Method and system for applying a liquid sample onto a substrate for image analysis

Abstract

The present disclosure relates to an automated method for applying a liquid sample (1) onto a substrate (2) for image analysis as well as an automated system (100) for performing the automated method. The automated method comprises providing data corresponding to a property of the liquid sample (1) from a property determination unit (20) to a workflow management unit (90); selecting, by the workflow management unit (90), sample preparation operations based on the property of the liquid sample (1); setting, by the workflow management unit (90), operational parameters based on the selected sample preparation operations and / or on the property of the liquid sample (1); controlling a sample preparation unit (10) to prepare the liquid sample (1) for image analysis by performing the sample preparation operations selected by the workflow management unit (90) and by using the operational parameters set by the workflow management unit (90).

Description

METHOD AND SYSTEM FOR APPLYING A LIQUID SAMPLE ONTO A SUBSTRATE FOR IMAGE ANALYSISTECHNICAL FIELD

[0001] The present disclosure relates to an automated method and an automated system for applying a liquid sample onto a substrate for image analysis. The present disclosure further relates to a computer program product comprising instructions to cause the automated system to execute the automated method. The automated method, automated system and computer program product according to the present disclosure may be used in the field of hematology. However, other fields of application are feasible.BACKGROUND

[0002] There is a general trend towards automating sample preparation and sample analysis in the field of hematology. For example, the Complete Blood Count (CBC), which is one of the most frequently performed panels of laboratory tests in hematology, can be assessed by automated devices, such as flow cytometers using impedance, fluorescence, and / or scattered light-based measurement techniques. Such devices allow determining a number of properties of a biological sample, e.g. red blood cell (RBC) count, white blood cell (WBC) count, platelet count, hemoglobin (Hb), hematocrit (Het), etc. In addition, they allow characterizing morphological features of RBCs, WBCs, and platelets by indirect measurements, based on light absorption and light scattering techniques and / or cytochemically based measurements.

[0003] Whenever an abnormality is detected in a biological sample by flow cytometric analysis, which may be an indication for a certain disease (for example an abnormally high WBC count or WBCs suspicious for infection), it is a standard procedure to further investigate the sample in order to complete the assessment of the morphological components of the CBC. In a typical hematology workflow, the in-vitro diagnostic (IVD) system flags the biological sample for which an abnormality has been detected, in order to indicate to a laboratory technician that further investigation is required. It is still common to perform such further assessments manually, which involves placing a portion of the biological sample on a substrate, for example a microscope slide, smearing the sample against the substrate to form a wedge smear, drying and staining the sample, and placing the sample under a microscope for visual examination. This challenging manualprocess is cost-intensive and time-consuming and often needs to be done by skilled laboratory technicians. Advancements have been made in the past years to automate this procedure. Currently, various products are available on the market that are designed to apply a liquid sample onto a microscope slide by an automated wedge preparation technique. The major disadvantage of wedge smear preparation however is that it yields samples with highly variable thicknesses and distribution of blood constituents. The wedge smear often has only a single narrow band with an appropriate cell density for examination and analysis. The location and shape of this band varies from slide to slide. In addition, these products often are stand-alone units, the consequence being that the samples have to be carried manually from the automated sample preparation unit to a microscope for visual inspection. US 2008 / 0050511 Al discloses an automated slide-making apparatus and a method for processing biological samples by quantizing a residue in a tube left behind by a blood sample that was advanced through said tube and - in response to said quantization - controlling the motion profile of a blood-smearing member of the automated slidemaking apparatus.

[0004] Other automated sample preparation methods and systems have been developed to overcome the disadvantages of wedge smears in order to produce a uniform, high-quality specimen to make visual evaluation of the sample both easier and more accurate. For example, US 10,764,538 discloses a system and a method utilizing an improved technique for applying a single layer of sample cells onto a substrate and generating a uniform distribution of cells on the substrate and further discloses an integrated digital microscope for automated visual inspection. The system makes use of an applicator instead of a spreading member to deposit the sample on the substrate. Physical characteristics of the sample material, for example viscosity or consistency, have an influence on how the sample fluidically behaves while being dispensed through the applicator onto the substrate and how it distributes over the substrate once dispensed thereon. The system of US 10,764,538 uses a predetermined optimized sample flow rate through the applicator tip in combination with a predetermined relative applicator speed and height of the tip above the substrate. However, sample characteristics such as viscosity, consistency, and density of constituents can vary from sample to sample, and become particularly prominent between different types of sample material. This poses a challenge to systems intended to process different types of sample materials, for example whole blood, serum, plasma, and other bodily fluids, such as urine, cerebrospinal fluid (CSF), bone marrow, etc. The system of US 10,764,538 proposes to analyze an image of a sample and to determine based on the number of counted cells in the image whether the sample requires further dilution or concentration. If so, the system would either dilute orconcentrate a portion of the sample followed by a sample re-test. In any event, the system would need to perform the entire sample preparation workflow on the same sample again. This reduces throughput of the system and increases the time to result, which ultimately presents an economical burden to the laboratories.GENERAL DESCRIPTION

[0005] It is against the above background that aspects of the invention according to the present disclosure provide certain unobvious advantages and advancements over the prior art. In particular, a need was recognized for an improved method and system for preparing liquid samples for image analysis.

[0006] Although the aspects of the invention according to the present disclosure are not limited to specific advantages or functionality, it is noted that certain aspects of the present disclosure may allow for an automated method and system for applying a liquid sample onto a substrate to generate an improved specimen for subsequent image analysis. In particular, certain aspects of the present disclosure may enable the formation of a monolayer of sample particles on the substrate while ensuring an adequate number of particles and a uniform distribution of particles on the substrate, for different liquid samples with different physical properties.

[0007] Certain aspects of the present disclosure may allow for an improved method of applying a liquid sample onto a substrate by selecting sample preparation operations and by setting operational parameters based on previously determined properties of each individual liquid sample. The method and system according to certain aspects of the present disclosure may further provide a simple and cost-effective technique to determine a property of the liquid sample. Other aspects of the present disclosure may provide a sample processing workflow that is faster and less resource-intensive as compared to solutions known in the art. Furthermore, the method and system according to certain aspects may enable a sample preparation procedure adaptable to a plurality of different sample materials.

[0008] In particular, the present disclosure relates to an automated method for applying a liquid sample onto a substrate for image analysis. The automated method comprises providing data corresponding to a property of the liquid sample from a property determination unit to a workflow management unit. The automated method further comprises selecting, by the workflow management unit, sample preparation operations based on the property of the liquid sample. Theautomated method further comprises setting, by the workflow management unit, operational parameters based on the selected sample preparation operations and / or on the property of the liquid sample. The automated method further comprises controlling a sample preparation unit to prepare the liquid sample by performing the sample preparation operations selected by the workflow management unit and by using the operational parameters set by the workflow management unit, wherein the selected sample preparation operations at least comprise the application of the liquid sample onto the substrate by controlling an applicator to move translationally in relation to the substrate while dispensing the liquid sample through the applicator onto the substrate, so that particles in the liquid sample settle onto the substrate.

[0009] The term “liquid sample” as used herein is a broad term and may refer to any liquid material that can be subjected to a diagnostic procedure, e.g. an in-vitro diagnostic (IVD) analysis, in order to determine a physical, chemical, physiological or immunological property thereof, or to detect one or more analytes of interest suspected to be present therein, e.g. polypeptides (like antigens, antibodies), nucleic acid, electrolytes, cells, cell surface proteins, coagulation factors, and the like. The term may refer to a biological sample, a quality control solution, a calibrator solution, a reference solution, or mixtures of one or more biological samples with one or more reagents and / or diluents or mixtures of a quality control solution or a calibration solution with one or more reagents and / or diluents. The liquid sample may be derived from any biological source, such as a physiological fluid, including blood, cerebrospinal fluid (CSF), urine, synovial fluid, peritoneal fluid, pleural fluid, tissue, bone marrow, or the like. A liquid sample may be used directly as obtained from the source or used following a pretreatment to modify the character of the sample, such as preparing plasma from blood, diluting viscous fluids, lysis or the like. Methods of treatment can involve filtration, distillation, concentration, centrifugation, inactivation of interfering components, and the addition of reagents. An initially solid or semi -solid biological material, e.g. tissue, may be rendered liquid by dissolving or suspending it with a suitable liquid medium. According to certain aspects of the present disclosure, the liquid sample may be a blood sample, such as whole blood, plasma, or serum. Alternatively and according to further aspects of the present disclosure, the liquid sample may be a body fluid sample, such as cerebrospinal fluid, synovial fluid, pleural fluid, ascites fluid, or bone marrow aspirate.

[0010] The term “reagent” as used herein may be any kind of liquid solution containing a reactant, typically a chemical compound or agent capable of binding to or chemically transforming one or more structures of sample constituents or analytes suspected in the liquid sample. Together with a corresponding detection method, a reagent enables detection of these structures or analytes.Examples of reactants are enzymes, enzyme substrates, dyes, conjugated dyes, protein-binding molecules, nucleic acid binding molecules, antibodies, chelating agents, promoters, inhibitors, epitopes, antigens, and the like. In particular, a reagent may comprise a dye or a mixture of dyes, where a mixture of dyes is commonly also referred to as a stain.

[0011] The term “dye” as used herein may refer to a detectable moiety that is suitable for staining cellular structures in a liquid sample for e.g. brightfield microscopy or fluorescence microscopy. Examples of dyes are chromogenic dyes, metallographic dyes, chromophorecontaining dyes, fluorescent dyes, phosphorescent dyes and nanomaterials (such as quantum dots). The term “stain” may refer to a solution comprising a plurality of dyes. A well-known example is the Romanowsky-type stain, which is a metachromatic stain useful for staining cytology samples, wherein the stain comprises a cationic thiazine dye (such as polychrome methylene blue, azure A, azure B, azure C, azure IV, sym- dimethylthionine, thionine, methylene violet Bemsthen, methylthionoline, toluidine blue, and combinations thereof) and an anionic halogenated fluorescein dye (such as eosin A, eosin Y, eosin G, and combinations thereof). Other examples include Malachowski stain, Giemsa stain, May-Gruenwald stain, May-Gruenwald-Giemsa (MGG) stain, Jenner stain, Wright stain, Leishman stain, and DIFF-QUICK (proprietary modified Wright stain).

[0012] The term “substrate” as used herein may refer to any type of support material with properties that allow a liquid sample to be applied thereon and that allow the liquid sample to be imaged and / or analyzed, e.g. through a microscope. Typically, a substrate has a planar receiving surface onto which the liquid sample can be applied. Substrates can include, but are not limited to, glass or plastic microscope slides, polished or mirrored ceramic slides, polished metal substrates, plastic or other flexible specimen films, and / or any other reflective materials capable of supporting a sample. The substrate may be a plate exhibiting a rectangular or circular form.

[0013] The term “property” as used herein with respect to a liquid sample may refer to any physical, chemical, physiological or immunological characteristic of the liquid sample. Examples include the sample type, number and / or type of particles in the sample, sample viscosity, sample density, sample color, turbidity, pH, etc., one or a plurality of which may be determined by a dedicated functional unit, i.e. a property determination unit. In particular and according to an aspect of the present disclosure, the property of the liquid sample comprises any or a combination of a liquid sample type, hematocrit, hematocrit range, sample viscosity, sample density or a particle count. Examples of possible sample types have already been mentioned above.

[0014] The term “hematocrit” as used herein may refer to the volume percentage (vol%) of red blood cells (RBCs) in blood. There are multiple methods to determine the hematocrit, some of them performed by fully automated devices. Most automated methods are indirect determination methods, where the hematocrit is derived, i.e. calculated, from other measurement parameters. For example, when applying a measurement method using the coulter principle, the number of RBCs in a known volume of a blood sample is determined as well as the mean cell volume of the RBCs. The RBC count is then multiplied by the mean cell volume and put in relation to the known sample volume to arrive at the hematocrit as volume percentage. An alternative method comprises estimating the hematocrit by a calculation using the measured hemoglobin amount. The term “hematocrit range” as used herein may refer to a predetermined range of hematocrit levels. For example, hematocrit ranges may correspond to well-known reference ranges that are defined for the use in laboratory diagnostics. One or more hematocrit ranges may be defined. For example, a “normal” hematocrit range may comprise all hematocrit levels considered being “normal” in a medical context, e.g. between 35% and 51%. A “low” hematocrit range may be determined comprising hematocrit levels below 35%, and a “high” hematocrit range may be determined comprising hematocrit levels above 51%. Hematocrit ranges may be determined based on the patient’s gender, e.g. a “normal range” for women may be between 36% and 45%, and a “normal range” for men between 40% and 51%. Further ranges may be determined with a higher resolution. Hematocrit ranges may be determined and implemented by the manufacturer or by the operator.

[0015] “Sample viscosity” as used herein may refer generally to a measure of a liquid sample’s resistance to deformation at a given rate. It quantifies the internal frictional force between adjacent layers of fluid that are in relative motion. Different factors can influence a liquid sample’s viscosity. One example is sample temperature. Another example is the number of constituents contained in a liquid sample. For example, a blood sample contains constituents, such as cells (for example RBCs, white blood cells (WBCs), platelets) or molecules (e.g. proteins, electrolytes, carbohydrates, fatty acids). Of these constituents, RBCs have the greatest effect on the blood sample’s viscosity. For example, the relative viscosity of blood may be four times higher than that of water. In a bone marrow aspirate, viscosity may be affected by, for example, the amount of present hematopoietic cells, adipose tissue (adipocyte), and supportive stromal cells. In yet another example, a cerebrospinal fluid sample may contain small numbers of WBCs, carbohydrates, electrolytes and proteins, affecting the sample’s viscosity. The viscosity of a liquid sample may be measured, for example, by a viscometer or a rheometer.

[0016] The term “sample density” may refer to the liquid sample’s mass per unit of volume, which can -similar to the sample viscosity- be affected by the liquid sample’s constituents. For example, according to Kenner, T. “The measurement of blood density and its meaning. ” Basic research in cardiology vol. 84,2 (1989): 111-24., blood density is proportional to hematocrit or to the total protein concentration of blood.

[0017] The term “particle count” refers to the number of a given type of particles in the liquid sample. Typically, the number of particles is reported in relation to a defined volume of the liquid sample. The term “particle” may thereby refer to any kind of particle in or constituent of the liquid sample. Particles in a blood sample may be any constituents smaller than 50 pm, according to an example smaller than 30 pm, according to another example smaller than 20 pm, such as cellular components like RBCs, WBCs, platelets, bacteria, parasitic protozoa, etc., or other structures, such as artifacts. In a clinical setting, the number of RBCs and / or WBCs may be determined in a predetermined volume of, for example, a patient’s blood or urine sample, in order to find possible irregularities, which indicate a potential medical condition. In another example, a cerebrospinal fluid sample may be analyzed by counting the number of WBCs contained therein, where an elevated WBC count may, for example, indicate inflammation or infection of the brain or spinal cord. The particle count may be determined by conventional methods, for example an impedancebased flow cytometer, using the coulter principle, or a (digital) microscope.

[0018] The term “unit” as used herein may refer to a functional entity that is dedicated to executing a specific set of operations, typically as part of a workflow. Accordingly, the unit is constructed and configured in a way to perform such operations optimally. It may be operated autonomously as a stand-alone apparatus, in cooperation with other units or as a sub -unit / module in a more complex apparatus. In the field of IVD, a unit may therefore refer to a standalone IVD analyzer, it may be connected to or work in cooperation with one or more other IVD analyzer(s), or it may refer to a part or module within an IVD analyzer or IVD system. For example, a unit may be a conveyor for conveying sample containers within a diagnostic device or between diagnostic devices. It may be a gripper, a centrifuge, a pipetting unit, an incubation unit, an analytical (measurement) unit, an imaging unit, a result-displaying unit, etc.

[0019] The term “property determination unit” as used herein is a broad term and may refer to any kind of apparatus, module or functional unit configured to determine a property or multiple properties of a liquid sample automatically. The property determination unit may be designed as stand-alone device that is communicatively connected to other devices via a network, for example via an intranet or internet network. Alternatively and preferably, the property determination unitmay be a module or a functional unit in a cluster of functional units forming a larger device, for example an IVD device, and may be communicatively connected to other functional units within that larger device. The property determination unit may be configured to use a certain measurement method to determine the property of the liquid sample. According to certain aspects of the present disclosure, providing data corresponding to a property of the liquid sample comprises controlling the property determination unit to determine the property of the liquid sample by any of an impedance measurement or a conductivity measurement or a resistance measurement or an electrochemical measurement or an optical measurement or a pressure measurement or a viscosity measurement or combinations thereof. For example, the property determination unit may be an impedance-based flow cytometer, using the coulter principle, to determine certain properties of a liquid sample. The coulter principle is well known in the art and will therefore not be elaborated in further detail. In another example, the property determination unit may be a blood gas analyzer, capable of performing conductivity measurements or electro-chemical measurements to determine a property of a liquid sample. Conductivity is the ability of a solution to transmit (conduct) electricity. The electrical current will increase in proportion to the number of ions (or charged particles) found in a solution, their electrical charge and their mobility (i.e. how easily the ions can move in the solution). In an electro-chemical measurement, a chemical reaction is translated into an electrical signal, which can then be measured. In another example, the property determination unit may be a photometer or a spectrophotometer, measuring transmission or absorption of light through a liquid sample, possibly with different wavelengths. According to an aspect of the present disclosure, the optical measurement is a light scatter measurement, for example in a flow cell, or an interferometric measurement. For example, the property determination unit may be a laserbased flow cytometer, measuring forward -scattered light (FSC) signals, side-scattered light (SSC) signals, and / or dye-specific fluorescence signals. Alternatively, the flow cytometer may be based on laser-interferometry, as for example described in Zhao, Y. et al., Self-Mixing Interferometry- Based Micro Flow Cytometry System for Label-Free Cells Classification. Appl. Sci. 2020, 10, 478. https: / / doi.org / 10.3390 / appl0020478. In yet another example, the property determination unit may be a viscometer or a rheometer. Typical viscometers used in the art are capillary viscometers or rotational viscometers. Another technique for determining viscosity of a liquid is by measuring the velocity of a magnetic sphere inside the liquid sample that is moved back and forth by a magnetic field. In a further example and according to an aspect of the present disclosure, the pressure measurement comprises controlling a pipetting probe to aspirate a portion of the liquid sample from a sample container. While aspirating the liquid sample, the method comprisesmeasuring pressure inside the pipetting probe by a pressure sensor operatively coupled to the pipetting probe. The method further comprises correlating the measured pressure with a hematocrit range and storing the measured pressure and correlated hematocrit range as property of the liquid sample in a data storage. Examples of hematocrit ranges to which the measured pressure may be correlated are described below. Alternatively, and according to another aspect of the present disclosure, the method may comprise correlating the measured pressure with any of a sample viscosity range, a sample density range or a particle count range. The method may further comprise storing the measured pressure and correlated viscosity range, sample density rang or particle count range, respectively, as property of the liquid sample in a data storage. Examples of respective ranges to which the measured pressure may be correlated are described below.

[0020] The term “pipetting probe” as used herein is a broad term and may refer to a functional unit configured to perform pipetting operations. For example, the pipetting probe may be configured to aspirate a liquid sample or a predetermined portion thereof from a sample container. Typically, a pipetting probe comprises a tubular section with an inner channel leading into at least one opening at or near a tip of the probe and at least one other opening at the opposite end of the tubular section. The pipetting probe further comprises a pump that is fluidically connected to the tubular section and allows for creating a negative pressure inside the inner channel to aspirate liquids into the inner channel. In addition or alternatively, the pump allows for creating a positive pressure inside the channel to dispense any present liquids out of the inner channel. The tubular section may be tapered at least partially, for example at the tip. The tubular section of the pipetting probe may be designed as a hollow needle with a sharpened tip to facilitate penetration through a lid of a closed sample container from where the liquid sample then is aspirated. The tubular section of the pipetting probe may be operated with a multi-use tip that requires to be cleaned regularly in order to prevent cross-contamination. Alternatively, it may be operated with single-use tips that are replaced between pipetting operations of different liquid samples.

[0021] The term “pressure sensor” as used herein may refer to any type of sensor configured to measure pressure within a certain vessel or compartment. According to an aspect of the present disclosure, the pressure sensor is operatively coupled to the pipetting probe, which enables measuring the pressure inside the pipetting probe. The pressure measurement may be performed while a liquid sample is being aspirated into the pipetting probe and / or while the liquid sample is being dispensed. The pressure measurement may therefore be conducted and recorded as a continuous measurement over time, for example from the beginning to the end of the aspiration operation, resulting in a pressure profile. Alternatively, it may be conducted and recorded once ormultiple times at predetermined time intervals during the aspiration operation. For example, pressure may be measured at the beginning of the aspiration operation and again at the end of the aspiration operation. Alternatively, pressure may be measured at intervals in a range anywhere between 0.05 seconds and 1 second. For example, a pressure measurement may be conducted and recorded every 0.1 seconds, every 0.2 seconds, or every 0.5 seconds. Various types of pressure sensing techniques are known in the art. The pressure sensor may be a resistive, a capacitive, a piezoelectric sensor, a sensor based on the MEMS technology (micro electro-mechanical system), or the like. The recorded pressure profile or pressure measurements may be compared to a reference profile or reference values. Depending on the deviation of the recorded pressure profile or pressure measurements from the reference profile or reference values, a calculation regarding certain properties of the liquid sample may be conducted, for example regarding the viscosity, the density, or the hematocrit of the liquid sample. In this manner and according to an aspect of the present disclosure, the measured pressure can be correlated to a hematocrit or hematocrit range. For example, while aspirating a liquid sample from a closed sample container, a negative pressure is created inside the pipetting probe in order to pull the liquid sample into the pipetting probe. If a liquid sample has a high hematocrit, and consequently a high viscosity, as compared to a liquid sample with a normal hematocrit, the negative pressure needs to be reduced even further, in order to aspirate the same volume of liquid sample in the same period of time. A pressure measurement inside the pipetting probe may detect such a variation in negative pressure.

[0022] The measurement data generated by the property determination unit are typically stored electronically, for example in a data storage. The term “data storage” as used herein is any kind of storage medium configured for digitally storing data or information, such as a computer memory, a network storage or a cloud storage. The property determination unit may therefore comprise a data storage or it may be communicatively connected to a remote data storage, such as a computer network or a cloud. The data “corresponding to a property of a liquid sample” may include any raw data and / or processed data that in any way reflect a property of said liquid sample, where examples of liquid sample properties have already been described above. Data processing may comprise a conversion step. For example, depending on the measurement method, the measurement parameters may be converted from an analog signal into a digital signal, for example by an analog-to-digital converter (ADC), before the data can be processed and / or stored. In another example, data may be processed by converting measurement values into other units, for example from g / L into mg / dL. In yet another example, data may be processed by stratification, for example by correlating or grouping one or a set of measurement results into predefined categories.Examples of a predefined categories are hematocrit ranges, such as a “normal” hematocrit range, which may comprise all hematocrit measurements between 35% and 51%, or a “high” hematocrit range comprising hematocrit levels above 51%, etc. (s. also above for more examples). As mentioned above, the property determination unit may be communicatively connected to other devices or functional units, for example a workflow management unit, via a direct connection, wired or wirelessly, or indirectly over a communications network to enable transmission or exchange of data or via other data management units, such as computers. Communication means and communication protocols for transferring data between two or more digital entities are well known in the art and will therefore not be further elaborated herein.

[0023] The term “workflow management unit” as used herein is a broad term and may refer to a hardware and / or software implemented system configured for managing workflows of one or more functional units that are in operational interaction with each other, for example within an IVD device or across a plurality of IVD devices. A workflow may comprise a sequence of operational steps, which may need to be synchronized across different functional units. The workflow management unit may comprise a programmable logic controller or processor running a computer-readable program provided with instructions to perform operational steps in accordance with the methods described herein. The workflow management unit may be integrated into an IVD device, it may be integrated into a unit, a sub-unit or a module of an IVD device, or it may be a separate logic entity in communication with the IVD device or its units, sub-units or modules via a direct connection, wired or wirelessly, or indirectly over a communications network, wired or wirelessly, such as a wide area network, e.g. the Internet or a Health Care Provider's local area network or intranet, via a network interface device. In some aspects, the workflow management unit may be integral with a controller or data management unit, e.g. implemented on a computing device such as a desktop computer, a laptop, a smartphone, a tablet, personal digital assistant (PDA), etc., or it may be comprised by a server computer and / or be distributed / shared across / between a plurality of IVD devices. For example, the workflow management unit may be configured to determine the processing order for a batch of liquid samples, possibly based on their respective test orders. In another example, the workflow management unit may instruct a functional unit, such as a conveyor, to transport a sample container containing a liquid sample to a specific position within the IVD device, and instruct a second functional unit, such as a pipetting probe, to aspirate the liquid sample or a portion thereof out of the sample container. The workflow management unit may therefore synchronize the operational steps performed by the conveyor with the operational steps performed by the pipetting probe for the sake of workflow efficiency and inorder to prevent certain risks, for example a collision between the pipetting probe and the sample container.

[0024] According to the present disclosure, the workflow management unit selects sample preparation operations based on the property of the liquid sample. The term “sample preparation operations” as used herein may refer to any kind of operational steps that are conducted in order to prepare a liquid sample for a following analytical assessment or diagnostic test. Examples of sample preparation operations are the dilution of a liquid sample, concentration of a liquid sample, mixing, dispersing, aliquoting, transfer of the liquid sample, e.g. from a sample container into a reaction vessel or from a sample container onto a substrate, staining of the liquid sample, etc. For the processing of each liquid sample, the workflow management unit typically selects a series of sample preparation operations, where the selected sample preparation operations may at least partially depend on the property of the liquid sample. In particular, sample preparation operations may be selected based on whether the data or data point corresponding to the property of the liquid sample falls within a predetermined range, lies above or below a predetermined threshold, or corresponds to a predetermined category or a reference value. For example, if the property of the liquid sample refers to an abnormally high hematocrit level, which may be defined as being above a predetermined upper threshold, for example above 55%, or falling into a predetermined range, for example between 55% and 60%, the workflow management unit may select the following sample preparation operations: dilution of the liquid sample, application of the diluted liquid sample onto the substrate, staining of the diluted liquid sample and fixation of the diluted liquid sample. According to an alternative aspect, the same sample preparation operations may be selected if the property corresponding to the liquid sample refers to a high sample viscosity, sample density or particle count. In another example, if the particle count of the liquid sample is low, which may be defined as a particle count below a predetermined lower threshold or falling into a predetermined range, for example if the number of erythrocytes is below 3.5 per pL or if the number of erythrocytes is between 2.5 and 3.5 per pL, the workflow management unit may select the following sample preparation operations: concentration of the liquid sample, for example by centrifuging the liquid sample and pipetting off the supernatant, application of the concentrated liquid sample onto the substrate, staining of the concentrated liquid sample and fixation of the concentrated liquid sample. According to an alternative aspect, the same sample preparation operations may be selected if the property corresponding to the liquid sample refers to a low hematocrit, hematocrit range, sample viscosity, or sample density. In yet another example, the workflow management unit may select sample preparation operations for applying the liquidsample onto the substrate and for staining and / or fixating the liquid sample on the substrate, if the property of the liquid sample is a hematocrit, a sample viscosity, a sample density or a particle count in a normal range. The selected sample preparation operations are then transmitted by the workflow management unit as instruction to the sample preparation unit to perform said selection of sample preparation operations.

[0025] According to the present disclosure, the workflow management unit sets operational parameters based on the selected sample preparation operations and / or the property of the liquid sample. The term “operational parameters” as used herein may refer to any kind of operational parameter according to which any operational step, in particular a sample preparation operation, is conducted. The operational parameters set by the workflow management unit may be transmitted as instructions to the sample preparation unit, which performs sample preparation operations using these operational parameters. Examples of operational parameters include rotational speed of a centrifuge and / or time of centrifugation, volume of a diluent to be pipetted to a liquid sample, distance between an applicator and a substrate, type and / or volume of stain to be used for staining, etc. In devices known in the art, operational parameters might be fixed, i.e. they might be predetermined by the manufacturer and therefore not adjustable during operation of the device. In other devices, operational parameters might be manually set by the device operator. The operational parameters according to the present disclosure however are automatically and dynamically adjustable based on variable factors, for example a property of the liquid sample. Liquid sample properties, examples of which are outlined above in more detail, have a great influence on the uniformity of liquid sample distribution on the substrate, which is why it is advantageous if the sample preparation operations and the corresponding operational parameters can flexibly be selected and adjusted.

[0026] According to aspects of the present disclosure, the operational parameters comprise at least one parameter chosen from the group of a distance between the applicator and the substrate, and / or a movement pattern of the applicator relative to the substrate, and / or a speed pattern of applicator relative to substrate movement, and / or a liquid sample dispensing volume, which is a portion of the aspirated liquid sample, and / or a dispensation rate, and / or a distance between rows of liquid sample on the substrate.

[0027] The term “distance between applicator and substrate” as used herein refers to the distance between the tip of the applicator and the substrate during the dispensing procedure of the liquid sample and has an influence on the uniformity of liquid sample distribution on the substrate, because of capillary forces within sample on the substrate contiguous with the sample beingdispensed from the applicator. The sample preparation unit is therefore configured to adjust the distance between applicator and substrate. For example, if the substrate is placed in the sample preparation unit in a horizontal orientation, the applicator may be movable in a vertical direction relative to the substrate, for example by a step motor or a pneumatic actuator. The distance between the applicator and substrate may be determined in a range between 8 to 20 pm (e.g., 10, 12, 14, 16, or 18 pm), depending on the property of the liquid sample. For example, if the liquid sample has a high hematocrit, for example a hematocrit above 52%, or alternatively a high sample viscosity, a high sample density or a high particle count, the workflow management unit may determine the distance between applicator and substrate at a value in the range between 8 to 12 pm, for example 10 pm, to enable uniform settling of particles onto the substrate. In another example, if the liquid sample is measured with a low viscosity, e.g. lower than 3 centipoise (cP), or alternatively with a low hematocrit, or a low sample density or a low particle count, the workflow management unit may determine the distance between applicator and substrate at a value in the range between 16 to 20 pm, for example 18 pm, to enable higher volumes of liquid sample being applied onto the substrate. Once the distance between the applicator and substrate is determined, the distance is maintained nearly constant during the entire dispensing procedure of the liquid sample, e.g. varying by no more than 2 pm.

[0028] The term “movement pattern” as used herein refers to the direction of movement of the applicator relative to the substrate over a period of time, e.g. during dispensation of the liquid sample. The skilled person will recognize that the movement pattern therefore also determines the settling pattern of the liquid sample, and consequently any particles included therein, on the substrate. According to an aspect of the present disclosure, the method comprises controlling the substrate and / or the applicator to move translationally in relation to each other. The applicator may therefore be movably coupled to guiding rails that enable a translational movement of the applicator in an x-y Cartesian plane relative to the substrate, while the substrate is held in a fixed position. Alternatively, the applicator may be installed in a fixed position and the substrate may be movable in an x-y Cartesian plane relative to the applicator. In another alternative, both the applicator and the substrate are movable in an x-y Cartesian plane relative to each other. The movement pattern is selected so that sample particles are distributed as uniformly as possible on the substrate. For example, the liquid sample may be dispensed onto the substrate in a linear alignment, forming a row. It may be dispensed in multiple rows next to each other. The multiple rows next to each other may be created in a singular continuous dispensation step, for example to form a boustrophedon pattern. Therefore, the applicator may be positioned e.g. at the lower leftcorner of the dispensation area on the substrate. As the liquid sample is dispensed through the applicator tip, the applicator is moved in x-direction to the lower right corner of the dispensation area on the substrate to create a first row. From there, while still dispensing liquid sample, the applicator is moved in y-direction for a predetermined distance, i.e. to create a distance between the rows, and then moved back in negative x-direction to the left side of the dispensation area on the substrate to create a second row next to the first. The applicator is moved in y-direction again with the same distance as before and moved in x-direction to the right side of the dispensation area on the substrate to create a third row next to the second. This procedure may be continued until the entire dispensation area is covered with liquid sample or until a predetermined liquid sample dispensing volume has been dispensed, resulting in a meander-shaped dispensation pattern. In another example, the liquid sample may be dispensed in concentric circles or in a spiral pattern.

[0029] The term “distance between rows” as used herein refers to a parameter that determines how far apart dispensation rows lie from each other in case the movement pattern contains multiple dispensation rows. In the example described above, the distance between rows is determined by the parameter for movement in y-direction, which may be in a range between 10 and 1000 pm, e.g. in a range between 30 and 800 pm, or according to another example in a range between 50 and 500 pm.

[0030] The term “liquid sample dispensing volume” as used herein refers to the volume of liquid sample to be dispensed in its entirety onto the substrate, particularly into the dispensation area on the substrate. Typically, a liquid sample is provided in a sample container, from which only a portion or aliquot is required for further processing. According to an aspect of the present method, the liquid sample dispensing volume is determined at a value between 0.1 and 10 pL. According to another aspect, the liquid sample dispensing volume is determined at a value between 0.3 and 8 pL. According to yet another aspect, a liquid sample dispensing volume is determined at a value between 0.5 and 1.5 pL. According to yet another aspect, a liquid sample dispensing volume is determined at about 1 pL.

[0031] The term “speed pattern” as used herein refers to the velocity, and adjustments thereof, of the applicator relative to the substrate during dispensation of the liquid sample. During the dispensation process, the velocity may be held constant or it may vary. For example and with regard to the boustrophedon movement pattern mentioned above, the applicator may be moved relative to the substrate in x-direction at a certain velocity. When approaching either side of the dispensation are on the substrate, velocity of applicator relative to substrate may be continuously decreased to ensure a smooth transition from movement in x-direction to a movement in y-direction, thereby preventing possible splashes of liquid sample, which might occur if applicator movement is stopped to abruptly. The speed pattern may therefore include variables such as the velocity of applicator relative to substrate in x-direction, velocity of applicator relative to substrate in y-direction, but also the rate of decelaration and / or acceleration.

[0032] The term “dispensation rate” as used herein refers to the volume of liquid sample dispensed out of the applicator tip per unit of time. The dispensation rate may be determined in relation to the applicator speed relative to the substrate or the speed pattern. For example, the dispensation rate may be determined at 0.1 pL per second while the applicator tip is moved at a speed of 30 mm per second over the substrate. According to an aspect of the present method, the dispensation rate may be set at a value between 0.01 pL / s and 1 pL / s, in particular at a value between 0.03 pL / s and 0.8 pL / s, more particularly at a value between 0.05 pL / s and 0.5 pL / s.

[0033] A “sample preparation unit” as used in the present disclosure is a broad term and may refer to any kind of apparatus, module or functional unit configured for automatically preparing a liquid sample for a following analytical assessment or diagnostic test. The sample preparation unit may be designed as a stand-alone device that may be in operational and / or communicative connection with other devices. Alternatively, the term may comprise a plurality of stand-alone devices that are in operational and communicative connection with each another and possibly with other devices, e.g. an analytical device. Alternatively and preferably, the sample preparation unit may be comprised in an IVD device, where the sample preparation unit may comprise one or more modules or functional units. The sample preparation unit may be configured for performing any kind or a combination of sample preparation operations. According to the present disclosure, the sample preparation unit prepares the liquid sample by performing the sample preparation operations selected by the workflow management unit and by using the operational parameters set by the workflow management unit, wherein the sample preparation operations comprise applying the liquid sample onto the substrate by controlling an applicator to translate in relation to the substrate while dispensing the liquid sample through the applicator onto the substrate, so that particles in the liquid sample settle onto the substrate, staining the liquid sample on the substrate and / or fixating the liquid sample on the substrate. The sample preparation unit therefore comprises an applicator for dispensing the liquid sample onto the substrate. The applicator may be a hollow and needle- or syringe-shaped element with an opening at the tip of the applicator. The applicator may be operationally connected to a pump, for example a peristaltic pump or a syringe pump that allows the dispensation of small volumes of liquid sample through the tip of the applicator onto the substrate. According to an aspect of the present disclosure, the tip may have an outside diameterof 2 to 5 mm and an inner diameter of about 0.5 mm. According to another aspect, the tip may have an outside diameter of 0.5 to 1.5 mm and an inner diameter of 0.1 to 0.45 mm. The tip may be disposable or washable. The tip may be rounded to facilitate insertion and cleaning of the tip. Translating the applicator in relation to the substrate may refer to moving the applicator translationally relative to the substrate, moving the substrate translationally relative to the applicator or moving both the applicator and the substrate translationally relative to each other. The sample preparation unit may further comprise a staining unit for staining the liquid sample on the substrate. The staining unit is configured for applying a stain or a dye onto the liquid sample on the substrate and may therefore comprise one or more dispensers, where the dispenser may be, for example, a peristaltic pump. Examples of compatible stains are Romanowsky stains, reticulocyte stains, stains using specific antibodies, hematoxylin and eosin, immunocytochemical stains, histochemical stains for viewing cellular components, and / or antibody, aptamer or other stains based on binding a ligand to an antigen (further examples of stains are mentioned above). The stain may be mixed with a diluent before being applied onto the substrate. Diluents useful for diluting whole blood for example, may include salt solutions or protein solutions. Salt solutions range from “physiological saline” (0.9N) to complex mixtures of salts. Protein solutions can range from simple solutions of bovine albumin to a commercial preparation with selected human plasma proteins. Additionally or alternatively, the sample preparation unit may comprise a fixating unit for fixating the liquid sample on the substrate. Fixating the liquid sample may comprise exposing the liquid sample on the substrate to a fixative, e.g. 85% methanol, and / or to a buffer solution. For some stains, an ethyl alcohol or formaldehyde-based fixative might be used. The fixating unit may therefore comprise a dispenser for dispensing the respective fixative to the liquid sample. An example of a staining unit and a fixating unit is disclosed in US 8,454,908. Alternatively, the substrate carrying the liquid sample may be immersed into one or more baths of the staining and fixing solutions. In another alternative, staining and / or fixing solutions may be moved across the substrate using capillary action.

[0034] According to an aspect of the present disclosure, the automated method further comprises controlling an imaging unit to generate a digital microscope image of one or more particles on the substrate, analyzing the one or more particles in the digital microscope image to generate an analytical result for the liquid sample, and displaying the data corresponding to the property of the liquid sample and / or the analytical result on a display device.

[0035] The term “imaging unit” as used herein refers to any kind of apparatus, module or functional unit configured to generate a digital microscope image of one or more particles on thesubstrate. The imaging unit may be designed as a stand-alone device that may be in operational and / or communicative connection with other devices, e.g. via an intranet or internet network. Alternatively and preferably, the imaging unit may be a module or a functional unit in a cluster of functional units forming a larger device, for example an IVD device, and may be connected operationally and communicatively to other functional units within that larger device. In order to generate a digital microscope image, the imaging unit may comprise a light emission device, a microscope and a light receiving device. A light emission device as referred to herein may be any kind of device configured for emitting light in any of the visible spectral range, the infrared spectral range or the ultraviolet spectral range in order to illuminate the liquid sample on the substrate. The light emission device may be configured for emitting light having a single wavelength or may be configured for simultaneously emitting light having different wavelengths. The light emission device may, for example, comprise a white light source or other multispectrum light sources such as a halogen bulb, florescent bulb, or incandescent bulb, etc. The light emission device may further comprise a filter, for example, in form of a filter wheel, in order to filter the multispectrum light into a single wavelength or a narrow band of wavelengths. Alternatively or additionally, the light emission device may comprise one or more lasers or light emitting diodes (LEDs). Lasers and LEDs typically produce narrow bandwidths of illumination. An advantage of using narrow -band illumination rather than broad-band illumination is that it increases the sharpness of the images generated by the light receiving device. A microscope as referred to herein may be any kind of device comprising at least one optical element configured for receiving light generated by the liquid sample on the substrate in response to illumination and / or for receiving light transmitted through the liquid sample on the substrate and for focusing the impinging light rays to produce an image. The microscope may comprise at least one lens, e.g. at least one imaging lens and / or at least one objective lens. The microscope may comprise a plurality of lenses such as a lens system. The microscope may be configured for projecting, e.g. in combination with the lens system, an image of the liquid sample on the substrate or a magnified image of areas of the liquid sample on the substrate to the light receiving device. The magnification of the microscope may range from 4x to lOOx. For example, the magnification may be lOx, or 20x, or 50x. The microscope may comprise at least one zoom lens and / or at least one zoom lens system. Examples for a microscope are a bright field microscope, fluorescence microscope, phase contrast microscope, spectral microscope, dark field microscope, Fourier ptychographic microscope, etc. A light receiving device as used herein may be any kind of device configured for receiving rays of light for recording or capturing optical data or information and for conveying said optical data or information to forma digital image. The light receiving device may therefore comprise a camera comprising an image sensor. As an example, the imaging sensor may be a CCD chip and / or a CMOS chip. The generated digital microscope image of the one or more particles on the substrate may then be stored in a data storage.

[0036] The digital microscope image may be transmitted to or accessed by a functional unit, e.g. an image processor, which is configured for analyzing the one or more particles in the digital microscope image to generate an analytical result for the liquid sample. The term “analyzing” as used herein may thereby comprise identifying, i.e. in the sense of locating, one or more particles of interest within the digital microscope image, possibly determining coordinates of the particles’ location in the digital microscope image or their location on the substrate, and identifying the type of the one or more particles, for example, based on certain characteristics the particles display in the digital microscope image. In order to perform this analysis in an automated way, the controller is provided with respective computer-readable instructions, e.g. image analysis algorithms. Examples of image analysis algorithms are described in US 7,689,038, US 7,881,532, US 2023028525A1, and WO2023118586A1. For example, in the field of hematology, particles of interest may be cellular components of blood, i.e. RBCs, WBCs, and platelets. When executed by the image processor, one or more image analysis algorithms enable the image processor to locate and differentiate RBCs, WBCs, and platelets from other particles in or from the background of the digital microscope image. They further enable the controller to identify the specific cell type, i.e. RBC, WBC, or platelet, and to further differentiate cell sub-types based on certain morphological features displayed in the digital microscope image. For example, sub-types of WBCs may include neutrophils, lymphocytes, atypical lymphocytes, hematopoietic progenitor cells, monocytes, immature granulocytes; granulocytes including basophils, eosinophils, neutrophils such as segmented neutrophils and band neutrophils, mast cells, blasts, promyelocytes, metamyelocytes, myelocytes, white blood cells with inclusions like Auer rods or Doehle bodies, etc. Sub-types of RBCs may include normal erythrocytes, nucleated red blood cells, polychromatic cells, reticulocytes, immature reticulocytes, schistocytes, spherocytes, stomatocytes, red blood cells with inclusions like Howell-Jolly bodies, Heinz bodies, Pappenheimer bodies, red blood cells with parasitic inclusions like plasmodia or babesia, etc.

[0037] The term “generating an analytical result” as used herein refers to using the data or information generated in the image analysis procedure by the imaging unit and based thereon creating an analytical result. An analytical result may therefore comprise any or a combination of one or more numerical values, any kind of descriptive information, or any kind of visualinformation regarding the analytes of interest in the tested liquid sample. For example, a Complete Blood Count (CBC) is one of the most frequently performed panel of laboratory tests in the field of hematology. An analytical result of a CBC typically comprises an RBC count, a WBC count, a platelet count, a hemoglobin concentration, a determination of the hematocrit, and determinations of the mean corpuscular volume (MCV), the mean corpuscular hemoglobin concentration (MCHC), the mean corpuscular hemoglobin (MCH), and the red blood cell distribution width (RDW). In another example, the analytical result of a white blood cell differential, which is another frequently performed laboratory test, may comprise an absolute count and relative percentage of WBC sub-types, such as neutrophils, lymphocytes, monocytes, eosinophils, and basophils. In another example, the analytical result may comprise the digital microscope images generated by the imaging unit and possibly processed by an imaging algorithm to make visible to a human observer certain features in the digital microscope images. In yet another example, the analytical result may include a measure of fluorescence, e.g. color and / or intensity, emitted by analytes in the liquid sample. Other examples of analytical results are a number and / or relative percentage of nucleated red blood cells (NRBC), mean platelet volume (MPV), number and / or relative percentage of reticulocytes, or a mean reticulocyte hemoglobin content. In many IVD devices known in the art, analytical results will be presented to a health-care professional, such as laboratory personnel or a physician, on a display device. The “display device” may therefore be any kind of device configured for displaying data or information in a human-readable manner. An example of displaying analytical results, in particular of images of cells, is disclosed in US 11,047,791. According to an aspect of the present disclosure, the automated method comprises a method step of displaying the data corresponding to the property of the liquid sample and / or the analytical result on the display device. For example, for a given liquid sample, a measure for sample viscosity and an analytical result of a CBC may be displayed. In another example, a particle count determined by the property determination unit may be displayed next to the analytical result of a CBC generated based on the digital microscope image, which comprises counts for RBCs, WBCs, and platelets.

[0038] According to an aspect of the present disclosure, the automated method further comprises generating an output value using the analytical result and the property of the liquid sample, which may allow for providing more accurate and reliable analytical results. The property of the liquid sample and the analytical result may refer to the same parameter, for example a hematocrit of the liquid sample. In such an event, the method comprises comparing the parameter measured by the property determination unit with the parameter determined by analysis of themicroscope image. This allows for verifying or validating the parameter determined by the property determination unit with the analytical result generated by the controller based on the microscope image, or vice versa. The “output value” may be generated by a calculation operation using the analytical result and the property of the liquid sample. For example, the output value may result in a subtraction of the analytical result from the property of the liquid sample, or vice versa, or the output value may refer to a relative difference between the two values. Alternatively, the output value may be generated by comparing a value calculated using the analytical result and the property of the liquid sample with a threshold or reference value. Alternatively, the output value may refer to a qualitative statement, for example indicating that the analytical result and the property of the liquid sample are the same or adequately similar. In another example, the output value may indicate that the two results are significantly different and that a quality issue in the sample preparation procedure may exist. An alert or warning message may be issued and displayed to inform a device operator of the quality issue.

[0039] According to aspects of the present disclosure, the operational parameters comprise a dilution parameter and / or a concentration parameter and the sample preparation operations further comprise a dilution or concentration of the liquid sample according to the dilution parameter or concentration parameter, respectively, before the application of the liquid sample onto the substrate. A “dilution parameter” as used herein may refer to any kind of settings, in particular adjustable settings, related to the sample preparation operation of diluting a liquid sample. Typical operational steps in a dilution operation may be, for example, to generate an aliquot of the liquid sample with a predetermined volume by pipetting said volume of liquid sample into a sample vessel. Another operational step may include pipetting a predetermined volume of diluent to the aliquot of the liquid sample. Yet another operational step may include mixing or vortexing of the diluted liquid sample. The dilution parameter may therefore comprise any or a combination of a volume of liquid sample, a volume of diluent, a ratio between volume of liquid sample and volume of diluent, a number of aliquots to be generated, a measure for mixing intensity, e.g. operational speed of a mixer, a time (duration) of mixing, a measure for vortexing intensity, etc. For example, the dilution parameter may comprise a volume ratio of liquid sample to diluent of 1 : 1, which indicates that a predetermined volume of the liquid sample is diluted in an equal volume of diluent. Hence, the dilution parameter may comprise a volume ratio of liquid sample to diluent anywhere in the range of 1 :0.05 to 1 :20 (liquid sample: diluent). For example, the dilution parameter may comprise a volume ratio between 1 :0.1 to 1 : 10, or it may comprise a volume ratio between 1 :0.5 to 1:8. According to a method step of the present disclosure, which is described in further detailabove, operational parameters are set by the workflow management unit based on the selected sample preparation operations and / or on the property of the liquid sample. For example, if the property of the liquid sample refers to a highly elevated hematocrit level, e.g. above a predetermined threshold, the workflow management unit may select sample preparation operations including a dilution of the liquid sample. The workflow management unit then sets the operational parameters, where the operational parameters include the dilution parameter. Since the property of the liquid sample refers to a highly elevated hematocrit level, the workflow management unit may set the dilution parameter in a manner that enables a high dilution, for example, comprising a volume ratio of 1 :5 (liquid sample: diluent), and possibly also comprising a high mixing intensity for mixing the diluted liquid sample. In another example, where the property of the liquid sample may refer to a moderately elevated hematocrit level, for example, between a predetermined lower and upper threshold, the workflow management unit may set the dilution parameter in a manner that enables a lower dilution than in the previous example, for example, comprising a volume ratio of 1 : 1 (liquid sample: diluent), and possibly comprising a low mixing intensity for mixing the diluted liquid sample.

[0040] A “concentration parameter” as used herein may refer to any kind of settings, in particular adjustable settings, related to the sample preparation operation of concentrating a liquid sample. “Concentrating” in the context of this disclosure refers to the process of bringing an analyte or particle of interest that is present in a solution at a certain concentration to a state of higher concentration, e.g. by removing a liquid part of the solution that does not contain the analyte or particle of interest. Typical operational steps in a concentration operation may be, for example, to generate an aliquot of the liquid sample with a predetermined volume by pipetting said volume of liquid sample into a sample vessel. Another operational step may include rotating or centrifuging the sample vessel containing the liquid sample so that particles of interest settle on the bottom of the sample vessel. Yet another operational step may include removing a supernatant, for example, by an automated pipette. Yet another operational step may include mixing the concentrated liquid sample. The concentration parameter may therefore comprise any or a combination of a volume of liquid sample for generating an aliquot, preferably in a range between 1 pL to 1000 pL, a number of aliquots to be generated, a rotational speed of a centrifuge or a sample vessel rotator, a time (duration) of centrifugation, a volume of supernatant to be removed, a mixing intensity, etc. For example, if the property of the liquid sample refers to a parameter that is below a predetermined threshold or in a range that is considered a low range, for example, if the particle count of the liquid sample is low, the workflow management unit may select samplepreparation operations including a concentration of the liquid sample. The workflow management unit then sets the operational parameters, where the operational parameters include the concentration parameter. For example, the workflow management unit may set the concentration parameter so that it comprises a rotational speed for a centrifuge of about 1500 to 2000 revolutions per minute (rpm) for a duration of 10 to 15 minutes, and a volume of supernatant to be removed after centrifugation, e.g. 500 pL. In another example, where the property of the liquid sample refers to a parameter that is moderately decreased, for example, a moderately decreased hematocrit level, the workflow management unit may set the concentration parameter so that it comprises a rotational speed for a centrifuge of about 1000 to 1500 rpm, and a volume of supernatant to be removed after centrifugation of 250 pL.

[0041] The present disclosure further refers to an automated system for preparing a liquid sample for image analysis according to the automated method described above. The automated system comprises a sample preparation unit comprising: an applicator for dispensing the liquid sample onto a substrate; and a staining unit for staining the liquid sample on the substrate and / or a fixating unit for fixating the liquid sample on the substrate. The automated system further comprises a workflow management unit configured to: receive data corresponding to a property of the liquid sample from a property determination unit; select sample preparation operations based on the property of the liquid sample; set operational parameters based on the selected sample preparation operations and / or on the property of the liquid sample, control the sample preparation unit to prepare the liquid sample by performing the sample preparation operations selected by the workflow management unit and by using the operational parameters set by the workflow management unit, wherein the sample preparation operations comprise the application of the liquid sample onto the substrate by controlling the applicator to move translationally in relation to the substrate while dispensing the liquid sample through the applicator onto the substrate, so that particles in the liquid sample settle onto the substrate.

[0042] The term “automated system” as used herein may refer to any kind of automated pre- analytical, analytical device or combination thereof. An analytical device is configured to obtain an analytical measurement value or analytical result from a patient’s sample in vitro in order to provide information on the health status of the patient. The analytical measurement values or analytical results can be qualitative, semi -quantitative and / or quantitative measures of analytes or particles of interest. It is designed to automatically conduct a set of processing operations optimized for the respective type of analysis, e.g. hematology analysis, coagulation analysis, clinical chemistry, immunochemistry, and can include operations such as pipetting, incubating,transporting, mixing, heating, cooling, measuring, detecting, cleaning, etc. The automated system may be or may comprise an IVD device. A pre-analytical device is configured to prepare the liquid samples or the sample containers holding the liquid samples in a way that they can be processed by an analytical device afterwards. This can include processing steps such as loading / unloading of sample containers and / or consumables, decapping, preliminary checks of the sample quality, filllevel checks, pipetting, aliquoting, centrifuging, diluting, labelling, sorting, incubating, and the like. The automated system may be operated as stand-alone device or in conjunction with one or more other devices, e.g. IVD devices. The automated system typically comprises a plurality of functional units, each dedicated to a specific task and cooperating with each other in order to enable automated sample processing and analysis. Such functional units may be a pipetting unit, a pump, a valve, a conveyor, a gripper, an incubation unit, an analytical measurement unit, a temperature regulating unit, a controller, etc.

[0043] According to an aspect of the present disclosure, the automated system further comprises: an imaging unit configured to generate a digital microscope image of one or more particles on the substrate; a display device; and an image processor configured for analyzing the one or more particles in the digital microscope image to generate an analytical result for the liquid sample, where the workflow management unit controls the automated system to display the property of the liquid sample and / or the analytical result on the display device.

[0044] The term “image processor” as used herein may refer to a programmable logic controller or processor running a computer-readable program provided with instructions to perform operations in accordance with an operation plan. The term can mean central processing units, microprocessors, microcontrollers, reduced instruction circuits (RISC), application specific integrated circuits (ASIC), logic circuits, and any other circuit or processor capable of executing the functions / methods described herein. Regardless of the type of processor, it is configured to execute one or more of the methods described herein. The image processor may be integrated into a unit, a sub-unit or a module of the automated system in communication with other units, subunits or modules of the automated system via a direct connection, wired or wirelessly, or indirectly over a communications network, wired or wirelessly. In some aspects, the controller might be integral with a data management unit, e.g. implemented on a computing device such as a desktop computer, a laptop, a smartphone, a tablet, PDA, etc., or it may be comprised by a server computer and / or be distributed / shared across / between a plurality of automated systems. Moreover, the systems can include remote devices, servers and cloud-based elements that communicate via wires or wirelessly (e.g. infrared, cellular, Bluetooth®), or a remote PC / server or a cloud-based system.In particular, the image processor is configured for analyzing particles in the digital microscope image of a liquid sample and to generate an analytical result for the liquid sample.

[0045] According to aspects of the present disclosure, the automated system further comprises the property determination unit configured to determine the property of the liquid sample by any of an impedance measurement or a conductivity measurement or a resistance measurement or an electro-chemical measurement or an optical measurement or a pressure measurement or a viscosity measurement.

[0046] According to an aspect of the present disclosure, the property determination unit further comprises a pipetting probe operatively coupled to a pressure sensor, wherein the controller is configured to: control the pipetting probe to aspirate a portion of the liquid sample from a sample container; while aspirating the liquid sample, measure pressure inside the pipetting probe by the pressure sensor; correlate the measured pressure with a hematocrit range; and store the measured pressure and correlated hematocrit range as property of the liquid sample in a data storage.

[0047] According to aspects of the present disclosure, the sample preparation unit comprises a dilution module and / or a concentration module configured to dilute or concentrate the liquid sample, respectively, according to the operational parameters set by the workflow management unit, wherein the operational parameters comprise a dilution factor and / or a concentration factor. An example of a concentration module may be a centrifuge for centrifuging the liquid sample and thereby concentrating particles in the liquid sample or separating different kinds of constituents of the liquid sample, e.g. like fractionating a whole blood sample. Dilution, suspension, or dispersion of the liquid sample may be performed in water or an appropriate buffer such as PBS (phosp hatebuffer saline), physiological sodium chloride solution or other buffers known to the skilled person. The sample preparation unit may therefore comprise respective functional units, for example, a pipette or dispenser for administering a dilution liquid or buffer into the liquid sample or vice versa. The sample preparation unit may further comprise appropriate reaction vessels, for example, for mixing and incubating liquid samples with reagents. An example of a dilution and / or concentration module is the automated system for processing particles as described in US 10,436,685.

[0048] The present disclosure further refers to a computer program product comprising instructions to cause the automated system as described above to execute the automated method as described above.BRIEF DESCRIPTION OF THE DRAWINGSFIG. 1 shows a flow diagram of an automated method for applying a liquid sample onto a substrate for image analysis according to the present disclosure;FIG. 2 shows a flow diagram of an automated method for applying a liquid sample onto a substrate for image analysis according to further aspects of the present disclosure;FIG. 3 shows a flow diagram of an automated method for applying a liquid sample onto a substrate for image analysis according to further aspects of the present disclosure;FIG. 4 shows a flow diagram of an automated method for applying a liquid sample onto a substrate for image analysis according to further aspects of the present disclosure;FIG. 5 shows a schematic illustration of an automated system according to the present disclosure;FIG. 6 shows a schematic illustration of an automated system according to further aspects of the present disclosure;FIG. 7 shows a schematic illustration of an automated system according to further aspects of the present disclosure;FIGS. 8A, 8B show microscope images of particles in a liquid sample on a substrate.

[0049] Skilled artisans appreciate that elements in the figures are illustrated schematically for simplicity and clarity and have not necessarily been drawn to scale. For example, the dimensions of some of the elements in the figures may be exaggerated relative to other elements to help improve understanding of the aspects of the present disclosure. Also, parts have been omitted that do not contribute to the teaching of this disclosure.DETAILED DESCRIPTION

[0050] FIG. 1 shows a flow diagram of an automated method for applying a liquid sample onto a substrate for image analysis. The automated method comprises providing data corresponding to a property of the liquid sample from a property determination unit to a workflow management unit 90 (step 102). For example, the property determination unit may be an impedance-based flow cytometer, using the coulter principle to determine the property of the liquid sample. The property of the liquid sample being determined may for example refer to a particlecount in the liquid sample, in particular the number of red blood cells (RBCs), the number of white blood cells (WBCs) and the number of platelets, as well as the hematocrit, which can be calculated from the number of RBCs multiplied with the mean cell volume of the RBCs and put in relation to the known sample volume. Any property of the liquid sample determined in this way is typically stored electronically, e.g. in a data storage. Therefore, the property determination unit may comprise a data storage (not shown) or it may be communicatively connected to a remote data storage (not shown), such as a computer network or a cloud. From there, the data corresponding to the property of the liquid sample is provided to the workflow management unit 90, via commonly known means of electronic data transfer, e.g. wired or wirelessly. Information regarding the identity of the liquid sample, such as an ID-number, may be provided to the workflow management unit 90 together with the data corresponding to the property of the liquid sample or it may be provided independently therefrom, e.g. by a local laboratory information system (LIS) (not shown).

[0051] Once the data corresponding to the property or properties of the liquid sample have been provided to the workflow management unit 90, the workflow management unit 90 selects sample preparation operations based thereon (step 104). According to a specific aspect of the present disclosure, the sample preparation operations at least include applying the liquid sample onto the substrate by controlling an applicator to translate in relation to the substrate while dispensing the liquid sample through the applicator onto the substrate, so that particles in the liquid sample settle onto the substrate, and staining the liquid sample on the substrate and / or fixating the liquid sample on the substrate. Other examples of sample preparation operations are mixing, dispersing, aliquoting, transferring, diluting or concentrating of the liquid sample, etc.

[0052] The workflow management unit 90 sets operational parameters based on the selected sample preparation operations and / or on the property of the liquid sample (step 106). According to an aspect of the present invention, the operational parameters comprise a dilution parameter and / or a concentration parameter. According to another aspect of the present disclosure, the operational parameters comprise at least one or a combination of parameters chosen from the group of a distance between the applicator and the substrate, a movement pattern of the applicator relative to the substrate, a speed pattern of applicator relative to the substrate, a liquid sample dispensing volume, a dispensation rate, and / or a distance between rows of liquid sample on the substrate.

[0053] The selected sample preparation operations and the operational parameters set by the workflow management unit 90 are transmitted as instructions to a sample preparation unit toprepare the liquid sample by performing the sample preparation operations selected by the workflow management unit 90 and by using the operational parameters set by the workflow management unit 90 (step 108).

[0054] FIG. 2 shows a flow diagram of an automated method for applying a liquid sample onto a substrate for image analysis according to an aspect of the present disclosure. Step 202 of providing data corresponding to the property of the liquid sample corresponds to step 102 in FIG. 1. Therefore, for a detailed description of step 202, reference is made to the description of FIG. 1

[0055] Step 204 corresponds to step 104 of selecting sample preparation operations based on the property of the liquid sample in FIG. 1, but shows further sub-steps of the selection process according to an aspect of the present disclosure. In general, the workflow management unit 90 selects the sample preparation operations by determining whether the data or data point corresponding to the property of the liquid sample lies above or below a predetermined threshold, whether it lies in a predetermined range, or whether it corresponds to a predetermined category or reference value. In the example shown in FIG. 2, the workflow management unit 90 determines which predetermined hematocrit range the data corresponding to the property of the liquid sample corresponds to, e.g. a normal, a high, or a low hematocrit range (step 2042). The hematocrit ranges may rely on the sample donor’s gender and age. For example, a normal range for adult women may be between 36% and 45%, a normal range for adult men between 40% and 51%. Therefore, the sample donor’s gender may be taken into consideration when determining the hematocrit range of a liquid sample. Depending on the outcome of the check in step 2042, the workflow management unit 90 selects respective sample preparation operations. For example, if the workflow management unit 90 determines in step 2042 that the hematocrit of the liquid sample is in a normal range, it selects the sample preparation operations of applying the liquid sample onto the substrate by controlling an applicator to translate in relation to the substrate while dispensing the liquid sample through the applicator onto the substrate, so that particles in the liquid sample settle onto the substrate, staining the liquid sample on the substrate and fixating the liquid sample on the substrate (step 2044). In another example and according to an aspect of the present invention, if the workflow management unit 90 determines in step 2042 that the hematocrit of the liquid sample is in an abnormally high range, which may, for example, be defined as being in a range between 55% to 60% or above a predetermined upper threshold of 55%, it selects the sample preparation operations of dilution of the liquid sample, applying the liquid sample onto the substrate, staining the liquid sample on the substrate and fixating the liquid sample on the substrate(step 2046). In yet another example and according to an aspect of the present invention, if the workflow management unit 90 determines in step 2042 that the hematocrit of the liquid sample is in an abnormally low range, which may, for example, be defined as being in a range between 20% to 30% or below a predetermined lower threshold of 30%, it selects the sample preparation operations of concentration of the liquid sample, applying the liquid sample onto the substrate, staining the liquid sample on the substrate and fixating the liquid sample on the substrate (step 2048).

[0056] Further, the workflow management unit 90 sets operational parameters based on the selected sample preparation operations and / or on the property of the liquid sample (step 206). In the above mentioned example, where the property of the liquid sample is a hematocrit in a normal range, the workflow management unit 90 sets one or more operational parameters selected from the group of a distance between the applicator and the substrate, a movement pattern of the applicator relative to the substrate, a speed pattern of applicator relative to substrate movement, a liquid sample dispensing volume, which is a portion of the aspirated liquid sample, a dispensation rate, a distance between rows of liquid sample on the substrate. In particular, the workflow management unit 90 may set the distance between the applicator and the substrate to 12 pm, the dispensation rate to 0.055 pL / s, and the speed pattern to 65 mm / s with regard to the applicator movement in x-direction (step 2064). According to another aspect, the operational parameters may include the volume and type of stain to be used for staining the liquid sample. In another example, where the liquid sample has a hematocrit in an abnormally high range, and where a dilution operation was selected, the workflow management unit 90 sets a dilution parameter, e.g. to a volume ratio of 1 :5 (liquid sample: diluent) (step 2066). Further operational parameters may be set in step 2066 analogous to the ones mentioned in the previous example (step 2064). In yet another example, where the liquid sample has a hematocrit in an abnormally low range, and where a concentration operation was selected, the workflow management unit 90 sets concentration parameters, e.g. the volume of an aliquot to 500 pL, the rotational speed for a centrifuge to about 2000 revolutions per minute (rpm) for a duration of 10 minutes, and the volume of supernatant to be removed after centrifugation to 250 pL (step 2068). Further operational parameters may be set in step 2068 analogous to the ones mentioned with regard to the previous examples (steps 2064 and 2066).

[0057] Step 208 corresponds to step 108 in FIG. 1, and refers to transmitting the selected sample preparation operations and the operational parameters set by the workflow management unit 90 as instructions to a sample preparation unit to prepare the liquid sample accordingly. Thisresults in a substrate, e.g. a glass microscope slide, carrying the liquid sample, where particles of the liquid sample, e.g. RBCs, WBCs and platelets, are uniformly distributed across the substrate in a monolayer. Furthermore, an adequate number of particles is available on the substrate for subsequent image analysis, thereby ensuring the generation of a reliable analytical result. According to an aspect of the present disclosure, the automated method further comprises controlling an imaging unit to generate a digital microscope image of the one or more particles on the substrate, to analyze the one or more particles in the digital microscope image in order to generate an analytical result for the liquid sample, and to display the data corresponding to the property of the liquid sample and / or the analytical result on a display device (step 210).

[0058] FIG. 3 shows a flow diagram of an automated method for applying a liquid sample onto a substrate for image analysis according to a further aspect of the present disclosure. Step 302 refers to providing data corresponding to a property of the liquid sample from a property determination unit to a workflow management unit 90. According to an aspect of the present disclosure, providing the data comprises controlling the property determination unit to determine the property of the liquid sample by any of an impedance measurement, a conductivity measurement, a resistance measurement, an electro-chemical measurement, an optical measurement, a pressure measurement or a viscosity measurement, and then transmitting said measurement data to the workflow management unit 90. For example, the property determination unit in the example in FIG. 3 may be a module or a functional unit within the automated system of the present disclosure, such as a photometer, a spectrophotometer, or a pressure sensor operatively coupled to a pipetting probe (further examples are described above). The data may be raw data, e.g. photometric information in the form of transmission or absorption data that needs to be interpreted or converted first by the workflow management unit 90 in order to derive information regarding the property of the liquid sample. Alternatively, the property determination unit may comprise a separate processor for processing the raw data before providing it to the workflow management unit 90.

[0059] In a first example of step 304 in FIG. 3, the hematocrit is found to be moderately elevated compared to a normal range, e.g. in a range between 51% and 55% (step 3042). In another example, the hematocrit is found to be moderately decreased compared to a normal range, e.g. in a range between 30% and 35% (step 3042). In yet another example, the hematocrit is found to be in a normal range, e.g. in a range between 35% and 51%. According to the aspect of the present invention shown in FIG. 3, the workflow management unit 90 selects the following sample preparation operations in the event of only moderately elevated or decreased hematocrit in a liquidsample: applying the liquid sample onto the substrate by controlling an applicator to translate in relation to the substrate while dispensing the liquid sample through the applicator onto the substrate, so that particles in the liquid sample settle onto the substrate, staining the liquid sample on the substrate and fixating the liquid sample on the substrate (steps 3046 and 3048). The selected sample preparation operations are the same as the ones selected for a liquid sample with a normal hematocrit (step 3044). In the event of only moderately elevated or decreased parameters, a designated dilution operation or concentration operation, as described in the examples of FIG. 2, may not be required.

[0060] The workflow management unit 90 then sets operational parameters based on the selected sample preparation operations and on the property of the liquid sample (step 306) in order to generate an improved specimen for subsequent image analysis. In particular, to enable the formation of a monolayer of sample particles on the substrate while ensuring an adequate number of particles and a uniform distribution of particles on the substrate. In the example of a liquid sample with a moderately elevated hematocrit, the workflow management unit 90 may set the distance between the applicator and the substrate to 12 pm, the liquid sample dispensing volume to 1 pL, the dispensation rate to a value in a range between 0.01 mL / s and 0.055 pL / s, e.g. 0.04 pL / s, and the distance between rows of liquid sample to range between 0.37 mm and 0.6 mm, e.g. to 0.5 mm (step 3066). In another example of a liquid sample with a moderately decreased hematocrit, the workflow management unit 90 may set the distance between the applicator and the substrate to 12 pm, the liquid sample dispensing volume to 4 pL, the dispensation rate to a value in a range between 0.1 mL / s and 0.4 pL / s, e.g. 0.22 pL / s, and the distance between rows of liquid sample to range between 0.1 mm and 0.37 mm, e.g. to 0.123 mm (step 3068). In the example of a liquid sample with a normal hematocrit, the workflow management unit 90 may set the distance between the applicator and the substrate to 12 pm, the liquid sample dispensing volume to 1 pL, the dispensation rate to a value in a range between 0.04 mL / s and 0.08 pL / s, e.g. 0.055 pL / s, and the distance between rows of liquid sample to range between 0.3 mm and 0.5 mm, e.g. to 0.37 mm (step 3064).

[0061] Other examples of possible operational parameter settings that were experimentally investigated are shown in table 1. The examples illustrate possible combinations of settings that, depending on the property of the liquid sample, enable monolayer formation of sample particles on the substrate and ensure an adequate number and uniform distribution of particles on the substrate.Table 1 : Examples of operational parameter settings

[0062] A microscope image of cells in a synovial fluid sample applied onto a substrate can be seen in FIG. 8A. In order to achieve an adequate number and uniform distribution of cells, the liquid sample dispensing volume was set to 9 pL, the dispensation rate to 0.495 pL / s, and the distance between rows to 0.053 mm. In comparison, FIG. 8B shows the same synovial fluid sample applied onto a substrate with the same operational parameters, except for using a liquid sample dispensing volume of 1 pL instead of 9 pL. As can be seen, the number of cells in FIG. 8B is lower than the number of cells in FIG. 8A and not adequate to perform a reliable analysis. In consequence and according to aspects of the present invention, the workflow management unit 90 sets the operational parameters according to the example in FIG. 8A if the property of the liquid sample is determined as being a synovial fluid sample and / or if the property of the liquid sample is determined as having a viscosity or density below a predetermined threshold.

[0063] With continued reference to FIG. 3, steps 308 and 310 correspond to steps 208 and 210 in FIG. 2. Therefore, reference is made to the description of FIG. 2 for a detailed description of steps 308 and 310.

[0064] FIG. 4 shows a flow diagram of an automated method for applying a liquid sample onto a substrate for image analysis according to another aspect of the present disclosure. The property determination unit in the example in FIG. 4 may be a device for generating data corresponding to a sample type of the liquid sample, e.g. a photometer or a spectrophotometer. The generated data corresponding to the property, i.e. the sample type, of the liquid sample is provided from the property determination unit to the workflow management unit 90 (step 402).

[0065] The automated method further comprises selecting, by the workflow management unit 90, sample preparation operations based on the property of the liquid sample (step 404). According to the aspect shown in FIG. 4, the workflow management unit 90 determines the sample type of the liquid sample based on the data received by the property determination unit, which in this example is a whole blood sample or a cerebrospinal fluid sample (step 4042). In the example related to the whole blood sample, the workflow management unit 90 selects sample preparation operations that include the application of the liquid sample onto the substrate, thestaining of the liquid sample on the substrate and the fixation of the liquid sample on the substrate (step 4044). In the example of the cerebrospinal fluid sample, which typically contains a low number of cells or particles, the workflow management unit 90 selects sample preparation operations that include the application of the liquid sample onto the substrate, the staining of the liquid sample on the substrate, the fixation of the liquid sample on the substrate, and - in contrast to the whole blood sample - the concentration of the liquid sample (step 4048), in order to increase the concentration of cellular material in the liquid sample relative to its volume.

[0066] The workflow management unit 90 then sets operational parameters based on the selected sample preparation operations and on the property of the liquid sample (step 406). In the example related to the whole blood sample and the sample preparation operation of applying the liquid sample onto the substrate, as selected in step 4044, the workflow management unit 90 sets the operational parameter defining the distance between the applicator and the substrate to 0.7 pm, the operational parameter concerning the liquid sample dispensing volume 1 pL, and the operational parameter concerning the dispensation rate to 0. 1 uL / s (step 4064). In the example related to the cerebrospinal fluid sample and the sample preparation operation of applying the liquid sample onto the substrate, as selected in step 4048, the workflow management unit 90 sets the operational parameter defining the distance between the applicator and the substrate to 0.7 pm, the operational parameter concerning the liquid sample dispensing volume 4 pL, and the operational parameter concerning the dispensation rate to 0.4 uL / s (step 4068). This allows for applying more liquid sample material onto the substrate, thereby compensating for the lower number of cellular material in the liquid sample. In relation to the concentrating operation that was selected in step 4048, the workflow management unit 90 sets the respective operational parameters, including for example the rotational speed of a centrifuge (step 4068) and / or the volume of supernatant to be removed.

[0067] Steps 408 and 410 correspond to steps 208 and 210 in FIG. 2 and steps 308 and 310 in FIG. 3. Reference is therefore made to the description of FIG. 2 and FIG. 3 for a detailed description of steps 408 and 410.

[0068] FIG. 5 shows a schematic illustration of an automated system 100 according to the present disclosure, where the automated system 100 comprises a sample preparation unit 10 and a workflow management unit 90. The sample preparation unit 10 comprises an applicator 12, which may be a hollow needle- or syringe-shaped element, through which the liquid sample 1 is dispensed onto a substrate 2. The substrate 2 is placed onto a substrate holder 4 for the process of applying the liquid sample 1 onto the substrate 2. The applicator 12 and the substrate 2 on thesubstrate holder 4 may be configured to move translationally relative to each other in an x-direction, a y-direction and a z-direction. This can be achieved by either moving the applicator 12 relative to a static substrate holder 4, by moving the substrate holder 4 relative to a static applicator 12, or by moving both the applicator 12 and the substrate holder 4. The sample preparation unit 10 further comprises a staining unit 14 for staining the liquid sample 1 on the substrate 2.

[0069] The workflow management unit 90 may be a separate entity, i.e. physically separated from, but in communicative connection with, the automated system 100. The workflow management unit 90 is configured to receive data corresponding to a property of the liquid sample 1 from a property determination unit 20. Based on the received data, the workflow management unit 90 is configured to select sample preparation operations and set operational parameters, which it then communicates to the sample preparation unit 10 to prepare the liquid sample 1 by performing the sample preparation operations selected by the workflow management unit 90 and by using the operational parameters set by the workflow management unit 90. The sample preparation operations comprise the application of the liquid sample 1 onto the substrate 2 by controlling the applicator 12 to move translationally in relation to the substrate 2 while dispensing the liquid sample 1 through the applicator 12 onto the substrate 2, so that particles 3 in the liquid sample 1 settle onto the substrate 2. According to an aspect of the present disclosure, the liquid sample may be a whole blood sample and the particles may be any kind of cellular components, such as RBCs, WBCs, or platelets.

[0070] The property determination unit 20 in FIG. 5 is a separate physical entity, e.g. a laserbased flow cytometer that measures forward-scattered light (FSC) signals, side-scattered light (SSC) signals and / or dye-specific fluorescence signals, and is communicatively connected to the automated system 100, in particular to the workflow management unit 90 of the automated system 100.

[0071] FIG. 6 shows a schematic illustration of an automated system 100 according to further aspects of the present disclosure. The property determination unit 20 is implemented as a module or a functional unit in the automated system 100 and communicatively connected to the workflow management unit 90. It may for example be a photometer or a spectrophotometer, measuring transmission or absorption of light of different wavelength through the liquid sample.

[0072] The sample preparation unit 10 in FIG. 6 corresponds the one shown in FIG. 5. Therefore, reference is made to the description of FIG. 5 for a detailed description of the sample preparation unit 10.

[0073] The automated system 100 in FIG. 6 further comprises an imaging unit 30 configured to generate a digital microscope image of one or more particles 3 in the liquid sample 1 on the substrate 2 and a display device 40. The imaging unit 30 may therefore comprise a light emission device (not shown), a microscope 32 and a light receiving device 34, such as a camera. The image processor 92 is configured for analyzing the one or more particles 3 in the digital microscope image to generate an analytical result for the liquid sample 1 and for displaying the property of the liquid sample 41 and / or the analytical result 42 on the display device 40. The image processor 92 is therefore in communicative connection with the imaging unit 30 and the display device 40, possibly via the workflow management unit 90. The displayed property of the liquid sample 41 and the analytical result 42 may refer to different parameters. For example, the property of the liquid sample may refer to the liquid sample’s hematocrit, while the analytical result may refer to the number and -based on their morphology- type of WBCs in the liquid sample. According to an alternative aspect of the present invention, the property of the liquid and the analytical result may refer to the same parameter, e.g. hematocrit. The workflow management unit 90 may compare, e.g. as part of a quality assessment, the hematocrit measured by the property determination unit 20 with the hematocrit determined based on the analysis of the microscope image. An output value using the analytical result and the property of the liquid sample would then be generated and displayed 41, 42 on the display device 40. The output value may, for example, indicate that the two hematocrit values are the same or adequately similar. Alternatively, if the two hematocrit values differ by a certain predetermined factor, the output value may indicate that the two results are significantly different and that a quality issue in the sample preparation procedure may exist. An alert or warning message may additionally or alternatively be displayed 41, 42 to inform a device operator.

[0074] The automated system 100 in FIG. 7 shows a further aspect of the present invention. The property determination unit 20 in the automated system 100 in FIG. 7 comprises a pipetting probe 22 operatively coupled to a pressure sensor 24. The controller 90 is configured to control the pipetting probe 22 to aspirate a portion of the liquid sample 1 from a sample container 26. While aspirating the liquid sample 1, pressure inside the pipetting probe 22 is measured by the pressure sensor 24 and the measured pressure is correlated with a hematocrit range. As described above, the property determination by pressure measurement is based on the principle that the required negative pressure for aspirating the same volume of liquid sample in the same time period into the pipetting probe 22 depends on the viscosity and / or density of the liquid sample. Determining the pressure during liquid sample aspiration allows estimating the liquid sample’sviscosity and / or density and thereby allows for correlating the pressure measurement to e.g. a low, normal or high hematocrit range. The measured pressure and correlated hematocrit range is then stored as property of the liquid sample in a data storage 80. The data storage 80 is communicatively connected to the workflow management unit 90.

[0075] The sample preparation unit 10 of the automated system 100 in FIG. 7 comprises a dilution module 16 and / or a concentration module 16 configured to dilute or concentrate the liquid sample 1, respectively, according to the operational parameters set by the workflow management unit 92, wherein the operational parameters comprise a dilution parameter and / or a concentration parameter. Examples of dilution modules 16, concentration modules 16, dilution parameters and concentration parameters are mentioned above.

[0076] In the preceding specification, devices and methods according to various aspects are described in detail. The devices and methods may be embodied in many different forms and should not be construed as limited to the aspects set forth and illustrated herein. It is therefore to be understood that the devices and methods are not to be limited to the specific aspects disclosed and that modifications and other aspects are intended to be included within the scope of the appended claims. Although specific terms are employed herein, they are used in a generic and descriptive sense only and not for purposes of limitation. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one skilled in the art to which the disclosure pertains. Although any methods and materials similar to or equivalent to those described herein can be used in the practice or testing of the methods, the preferred methods and materials are described herein.

[0077] Moreover, reference to an element by the indefinite article “a” or “an” does not exclude the possibility that more than one element is present, unless the context clearly requires that there be one and only one element. The indefinite article “a” or “an” thus usually means “at least one.” Likewise, the terms “have,” “comprise” or “include” or any arbitrary grammatical variations thereof are used in a non-exclusive way. Thus, these terms may both refer to a situation in which, besides the feature introduced by these terms, no further features are present in the entity described in this context and to a situation in which one or more further features are present. For example, the expressions “A has B,” “A comprises B” and “A includes B” may refer both to a situation in which, besides B, no other element is present in A (i.e. a situation in which A solely and exclusively consists of B) or to a situation in which, besides B, one or more further elements are present in A, such as element C, elements C and D, or even further elements.

[0078] Also, reference throughout the specification to "one aspect", "an aspect", "one example" or "an example", means that a particular feature, structure or characteristic described in connection with the aspect or example is included in at least one aspect. Thus, appearances of the phrases "in one aspect", "in an aspect", "one example" or "an example", in various places throughout this specification are not necessarily all referring to the same aspect or example.

[0079] Furthermore, the particular features, structures, or characteristics may be combined in any suitable combinations and / or sub-combinations in one or more embodiments or examples.

Claims

CLAIMS1. An automated method for applying a liquid sample (1) onto a substrate (2) for image analysis, comprising:- providing data corresponding to a property of the liquid sample (1) from a property determination unit (20) to a workflow management unit (90); selecting, by the workflow management unit (90), sample preparation operations based on the property of the liquid sample (1); setting, by the workflow management unit (90), operational parameters based on the selected sample preparation operations and / or on the property of the liquid sample (1); controlling a sample preparation unit (10) to prepare the liquid sample (1) for image analysis by performing the sample preparation operations selected by the workflow management unit (90) and by using the operational parameters set by the workflow management unit (90), wherein the selected sample preparation operations at least comprise: applying the liquid sample (1) onto the substrate (2) by controlling an applicator (12) to translate in relation to the substrate (2) while dispensing the liquid sample (1) through the applicator (12) onto the substrate (2), so that particles (3) in the liquid sample (1) settle onto the substrate (2), and staining the liquid sample (1) on the substrate (2) and / or fixating the liquid sample (1) on the substrate (2).

2. The automated method according to claim 1, further comprising: controlling an imaging unit (30) to generate a digital microscope image of one or more particles (3) on the substrate (2); analyzing the one or more particles (3) in the digital microscope image to generate an analytical result for the liquid sample (1); and displaying the data corresponding to the property of the liquid sample (1) and / or the analytical result on a display device (50).

3. The automated method according to claim 2, further comprising generating an output value using the analytical result and the property of the liquid sample.

4. The automated method according to any of the preceding claims, wherein the operational parameters comprise a dilution parameter and / or a concentration parameter and whereinthe sample preparation operations further comprise a dilution or concentration of the liquid sample according to the dilution parameter or concentration parameter, respectively, before applying the liquid sample (1) onto the substrate (3).

5. The automated method according to any of the preceding claims, wherein the operational parameters comprise at least one parameter chosen from the group of: a distance between the applicator (12) and the substrate (2), and / or a movement pattern of the applicator (12) relative to the substrate (2), and / or a speed pattern of applicator (12) relative to the substrate (2), and / or a liquid sample (1) dispensing volume, and / or a dispensation rate, and / or a distance between rows of liquid sample (1) on the substrate (2).

6. The automated method according to any of the preceding claims, wherein the property of the liquid sample (1) comprises any or a combination of a liquid sample type, hematocrit, hematocrit range, sample viscosity, sample density or a particle count.

7. The automated method according to any of the preceding claims, wherein providing data corresponding to a property of the liquid sample (1) comprising controlling the property determination unit (20) to determine the property of the liquid sample (1) by any of an impedance measurement or a conductivity measurement or a resistance measurement or an electro-chemical measurement or an optical measurement or a pressure measurement or a viscosity measurement.

8. The automated method according to claim 7, wherein the pressure measurement comprises: controlling a pipetting probe (22) to aspirate a portion of the liquid sample (1) from a sample container (26);- while aspirating the liquid sample (1), measuring pressure inside the pipetting probe (22) by a pressure sensor (24) operatively coupled to the pipetting probe (22); correlating the measured pressure with a hematocrit range; and storing the measured pressure and correlated hematocrit range as property of the liquid sample (1) in a data storage (80).

9. An automated system (100) for preparing a liquid sample (1) for image analysis according to any of claims 1 to 8, comprising: a sample preparation unit (10) comprising: an applicator (12) for dispensing the liquid sample (1) onto a substrate (2); and a staining unit (14) for staining the liquid sample (1) on the substrate (2) and / or a fixating unit (14) for fixating the liquid sample (1) on the substrate (2); a workflow management unit (90) configured to:- receive data corresponding to a property of the liquid sample (1) from a property determination unit (20); select sample preparation operations based on the property of the liquid sample (1); set operational parameters based on the selected sample preparation operations and / or on the property of the liquid sample (1);- control the sample preparation unit (10) to prepare the liquid sample (1) by performing the sample preparation operations selected by the workflow management unit (90) and by using the operational parameters set by the workflow management unit (90), wherein the sample preparation operations comprise the application of the liquid sample (1) onto the substrate (2) by controlling the applicator (12) to move translationally in relation to the substrate (2) while dispensing the liquid sample (1) through the applicator (12) onto the substrate (2), so that particles (3) in the liquid sample (1) settle onto the substrate (2).

10. The automated system (100) according to claim 9, further comprising: an imaging unit (30) configured to generate a digital microscope image of one or more particles (3) on the substrate (2); a display device (40); and an image processor (92) configured for analyzing the one or more particles (3) in the digital microscope image to generate an analytical result for the liquid sample (1), wherein the workflow management unit (90) controls the automated system (100) to display the property of the liquid sample (1) and / or the analytical result on the display device (40).

11. The automated system (100) according to claims 9 or 10, further comprising the property determination unit (20) configured to determine the property of the liquid sample (1) by any of an impedance measurement or a conductivity measurement or a resistance measurement or an electro-chemical measurement or an optical measurement or a pressure measurement or a viscosity measurement.

12. The automated system (100) according to claim 11, wherein the property determination unit (20) comprises a pipetting probe (22) operatively coupled to a pressure sensor (24), wherein the workflow management unit (90) is configured to: control the pipetting probe (22) to aspirate a portion of the liquid sample (1) from a sample container (26);- while aspirating the liquid sample (1), measure pressure inside the pipetting probe (22) by the pressure sensor (24); correlate the measured pressure with a hematocrit range; and store the measured pressure and correlated hematocrit range as property of the liquid sample (1) in a data storage (80).

13. The automated system (100) according to any of claims 9 to 12, wherein the sample preparation unit (20) further comprises a dilution module (16) and / or a concentration module (16) configured to dilute or concentrate the liquid sample (1), respectively, according to the operational parameters set by the workflow management unit, wherein the operational parameters comprise a dilution parameter and / or a concentration parameter, respectively.

14. A computer program product comprising instructions to cause the automated system (100) according to any of claims 9 to 13 to execute the automated method according to any of claims 1 to 8.

Images