the current disclosure, the person skilled in the art will understand that commercially available luminescent and fluorescent dyes may be used herein without limiting or deviating from the scope of the disclosure. The person skilled in the art will also understand that when referring to luminescent or fluorescent signals or wavelengths a range of wavelengths may be covered. Furthermore, the term ‘signal’ may refer to light with a wavelength or comprising a spectrum of wavelengths. However, the term signal may also/additionally refer to a signal which represents or contains information relating to light with a wavelength or comprising a spectrum of wavelengths. Such a signal may for example be a digital signal and processed on a computer, e.g. in a computer implemented method.
In some liquid handling devices, a separate luminescent registration device is arranged on the worktable, where a cartridge or well plate has to be inserted for luminescent registration. By arranging the luminescence tool above the worktable, e.g. on the carrier arm, as discussed, the footprint of the worktable can be kept small since separate luminescent registration devices on the worktable can be avoided. In another aspect there is disclosed a liquid handling device for transferring liquid and handling liquids. The liquid handling device comprises an operating chamber arranged within a housing and a worktable area arranged in the operating chamber for receiving/arranging labware. As discussed previously the labware may comprise an at least one well plate, at least one pipette tip container containing at least one pipette tip and/or at least one dosing tool.
The luminescence tool may comprise at least one probe head with at least one light emitter for emitting a luminescent signal and at least one light receiver for detecting a luminescent signal, wherein the probe head is configured to be moved around in the operating chamber above the worktable area. The probe head can thus be moved to a sample in e.g. a well plate for luminescent registration.
Alternatively, the well plate could be moved to the probe head for the luminescent registration or both could be moved in order to arrange the probe head and the sample in correct relative position for luminescent registration.
Similar to the above, providing a luminescence tool that may be moved around the operating chamber of the liquid handling device removes the need for using a separate luminescent registration device and thus avoids unnecessary handling of and/or movement of the sample away from the worktable area of the liquid handling device.
In a further aspect there is disclosed a liquid handling system comprising the liquid handling device disclosed herein and at least one labware arranged on the worktable area.
The at least one labware can for example be an at least one well plate, at least one pipette tip container containing at least one pipette tip and at least one dosing tool.
As discussed, performing a luminescent registration directly in a sample provided in a well plate on the worktable of a liquid handling device using the luminescence tool removes the need of moving the sample away to a separate luminescent registration device.
Thus, by measuring directly in the well plate other peripheral and/or customized devices may be avoided. For example, using cartridges and microfluidic systems for luminescent registration may thus generally be avoided by the liquid handling devices, systems and methods described herein. In an even further aspect, a luminescence tool configured to be arranged above a worktable area, e.g. on a carrier arm, of a liquid handling device, is disclosed, wherein the luminescence tool comprises at least one probe head with a light emitter for emitting a luminescent signal and a light receiver for detecting a luminescent signal.
In one embodiment of the luminescence tool the tool is a fluorescence tool, and the probe unit in which the probe head may be integrated, measures the fluorescent intensity in dependency of different voltages applied on the probe. This is for example described in W02013/056820A1 and provides a very precise determination of the sample concentration.
After the detection of the fluorescent signal, the sample concentration can be calculated based on the fluorescent intensity of the standards.
In a further aspect a luminescent measuring shield system is disclosed comprising an at least one probe head with a light emitter for emitting a luminescent signal and a light receiver for detecting a luminescent signal, wherein the shield system further comprises a cover element enclosing the probe head, wherein the cover element comprises a barrier opaque to the light, such as the luminescent signal, and a cover opening provided in the barrier allowing light, such as the luminescent signal, to pass, wherein the probe head and the cover opening are movable relative to each other.
In one embodiment the probe head and the cover opening are movable relative to each other at least along the light emitting direction of the probe head.
This aspect provides a solution where the immersion depth of the light, e.g. the focal point of the light, in a sample can be controlled while reducing or even eliminating noise from ambient light during measuring.
Furthermore, the cover element will also facilitate cleaning, in particular in embodiments where the cover element completely covers the probe head as is disclosed further herein.
One embodiment of the luminescent measuring shield system may further provide a luminescence tool configured to be arranged above a worktable area, e.g. on a carrier arm, of a liquid handling device, wherein the luminescence tool comprises at least one probe head with a light emitter for emitting a luminescent signal and a light receiver for detecting a luminescent signal is disclosed wherein, the luminescence tool comprises a tool housing at least partly enclosing the probe head, and where the tool housing supports a probe unit comprising the probe head and where the probe unit is slidably arranged in the tool housing.
In one embodiment the luminescence tool may further comprise an electric circuit for communicating with the probe unit arranged on the tool frame for electrical coupling with a corresponding electrical coupling on a carrier arm. Similarly, the luminescence tool may comprise a mechanical coupling on the tool frame for mechanical coupling with a corresponding mechanical coupling interface in a carrier arm. The electrical and mechanical coupling may be separated or may be integrated into a common coupling interface providing both mechanical and electrical connection in the same coupling structure. In yet a further embodiment the coupling may for example be magnetic or a combination of magnetic, mechanical and/or electrical coupling.
In another aspect there is disclosed a method for using the luminescent shield system as discussed. The disclosed comprises a method for controlling the relative position, between a first relative position and a second relative position, of the probe head and the cover opening of the luminescent shield system as disclosed herein, and a distance between the cover opening and a well plate comprising at least one well comprising a sample, where the cover opening and the well plate are arranged so that the cover opening is placed between the well plate along the axis A - A of the luminescent shield system and the at least one probe head, and the at least one well opens towards the cover opening, wherein the method comprises
- maintaining a well distance between the cover opening and the well plate at a predetermined well distance,
- determining the level of the surface of a sample in the well,
- moving the probe head relative to the cover opening so that the distance between the probe head and the surface of the sample in the well is at a predetermined surface distance.
As will be understood applying the above method to the luminescent measuring shield system the risk of ambient light interfering with the measurement is reduced considerably, while at the same time it is possible to adapt the system to measure on individual sample volumes provided in the wells of a well plate. In an alternative aspect the method for using the luminescent shield system comprises moving the sample relative to the cover opening so that the distance between the probe head and the surface of the sample in the well is at a predetermined surface distance.
In one embodiment thereof, the predetermined surface distance is determined based on the focus length of the luminescent signal emitted from the probe head, the level of the surface of the sample in the well and a desired emergence depth of the luminescent signal emitted from the probe head, where the emergence depth is a distance from the surface of the sample to the focus point of the luminescent signal in the sample.
In some embodiments it has been found that an emergence depth of 3,5 mm is preferred. However, this may depend on the sample type, the dye used and other parameters. For example the emergence depth can be between 2 to 5 mm; 2 to 4 mm; 2 to 3,5 mm; 2 to 3 mm or 2 to 2,5 mm.
The level of the surface of a sample in the well can be determined in different ways. In one embodiment it is determined by detecting a change in the luminescent signal as the probe is moved relative to the sample or detecting a change in refraction in the light.
In another aspect there is disclosed a method for measuring the luminescence of a sample in a well plate arranged on a worktop area of a liquid handling device.
The method comprises steps of,
- arranging a probe head of a luminescence tool at the sample, where the luminescence tool comprising at least one probe head with a light emitter for emitting a luminescent signal and a light receiver for detecting a luminescent signal,
- exciting the sample by emitting the luminescent signal using the light emitter,
- detecting the subsequent luminescent signal emitted by the sample using the light receiver,
In one embodiment the method further comprises adding a luminescent dye to the sample. A standard solution may also be provided in one embodiment. Typically, the sample with the dyes and in relevant cases the standard solution is mixed and incubated for a certain time, which is determined by the specific application, sample, dye and/or the standard solution.
Having registered the luminescence of the samples it is possible to determine a normalization value of each sample. Accordingly, in one embodiment the method further comprises the steps of
- determining normalization volume, and
- normalizating the sample volume.
In one embodiment the user enters the values for the normalization before the luminescent registration is initiated. The user enters values for target concentration of normalized sample, target volume of normalized sample etc. Based on these predefined values and the determined sample concentration the system can calculate volumes which are required from the sample and/or the diluent to do the normalization.
Furthermore, in one aspect, the method for measuring the luminescence of a sample as disclosed herein may be computer implemented. In particular, the method for determining normalization of a sample based on a luminescent registration may be computer implemented.
As will be understood, one aspect herein discloses also using the liquid handling system, the liquid handling device and the luminescence tool as disclosed herein for measuring luminescence of a sample in a well plate. In particular, they are used for determining normalization of a sample based on a luminescent registration.
Detailed description
As discussed, the disclosure relates to a liquid handling device or a liquid handling system having an operating chamber wherein a luminescence tool can be moved around above a worktable to interact with different labware. The luminescence tool can for example be moved around by arranging the luminescence tool on a carrier arm.
In one embodiment the luminescence tool has a coupling interface for coupling and decoupling the luminescence tool to the carrier arm. As discussed herein such coupling may comprise both electrical and mechanical coupling. This allows for the liquid handling device to use different tools, for example from a selection of tools arranged on the worktable. Typical tools are different types of dosing tools such as dosing tools, for example one dosing tool provides a single channel pipetting tool, where another dosing tool provides an eight channel pipetting tool. By using different tools during liquid handling the device may in one situation be able to handle bulk pipetting actions by using the eight channel dosing tool, whereas individual sample handling can be done by using the single channel dosing tool.
By providing the luminescence tool which is able to couple and decouple from the carrier arm further flexibility is provided to the tool set of the liquid handling device.
Even further, in another embodiment, different luminescence tools could be provided. For example, similar to the dosing tools discussed above, one luminescence tool is provided with a single probe head for registering the luminescence for a single sample at a time, for example by measuring it in a well of the well plate. Another luminescence tool may comprise eight probe heads spaced such that they may register the luminescence from eight wells in a row.
Typically, the spacing of the probe heads would be nine millimeters as this is the standard distance between the wells in a 96-well plate. The person skilled in the art will understand the different types and sizes of well plates and dimensions thereof to which a luminescence tool as discussed herein may be configured and dimensioned. In particular, the person skilled in the part will look towards positions and dimensions as set out in recognized standards, for example the ANSI SLAS 4-2004 (R2012), provided by the Society for Laboratory Automation and Screening (SLAS) Microplate Standard Advisory Committee as directed by the American National Standards Institute (ANSI).
Accordingly, it is understood that embodiments may be provided wherein the one or more luminescence tools are provided comprising multiple probe heads, such as 8, 12, 96, 384 and/or 1536 probe heads.
In another embodiment it may be desirable to provide a liquid handling device wherein the luminescence tool is integrated in the carrier arm. This may be an advantage if the liquid handling device is designed for specific liquid handling processes where luminescent registration is frequently used, such as NGS (Next Gen Sequencing), where processes as normalization are frequently used. Thus, by integrating the luminescence tool in the carrier arm the step of switching between tools is removed, however, the carrier arm may become more bulky and the versatility of selecting between different luminescence tools is traded for a more dedicated system.
Although it has shown that design of the liquid handling device may be provided where cross talk and ambient noise interfering with the luminescent registration is negligent it may be desired to further ensure that such risks are reduced.
Accordingly, in one embodiment the at least one well plate comprises a material opaque to luminescent wavelengths, e.g. the wavelength of the light emitted by the light emitter of the probe head and/or the wavelength of the light detected by the light receiver of the probe head. Typically, dyed plastic, e.g. black, may be used for manufacturing the well plate in order to render it opaque to the luminescent wavelengths used.
In yet another embodiment the at least one pipette tip comprises a material opaque to luminescent wavelengths, e.g. the wavelength of the light emitted by the light emitter of the probe head and/or the wavelength of the light detected by the light receiver of the probe head.
In one embodiment the interface between the probe head and the well plate may be shielded when the luminescent registration is performed. For example, in one embodiment the probe head may be at least partly enclosed by a cover opaque to the luminescent wavelength, e.g. the wavelength of the light emitted by the light emitter of the probe head and/or the wavelength of the light detected by the light receiver of the probe head. This may provide a cover skirt which is configured to cover the well top when the probe head is arranged at or inserted into the well of the at least one well plate, thus reducing the risk of undesired luminescent illumination occurring during the luminescent registration.
In a further additional or alternative embodiment the housing of the liquid handling device may comprise a material opaque to luminescent wavelengths, e.g. the wavelength of the light emitted by the light emitter of the probe head and/or the wavelength of the light detected by the light receiver of the probe head, thus shielding the entire operating chamber from ambient luminescent illumination.
Luminescence is a generally well understood process. The phenomenon involves the absorption of light at a specific wavelength (emitted by the probe head) by a chemical molecule. This is also referred to the excitation wavelength. An emission of light subsequently occurs from the chemical molecule at a different wavelength (received by the probe head), typically a longer wavelength.
For example in one embodiment the luminescent wavelength of the signal emitted by the light emitter has a wavelength of 520 or 600nm. Such wavelengths are generally used in sequencing, e.g. DNA and RNA sequencing. It is well understood from the current disclosure that other wavelengths can be used for other applications.
In one embodiment the probe head is arranged such that it faces the worktable, e.g. so that it will face a sample in a well plate during use and when operated by the carrier arm.
For example, at least a proximal end of the probe head facing the worktable area during operation has a smaller cross-section than the cross section of a well of at least one well plate. This allows the probe head to be at least partly inserted into the well of a well plate to ensure that it is close to the sample, and also reduces the risk that ambient luminescent illumination interferes with the registration.
In one embodiment the probe head is arranged at a distance of 2 - 5 mm from the top opening of the well when the registration occurs.
As discussed, the luminescence tool may for example be configured to be arranged on the carrier arm or it may be integrated into the carrier arm.
For example, in one embodiment a first coupling interface is arranged on the fluorescence tool configured to couple and decouple with a corresponding second coupling interface on the carrier arm of the liquid handling device.
In an even further embodiment, the luminescence tool comprises a tool frame for supporting a probe unit comprising the probe head and an electric circuit for communicating with the probe unit and where a coupling interface is arranged on the tool frame for mechanical and electrical coupling with a corresponding coupling interface.
The electric circuit may in one embodiment comprise a dedicated system providing a protocol for communication between the luminescent sensor and the liquid handling device. In one embodiment the probe head is designed to emit and receive luminescent signals at specific wavelengths or wavelength bands. As discussed above the person skilled in the art will understand that the luminescent signals will be emitted and received as light at specific wavelengths, but in some embodiments it will be further processed as digital signals representing or containing information representing or relating to the wavelengths emitted or received by the probe head.
In another embodiment the luminescence tool may be provided as a kit where the probe head is configured to emit and receive luminescent signals within a desired wavelength band or at a specific wavelength. In one embodiment the probe head and probe unit itself is configured to emit and receive luminescent signals within a broad range of wavelengths, however, filter sets are provided and can arranged in the optical axis of the probe head to configure the system such that the desired wavelength, or wavelength band, emitted and received is registered. For example, a filter set may be provided for fluorescent registration where the emitted wavelength is 520 or 600 nm.
The probe unit can in one embodiment be slidably arranged in the tool frame. This allows for minor adjustment of the probe head relative to the sample in order to optimize the luminescent registration.
In relation to the luminescent measuring shield system (also referred to more briefly as ‘shield system’ herein) further embodiment thereof may be provided.
For example, in one embodiment the probe head has an annular shape and the cover opening has an annular shape.
Arranging the cover opening and the probe head may be done in several ways, for example the probe head and the cover opening may be co-axially arranged along the axis A - A.
In one embodiment, the system may be designed such that the probe head and the cover opening are movable relative to each other along the axis A - A. In particular the probe head and cover opening may be limited to movement relative to each other along the axis A - A, this avoids any undesired movement in other directions than along the axis A - A.
In some embodiments an optical axis of the probe head corresponds to the axis A - A. The cover opening and the probe head may in some embodiment be annular and the radius of the cover opening and the radius of the probe head may be the same.
Different considerations should be taken into account in order to further improve the shield system. E.g. in one embodiment the shield system further comprises a well plate, comprising at least one well, wherein the well plate is arranged so that the cover opening is arranged between the well plate and the at least one probe head and the well opens towards the cover opening.
In such embodiment a first distance between the cover opening and the at least one well in the first relative position of the system may be the same as a second distance between the cover opening and the at least one well in the second relative position of the system. Or in other words, the position of the cover opening is maintained relative to the well, while the probe head may be moved relative to the well.
In one embodiment it is ensured that the first and the second distance are maintained in order to further obtain comparable measurements.
In order to further shield from ambient light at least a part of the barrier facing the well plate may in one embodiment have a cover width, transverse to the axis A - A, that corresponds to at least three times the width of the well.
In a further embodiment at least a part of the barrier facing the well plate may have a cover length, transverse to the axis A - A and perpendicular to the width, the corresponds to at least three times of the length of the well.
The cover opening may in some embodiments be arranged in the center of the part of the barrier facing the well plate having the cover width and/or the cover length.
The luminescent registration as described herein is particular useful for normalizing sample, e.g. when performing sequencing such as NGS . Sample normalization is well known and refers to adjusting the sample volume or concentration prior to or after data acquisition in order to be able to compare the sample.
With regards to luminescent registration the challenge has been that this has been done away from the automated liquid handling device and without proper integration the data had to be read out separately, the normalization calculated and programmed in to the liquid handling devices which result in a long cumbersome process which also has a high risk of error as each input is done manually with the risk of errors when entering data.
By providing the fluorescence detection in the automated liquid handling devices as disclosed the normalization can be automated as the data can be relayed and processed internally in the system without external input from e.g. external data sources or manual input from the user.
Description of the drawings
In the following, embodiments and examples will be described in greater detail with reference to the accompanying drawings:
Fig. 1 shows an embodiment of an automated liquid handling device suitable for use with a luminescence tool as disclosed herein.
Fig. 2a and 2b shows one embodiment of a fluorescence tool as disclosed herein, and
Fig. 3a and 3b shows the above fluorescence tool during fluorescent registration of a sample in a well plate.
Detailed description of the drawings
Fig. 1 shows an automated liquid handling device 100 having a housing 101 defining an operating chamber 102 wherein a worktable 103 is arranged. The worktable is configured to receive a plurality of labware (not shown), such as well plates, mixers, heaters etc. in the designated areas 104.
Within the operating chamber there is also arranged a carrier arm 105. The carrier is moveable in the x (width), y (depth) and z (height) direction within the volume of the operating chamber.
In the current embodiment this is enabled by providing a rail system comprising a first rail arm 106 on the back of the housing 101 which extends along the width of the operating chamber, indicated by the x-direction. Transverse to the first rail arm 106 there is a second rail arm 107 which extends transversely to the first rail arm in the depth of the operating chamber, indicated by the y-direction. The second rail arm is arranged on the first rail arm so that it can be controlled and moved in the width direction. A third rail arm 108 is slidable arranged on the second rail arm 107. The third rail arm extends in the height direction, indicated by the z-direction. The third rail arm 108 is arranged on the second rail arm so that it can be controlled and moved in the depth direction along the second rail arm.
The carrier arm 105 is arranged on the third rail arm 108 whereon it can be moved in the height direction.
This provides full operability of the carrier arm 105 within the operating chamber in all three dimensions. The rail system 106, 107, 108 described for operating the carrier arm 105 arrangement is generally known and one example of enabling movement of the carrier arm within the operating chamber. Other construction may be provided by the person skilled in the art, e.g. a robotic arm.
The carrier arm is configured to releasable engage different tools (not shown) via a coupling structure (e.g. coupling structure 105’ in Fig.1 ). Such tools could for example be a single or an eight channel dosing tool. The carrier arm is also configured to releasable engage a fluorescence tool as discussed herein. The worktable further comprises a tool holding section 109 in which the different tools may be placed when not in use. The system may then move to the tool holding section and switch between tools during operation as instructed.
The automated liquid handling device 100 further comprises a waste basket 110 arranged in the worktable where the carrier arm may dispense of used products, in particular used pipettes. In order to gain access to the operating chamber and worktable the front 114 of the housing 101 may be opened.
A terminal 115, comprising a monitor 111 , a keyboard 112 and a mouse 113 is linked to the automated liquid handling device where an operator can program the device, monitor it during operation and/or interfere in a running program if necessary. The terminal may be linked to the automated liquid handling device in a wired manner or wirelessly. Other types of terminals could be considered, e.g. a laptop, a smartphone or a tablet.
Fig. 2a and 2b shows an embodiment of a fluorescence tool 200. The fluorescence tool comprises a housing 201 , where part of the housing has been opened to show a probe unit 202, comprising a probe head 203. The probe unit is slidable arranged in the housing along a focusing direction 204 between a first position 205 as shown in Fig. 2a and a second position 206 as shown in Fig. 2b. The focusing direction is parallel to the optical axis A - A of the probe unit. In current embodiment the optical axis can be considered to be the rotational symmetry axis of the light emitted from the probe head. Said light comprising a wavelength or wavelength band desired to excite a fluorescent dye.
The probe unit 202 is fixed on a mounting plate 207, where the mounting plate is provided with first and a second longitudinal extending grooves 208, 209, which extend parallel to each other. First and second pins 210, 211 are provided in the housing and arranged to guide each groove respectively, i.e. the first pin is received in the first groove and the second pin is received in the second groove.
By providing two groove and pin configurations as discussed, the probe unit and the housing are locked relative to each other only allowing relative movement along the longitudinal extent of the grooves. In the current embodiment the pins of the housing are arranged on a line parallel/along the focus direction, which results in the grooves being arranged so that the longitudinal extent is along the focus direction. Accordingly, this provides a configuration where the movement of the probe unit relative to the housing is limited by the longitudinal grooves in the focus direction of the fluorescence tool in Fig. 2a and 2b.
The housing 201 defines a barrier part 212 that is opaque to ambient light and encloses the probe head. The barrier part is furthermore formed of a material having a low reflectance, i.e., the material will absorb or disperse the light so the risk of ambient light being reflected into the sample or the probe head is minimized or removed.
An opening 213 which allows for light to pass is arranged coaxially with the optical axis in the proximal end 214 of the housing. The proximal end of the housing is the end which is closest to the well plate during measurement. Thus, as will be shown light from the probe head may pass through the opening and into the well plate and light emitted from the sample may pass through the opening and be received by the probe head.
The opening 213 is closed by a glass element 219 which allows light to pass. The glass element works as a physical barrier together with the rest of the housing so that foreign object does not enter into the housing and contaminates or blocks the probe head or otherwise compromises the function of the luminescence tool. This also facilitates cleaning of the fluorescence tool 200.
In order to activate the movement of the probe unit within the housing a shaft 215 is connected to the probe unit (or the mounting plate 207) at a connection end 216 and forms part of a spindle drive at a spindle end (not shown).
The housing 201 is at its distal end 217, i.e. opposite the proximal end, provided with a coupling interface 218 which is configured to couple with a matching coupling part provided on a carrier arm (not shown in the current figures, but shown in one embodiment in Fig. 1 as coupling structure 105’).
Also, though not shown, the probe unit is powered and communicates with a processor via electrical couplings and an electrical coupling interface provided in the coupling interface in order to power, control and read data from the probe unit.
Figures 3a and 3b illustrates, in section, the fluorescence tool 200 in use in the first position 205 in Fig. 3a as it measures a relative small sample 300 in a well 301 of a well plate 302 and in the second position 206 in Fig. 3b as it measure a relative large sample 310 in a well 31 1 of a well plate 312.
In both positions it can be seen that the proximal end 214 of the housing is maintained at the same distance relative to the well plate. The distance d between the surface 220 of the proximal end 214 and the top 303, 313 of the well plates 302, 312 is so small that ambient light does not affect the measurement.
In other word, during operation the proximal end of the housing functions as a shield, reducing or removing ambient light from entering the well wherein a sample is measured and/or from reaching the probe head.
As can be understood, because the sample size is different in the two wells of Figs. 3a and 3b it is possible to control the focus point, ‘p’, of the emitted light 320 emitted by the probe unit 202 and thus the desired immersion depth of the emitted light in the sample can be controlled by simultaneous minimizing the impact of environmental light on the fluorescence measurement since the proximal end 214 functions as a barrier preventing ambient light to interfere. In one embodiment the focus distance f of the emitted light 320, i.e.. the distance from the probe head to the focus point p’ is 18 mm, and the moving distance ‘m’, ie. the relative distance travelled between the probe unit and the housing 201 when moving from the first position to the second position or vice versa is 5,5 mm. This allows for a system where an immersions depth , ie. the distance from the surface of the sample in the well and the focus point of the emitted light, can be maintained at 3,5 at the first and second positions and all the positions in between.
Reference numbers automated liquid handling device 100 housing 101 operating chamber 102 worktable 103 designated areas 104 carrier arm 105 coupling structure 105’ first rail arm 106 second rail arm 107 third rail arm 108 tool holding section 109 waste basket 110 front 114 of the housing 101 terminal 1 15 comprising a monitor 111 a keyboard 112 mouse 113 fluorescence tool 200 housing 201 probe unit 202 probe head 203 focusing direction 204 first position 205 second position 206 mounting plate 207 first longitudinal extending groove 208 second longitudinal extending groove 209 first pin 210 second pin 211 barrier part 212 opening 213 proximal end 214 of the housing 201 glass element 219 shaft 215 connection end 216 distal end 217 of the housing coupling interface 218 surface 220 of the proximal end 214 small sample 300 well 301 ; 311 well plate 302; 312 large sample 310 top 303; 313 of the well plates 302; 312 emitted light 320
Embodiments
1. A liquid handling device for transferring liquid and handling liquids, comprising an operating chamber arranged within a housing, a worktable area arranged in the operating chamber for receiving/arranging labware, wherein the labware comprises an at least one well plate, at least one pipette tip container containing at least one pipette tip and at least one dosing tool, wherein the operating chamber further comprises
- a carrier arm for moving tools between labware arranged on the worktable area, and
- a luminescence tool comprising at least one probe head with at least one light emitter for emitting a luminescent signal and at least one light receiver for detecting a luminescent signal, wherein the luminescence tool is configured to be arranged on the carrier arm.
2. The liquid handling device according to embodiment 1 , wherein the luminescence tool has a coupling interface for coupling and decoupling the luminescence tool to the carrier arm.
3. The liquid handling device according to embodiment 1 , wherein the luminescence tool is integrated in the carrier arm.
4. The liquid handling device according to embodiment 1 , 2 or 3, wherein the at least one well plate comprises a material opaque to luminescent wavelengths.
5. The liquid handling device according to any one of the embodiments 1 - 4, wherein the at least one pipette tip comprises a material opaque to luminescent wavelengths.
6. The liquid handling device according to any one of the embodiments 1 - 5, wherein the probe head is at least partly enclosed/surrounded/encircled by a skirt/cover/shield opaque to luminescent wavelength.
7. The liquid handling device according to embodiment 6, wherein the skirt is configured to cover the well top when the probe head is arranged at or inserted into the well of the at least one well plate.
8. The liquid handling device according to any one of the preceding embodiments, wherein the housing comprises a material opaque to luminescent wavelengths. 9. The liquid handling device according to any one of the preceding embodiments, wherein luminescent wavelength emitted by the light emitter has a wavelength of 520 or 600nm.
10. The liquid handling device according to anyone of the preceding embodiments, wherein the luminescence tool comprises multiple probe heads, such as 8, 12, 96, 384 and/or 1536 probe heads.
11 . The liquid handling device according to any one of the embodiments 1 - 10, wherein the at least one light emits a fluorescent or phosphorescent signal and the at least one light receiver receives a fluorescent or phosphorescent signal.
12. The liquid handling device according to any one of the embodiments 1 - 11 , wherein the probe head faces the worktable when operated by the carrier arm.
13. The liquid handling device according to embodiment 12, wherein at least a proximal end of the probe head facing the worktable area during operation has a smaller crosssection than the cross section of a well of at least one well plate.
14. The liquid handling device according to anyone of the preceding embodiments, wherein the luminescence tool comprises a tool frame for supporting a probe unit comprising the probe head and an electric circuit for communicating with the probe unit and where a coupling interface is arranged on the tool frame for mechanical and electrical coupling with a corresponding coupling interface.
15. The liquid handling device according to embodiment 14, wherein the probe unit is slidably arranged in the tool frame.
16. A liquid handling device for transferring liquid and handling liquids, comprising an operating chamber arranged within a housing, a worktable area arranged in the operating chamber for receiving/arranging labware, wherein the labware comprises as at least one well plate, at least one pipette tip container containing at least one pipette tip and at least one dosing tool, wherein the operating chamber further comprises
- a luminescence tool comprising at least one probe head with at least one light emitter for emitting a luminescent signal and at least one light receiver for detecting a luminescent signal, wherein the probe head is configured to be moved around in the operating chamber above the worktable area. 17. A liquid handling system, comprising the liquid handling device according to any one of the embodiments 1 - 16 and at least one labware arranged on the worktable area.
18. A liquid handling system according to embodiment 17, wherein the at least one labware comprises at least one well plate, at least one pipette tip container containing at least one pipette tip and at least one dosing tool.
19. A liquid handling system according to any one of the preceding embodiments, wherein when operating the system, the probe head is arranged in a distance of 2 - 5 mm from the top opening of the well.
20. A luminescence tool configured to be arranged above a worktable area, e.g. on a carrier arm, of a liquid handling device, wherein the luminescence tool comprises at least one probe head with a light emitter for emitting a luminescent signal and a light receiver for detecting a luminescent signal.
21. The luminescence tool according to embodiment 20, wherein the luminescence tool is configured to be arranged on the carrier arm.
22. The luminescence tool according to embodiment 21 , wherein the luminescence tool is integrated into the carrier arm.
23. The luminescence tool according to embodiment 20 or 21 , wherein a first coupling interface is arranged on the luminescent tool configured to couple and decouple with a corresponding second coupling interface on the carrier arm of the liquid handling device.
24. The luminescence tool according to anyone of the embodiments 20, 21 , 22 or 23, wherein the luminescence tool comprises a tool frame for supporting a probe unit comprising the probe head and an electric circuit for communicating with the probe unit and where a coupling interface is arranged on the tool frame for mechanical and electrical coupling with a corresponding coupling interface.
25. The luminescence tool according to embodiment 24, wherein the probe unit is slidably arranged in the tool frame.
26. A method for measuring the luminescent of a sample in a well plate arranged on a worktop area of a liquid handling device, wherein the method comprises the steps of, - arranging a probe head of a luminescence tool at the sample, where the luminescence tool comprising at least one probe head with a light emitter for emitting a luminescent signal and a light receiver for detecting a luminescent signal,
- exciting the sample by emitting the luminescent signal using the light emitter, and
- detecting/recording the subsequent luminescent signal emitted by the sample using the light receiver.
27. A method according to embodiment 26, wherein the method further comprises the steps of
- determining normalization volume
- normalization the sample volume.
28. A computer implemented method for determining normalization of a sample based on luminescent measurement according to embodiment 26 or 27.
29. Using the liquid handling device according to any one of the embodiments 1 - 16, the liquid handling system according to any one of the embodiments 17 - 19 or the luminescence tool according to any one of the embodiments 20 - 25 for measuring luminescence of a sample in a well plate.
30. A luminescent measuring shield system comprising an at least one probe head with a light emitter for emitting a luminescent signal and a light receiver for detecting a luminescent signal, wherein the shield system further comprises a cover element enclosing the probe head, wherein the cover element comprises a barrier opaque to the light and a cover opening provided in the barrier allowing light to pass along an axis A - A, wherein the probe head and the cover opening are movable relative to each other between a first relative position and a second relative position.
31. The luminescent measuring shield system according to embodiment 30, wherein the luminescent measuring shield system comprises a luminescence tool configured to be arranged above a worktable area, e.g. on a carrier arm, of a liquid handling device, wherein the luminescence tool comprises at least one probe head with a light emitter for emitting a luminescent signal and a light receiver for detecting a luminescent signal is disclosed wherein, the luminescence tool comprises a tool housing, wherein the cover element and the cover opening is provided so that the tool housing at least partly encloses the probe head, and where the tool housing supports a probe unit comprising the probe head and where the probe unit is slidably arranged in the tool housing between the first relative position and the second relative position.
32. The luminescent measuring shield system according to embodiment 31 , wherein the lluminescence tool further comprise an electric circuit for communicating with the probe unit and/or a coupling interface is arranged on the tool frame for mechanical and electrical coupling with a corresponding coupling interface.
33. The system according to any one of the embodiments 30, 31 or 32, wherein the probe head has an annular shape and the cover opening has an annular shape.
34. The system according to any one of the embodiments 30 - 33, wherein the probe head and the cover opening are co-axially arranged along the axis A - A.
35. The system according to any one of the embodiments 30 - 34, wherein the probe head and the cover opening are movable relative to each other along the axis A - A.
36. The system according to embodiment 35, wherein the probe head and cover opening are limited to movement relative to each other along the axis A - A.
37. The system according to any one of the embodiments 30 - 36, wherein an optical axis of the probe head corresponds to the axis A - A.
38. The system according to any one of the embodiments 33 - 37, wherein the cover opening and the probe head are annular and the radius of the cover opening and the radius of the probe head are the same.
39. The system according to any one of the embodiments 30 - 38, wherein the system further comprises a well plate, comprising at least one well, wherein the well plate is arranged so that the cover opening is arranged between the well plate and the at least one probe head and the well opens towards the cover opening.
40. The system according to embodiment 39, wherein a first distance between the cover opening and the at least one well in the first relative position of the system is the same as a second distance between the cover opening and the at least one well in the second relative position of the system. 41 . The system according to embodiment 39 or 40, wherein at least a part of the barrier facing the well plate has a cover width, transverse to the axis A - A, that corresponds to at least three times the width of the well.
42. The system according to embodiment 41 , wherein at least a part of the barrier facing the well plate has a cover length, transverse to the axis A - A and perpendicular to the width, that corresponds to at least three times of the length of the well.
43. The system according to embodiment 41 or 42, wherein the cover opening is arranged in the center of the part of the barrier facing the well plate having the cover width and/or the cover length.
44. A method for controlling the relative position, between a first relative position and a second relative position, of the probe head and the cover opening of the luminescent shield system according to any one of the embodiments 30 - 43, and a distance between the cover opening and a well plate comprising at least one well comprising a sample, where the cover opening and the well plate is arranged so that the cover opening is placed between the well plate along the axis A - A, and the at least one probe head and the at least one well opens towards the cover opening, wherein the method comprises
- maintaining a well distance between the cover opening and the well plate at a predetermine well distance,
- determining the level of the surface of a sample in the well,
- moving the probe head relative to the cover opening so that the distance between the probe head and the surface of the sample in the well is at a predetermined surface distance.
45. The method according to embodiment 44, wherein the predetermined surface distance is determined based on the focus length of the luminescent signal emitted from the probe head, the level of the surface of the sample in the well and a desired emergence depth of the luminescent signal emitted from the probe head, where the emergence depth is a distance from the surface of the sample to the focus point of the luminescent signal in the sample.
46. The method according to embodiment 45, wherein the emergence depth is 3,5 mm.