Fully automatic staining device

Abstract

An automatic staining device for dispensing staining reagents over one or more biological samples is disclosed. The device utilizes a pressure-driven pumped flow to deliver reagents and comprises a processor in communication with a memory, at least one camera, one or more sample holder supports, reagent vials with movable lids, an air blowing unit, and a robotic arm with at least one dispensing probe. The device is configured to withdraw reagents from the reagent vials, optionally mix them, and dispense the reagents over the sample holders. The air blowing unit is designed to maintain an adequate moisture level on the sample to facilitate further reagent dispensing, determined by an air flow sensor. Preferably the processor, utilizing machine learning models or geometric algorithms, processes images captured by the camera to determine the presence, position, and orientation of sample holders and reagent vials, as well as the open or closed state of vial lids and preferably to read the labels comprised in the sample holders or reagent vials.

Description

[0001] FULLY AUTOMATIC STAINING DEVICE

[0002] Technical field of the invention

[0003] The present invention relates to the field of laboratory automation and sample processing technology, specifically to automated devices for dispensing staining reagents onto biological samples. More particularly, it relates to methods, systems and apparatus for performing histological staining, immunoenzymatic staining and / or hybridisation on tissue sections or cell smears.

[0004] Background of the invention

[0005] The field of biological sample processing, particularly in histology and pathology laboratories, is continuously evolving to enhance efficiency, accuracy, and reliability. Fully automatic staining machines play a crucial role in modern laboratories by automating the application of staining reagents to biological samples. These machines are essential for high-throughput processing, reducing manual labour, minimizing human error, and ensuring consistent staining results, which are vital for accurate diagnosis and research outcomes.

[0006] Current automatic staining machines predominantly rely on capillarity-based mechanisms to deliver reagents to samples. While these systems have introduced a degree of automation, they present several significant drawbacks. The reagents used often contain high concentrations of salts and other substances that tend to crystallize or solidify within the capillary channels. This crystallization necessitates frequent and thorough cleaning to maintain the functionality of the machine. However, in practical laboratory settings — especially in research labs with busy schedules and inexperienced users such as PhD students, or researchers that will be there only temporarily — the meticulous cleaning required is often neglected. This oversight leads to the machines becoming non-functional within months, requiring maintenance and causing downtime and ever-growing expenses.

[0007] Moreover, capillary-based devices are typically closed systems that are complicated to clean. Accessing the internal components for thorough cleaning involves disassembling several parts, which is cumbersome and time-consuming. This complexity discourages users from performing regular maintenance. Additionally, the closed design limits accessibility, making it challenging to monitor and intervene during the staining process if necessary.

[0008] Attempts to develop automatic staining machines without capillarity have aimed to create open systems that are easier to access and clean, thereby avoiding crystallization problems. However, these non-capillary approaches have encountered several challenges. Unlike capillary devices, which are relatively insensitive to the stability and inclination of the machine due to the passive nature of capillary action, non-capillary are affected over the device's stability and angle of operation. Maintaining consistent inclination is critical to ensure uniform reagent distribution.

[0009] Furthermore, non-capillary systems often require significantly more reagent volume to achieve adequate coverage of the samples with respect to capillary systems. This increased usage not only raises operational costs but also presents practical obstacles in handling and disposing of excess reagents. Removing reagents can result in samples becoming too dry, hindering the proper diffusion of subsequent reagents and affecting staining quality. Additionally, some reagents are highly viscous and do not spread easily, leading to uneven staining or the need for the use of excessive reagent quantities to ensure full coverage.

[0010] Another limitation of existing non-capillary automatic staining machines is the lack of precision in reagent dispensing. These devices may dispense reagents over the entire sample holder, wasting valuable reagents, increasing the risk of spills, and heightening the potential for crosscontamination between samples. The inefficiency in reagent usage not only adds unnecessary costs but also complicates waste disposal procedures due to the larger volumes of hazardous materials involved.

[0011] Moreover, current automatic staining devices do not fully automate the entire staining process. Users are often required to manually verify that all reagents and samples are correctly placed and that the system is properly configured before operation. In laboratories processing numerous samples — such as managing 42 samples with 56 different reagents — this manual oversight is prone to errors. Mistakes in reagent placement or sample loading can lead to incorrect staining, wasted resources, and the need for repeat experiments, thereby reducing overall laboratory efficiency.

[0012] There is, therefore, a pressing need for a fully automated, smart staining device that operates without the limitations of capillarity. Such a device should eliminate the issues associated with capillary systems, including reagent crystallization and cumbersome maintenance. It should also address the challenges faced by non-capillary systems, such as the need for excessive reagent volumes, difficulties in removing excess reagents without over-drying samples, and problems with viscous reagents not spreading adequately. The device should ensure precise dispensing of reagents without relying only on covering the full surface of the sample holder, of which the sample occupies only a fraction, minimizing thus waste and reducing the risk of contamination. Additionally, it should automate the full staining process, from the withdrawal of reagents to the mixing and the dispensing, minimizing user intervention and thus the potential for human error. By meeting these needs, the device would significantly improve laboratory efficiency, reduce operational costs, and enhance the reliability of staining results. Summary of the invention

[0013] To overcome the above-mentioned limitations of the staining devices that use capillarity, and that are not completely automated and require user supervision, as well as to tackle the concomitant problems that arise from not using capillarity, herein it is disclosed a smart, fully automated staining device for dispensing staining reagents over one or more biological samples, wherein the reagents are delivered through a pressure-driven pumped flow.

[0014] By using a pumped flow, instead of capillarity, the problems arising from reagent crystallization and continuous and complex maintenance of the automatic staining device is solved. Regarding the issues that arise from not using capillarity, several innovations are herein disclosed that alone or in combination help overcome said challenges. In some embodiments, the automatic staining device comprises an air blower unit which an air blower and an air flow sensor, which makes it possible to regulate the air flow, and therefore this way it is possible to spread the reagent on the sample support, and also to remove excess reagent just until an optimum point of moisture, which allows to continue the staining procedure, as if it the sample or sample holder was left too dry, the new reagents dispensed would present difficulties to spread and distribute over the sample and / or sample holder. Besides, in some preferable embodiments of the invention, the air blower presents a size and positioning in the air exits that allows for an even distribution of the reagent over the sample and / or sample holder.

[0015] For reagents that are particularly viscous, in some preferred embodiments of the invention it is used a spreading element that can mechanically sweep the sample and / or sample holder, mechanically spreading thus the reagent, which also allows to localize the dispensing of the reagent to desired parts of the sample and / or sample holder, such as for example the half, third or quarter of the sample holder wherein the sample is comprised, allowing thus a directed dispensation that reduces the waste of expensive and possibly hazardous reagents.

[0016] Furthermore, in some embodiments, to achieve a full automatization, the automatic staining device of the invention comprises a camera that can take pictures of the sample holders and of the reagent vials, allowing to determine, through the use of mathematical algorithms and / or or artificial intelligence, important information necessary to start the process, such as if the sample holders in place and in the correct orientation, read the labels of each sample holder to, for example, determine which reagents need to be dispensed in each sample holder, and furthermore it can determine if said needed reagents are in place, and if the lids are open, which is a vital information to automatically start the process. Not only that, in preferred embodiments of the invention the smart automatic staining device can, through analysing the images taken by the camera with a machine learning algorithm trained to recognize image features, recognize the size and position of the sample in a given sample holder to direct a smart dispensing only over the section of the sample holder comprising the sample, saving thus expensive and potentially hazardous reagents.

[0017] Additionally, in some preferred embodiments of the invention, the disclosed automatic staining device comprises inclination sensors, which can be very useful to determine if the sample holders are completely horizontal, which is of paramount importance in devices that do not use capillarity, and also if the staining device is stable, and the movement of the robotic arm does not cause the whole device to move or lose stability, which again can negatively affect the staining process in these types of devices.

[0018] In a first aspect of the invention, an automatic staining device (1) for dispensing staining reagents over one or more biological samples is disclosed, wherein one or more reagents are delivered through a controlled pressure-driven flow. The automatic staining machine comprises a pressure-driven fluid displacement mechanism, a processor in communication with a memory, at least one camera, one or more sample holder supports suitable to support one or more sample holders, the sample holders preferably comprising at least one biological sample, one or more reagent vials comprising a lid, wherein the lid is movable between an open and a closed state, one air blowing unit comprising an air blower and an air flow sensor, wherein the air blowing unit is configured to remove reagent from the sample and / or sample holders by blowing air over the reagent, preferably one or more mixing vials, and a robotic arm comprising at least one dispensing probe, wherein the dispensing probe is fluidly connected to the pressure-driven fluid displacement mechanism.

[0019] In some embodiments, the automatic staining device is further characterized in that the air blowing unit is configured to blow air over the reagent only until a moisture level is left on the sample and / or sample holder adequate to facilitate further reagent dispensing over the sample and / or sample holder. The adequate moisture level is determined based on the flow of air sensed by the air flow sensor. The memory in communication with the processor comprises data representing a set of instructions that, when executed by the processor, cause the processor to receive one or more images taken by one of the at least one camera comprising the image of one or more sample holders, and determine the presence, the position, and / or the orientation of the sample holders based on said images. The determination of the position and / or orientation is made through geometric algorithms or through a first trained machine learning model. The set of instructions further causes the processor to read one or more labels comprised in the image of the one or more sample holders through the first or a second trained machine learning model, wherein said labels comprise at least data relating to a staining procedure. In some embodiments, the processor is also configured to receive one or more images taken by one of the at least one camera comprising the image of one or more reagent vials and determine the presence of the one or more reagent vials and / or the open or closed state of the lid of said reagent vials based on said images. The determination of the presence of the one or more reagent vials and / or the open or closed state of the lid is made through the first, the second, or a third trained machine learning model. Preferably, the set of instructions causes the processor to read one or more labels comprised in the image of the one or more reagent vials through the first, second, third, or a fourth trained machine learning model, wherein said labels comprise at least data relating to a reagent.

[0020] In some embodiments, the dispensing by the dispensing probe comprised in the robotic arm, via a pressure-driven fluid displacement, depends on the determination of the presence, position, and / or orientation of the sample holder and the reading of the one or more labels from the sample holder, and on the determination of the presence and open or closed state of the lid of one or more reagent vials, and optionally from the reading of the one or more labels from the reagent vials.

[0021] In a preferred embodiment of the first aspect of the invention, the automatic staining device further comprises a spreading unit comprising a supporting element and a spreading element, the spreading unit preferably being part of the robotic arm. The longitudinal axis of the spreading element is or can be placed parallel to the surface of the one or more sample holders and / or the surface of the sample holder supports, and the spreading element is supported by a supporting element that can be displaced relative to the spreading element in a direction perpendicular to the longitudinal axis of the spreading element and parallel to the longitudinal axis of the supporting element, between a first configuration wherein the supporting element supports the weight of the spreading element, and a second configuration wherein the spreading element is not supported by the supporting element, such as when it is supported by the sample holder and / or the sample holder support.

[0022] According to a preferred embodiment, the automatic staining device further comprises one or more inclination sensors to detect the inclination of the one or more sample holders with respect to the vertical direction, the gravitational pull or the surface on which they are placed.

[0023] In another preferred embodiment, the sample holder supports comprise a grid of at least 6 apertures to allow excess reagent to flow through instead of accumulating between the sample holder and the sample holder support, preferably comprising at least 9 apertures, more preferably comprising a grid of at least 3x6 apertures, and even more preferably comprising a grid of at least 3x12 apertures. According to yet another preferred embodiment, the set of instructions, when executed by the processor, further cause the processor to receive one or more images taken by one of the at least one camera comprising the image of one or more sample holders, which can be the same or different from the images received previously, and provide said one or more images to the first, second, third, or a fourth trained machine learning model, and determine the position and size of the sample in the one or more sample holders based on the output received from the machine learning model. The withdrawal, mixing, and / or dispensing by the automatic staining device further depend on the determination of the position and size of the sample in the one or more sample holders.

[0024] In a preferred embodiment of the first aspect of the invention, the automatic staining device is configured to dispense an amount of reagent proportional to the size of the sample determined, and is further configured to dispense said reagent in a position on the sample holder depending on the position of the sample determined.

[0025] According to another preferred embodiment, the set of instructions, when executed by the processor, further cause the processor to receive one or more images taken by one of the at least one camera comprising the image of one or more mixing vials, provide said one or more images to the first, second, third, fourth, or a fifth trained machine learning model, and determine the presence and position of the one or more mixing vials based on the output received from the machine learning model. The mixing by the automatic staining device depends on the determination of the presence and position of the one or more mixing vials.

[0026] In a preferred embodiment, the automatic staining device is configured such that the longitudinal length of the spreading element is substantially the same as the width of the sample holders and / or sample holder supports, or has a difference in length with said width of ± 25%.

[0027] According to another preferred embodiment, the spreading element is cylindrical and comprises a distal part, a proximal part, and a central part, wherein the diameter of the distal and proximal parts is the same for both the distal and proximal parts and is bigger than the diameter of the central part. The diameter of the distal and proximal parts is between 0.02 and 2 mm longer than the diameter of the central part, preferably between 0.06 and 0.6 mm longer, more preferably between 0.1 and 0.3 mm longer.

[0028] In a further preferred embodiment, the air blower has a shape and orientation adequate to evenly spread the reagents over the sample and / or sample holder, preferably such that the air is blown at an angle between 10 to 80 degrees with respect to the normal to the surface of the sample holder, more preferably between 25 and 65 degrees, even more preferably between 35 and 55 degrees.

[0029] According to a preferred embodiment of the invention, the air blower comprises one or more orifices to blow the air, and the longitudinal axis of the air blower has a length substantially the same as the width of the sample holders and / or sample holder supports, or has a difference in length with said width of ± 25%. If the air blower comprises only one orifice, said orifice is elongated in the direction of the longitudinal axis of the air blower, wherein the maximum length of said orifice is substantially the same as the width of the sample holders and / or sample holder supports, or has a difference in length with said width of ± 25%. If the air blower comprises more than one orifice, said orifices are placed along the longitudinal axis of the air blower, wherein the two most distanced orifices with respect to each other are separated such that at least part of one of the orifices is separated from at least part of the other orifice by a distance substantially the same as the width of the sample holders and / or sample holder supports, or has a difference in length with said width of ± 25%.

[0030] In a preferred embodiment, one of the at least one cameras is comprised in the robotic arm, and the central axis of the lens of said camera presents an inclination angle with respect to the normal axis of the plane of the sample holders and / or sample holder supports of between 10 and 80 degrees, preferably between 25 and 65 degrees, more preferably between 30 and 60 degrees, even more preferably around 45 degrees.

[0031] According to a further preferred embodiment, one of the one or more inclination sensors is configured to detect the inclination of the automatic staining device with respect to the vertical direction, preferably wherein said inclination is detected as a function of time or compared with previous measurements to determine the stability of the automatic staining device.

[0032] In another preferred embodiment, the automatic staining device further comprises a washing receptacle suitable to fit the length of the spreading element.

[0033] In a second aspect of the invention, an automatic staining device for dispensing staining reagents over one or more biological samples is disclosed, wherein one or more reagents are delivered through a controlled pressure-driven flow. The automatic staining device comprises a pressure-driven fluid displacement mechanism; a memory and a processor in communication with said memory, wherein the memory comprises at least one of information of at least one protocol or dispensing protocol, said protocol comprising information related to at least one reagent dispensing step and at least one blowing step, and information related to the one or more biological samples; one or more sample holder supports suitable to support one or more sample holders; one or more reagent vials; an air blowing unit comprising an air blower and, preferably, an air flow sensor; and a robotic arm comprising at least one dispensing probe, wherein the dispensing probe is fluidly connected to the pressure-driven fluid displacement mechanism, preferably wherein the air blowing unit is also comprised in the robotic arm. The memory comprises instructions that, when executed by the processor, cause the processor, during a blowing step, to regulate the flow or speed of air in the air blowing unit or the time duration of air blowing, the robotic arm displacement or speed during the air blowing step and / or the distance of the air blower to the sample or sample holder, based on information stored in the memory regarding a dispensing step of the protocol, preferably a previous, current or following reagent dispensing step, preferably a following or next step, and / or, optionally, based on information regarding the biological sample.

[0034] According to another preferred embodiment of the second aspect of the invention, during an air blowing step, the flow or speed of air in the air blowing unit, the time duration of air blowing, the robotic arm displacement or speed, and / or the distance of the air blower (60) to the sample or sample holder (41) is regulated based on a reagent dispensing step, preferably based on a previous, current or following reagent dispensing step, more preferably based on a following reagent dispensing step of the protocol.

[0035] According to another preferred embodiment of the second aspect of the invention, the air blowing unit is configured to remove a reagent from the sample and / or sample holders by blowing air over said reagent; and / or blow air over the reagent only until a moisture level is left on the sample and / or sample holder adequate to facilitate further reagent dispensing or reagent spreading over said sample and / or sample holder.

[0036] According to another preferred embodiment of the second aspect of the invention, the air blower has a shape and orientation adequate to evenly spread the reagents over the sample and / or sample holder, preferably such that the air is blown at an angle between 10 to 80 degrees with respect to the normal to the surface of the sample holder, more preferably between 25 and 65 degrees, even more preferably between 35 and 55 degrees.

[0037] According to another preferred embodiment of the second aspect of the invention, the air blower comprises one or more orifices to blow the air, and the longitudinal axis of the air blower has a length substantially the same as the width of the sample holders and / or sample holder supports, or has a difference in length with said width of ± 25%; and / or if the air blower comprises only one orifice, said orifice is elongated in the direction of the longitudinal axis of the air blower, wherein the maximum length of said orifice is substantially the same as the width of the sample holders and / or sample holder supports, or has a difference in length with said width of ± 25%; and / or if the air blower comprises more than one orifice, said orifices are placed along the longitudinal axis of the air blower, wherein the two more distanced orifices with respect to each other are separated such that at least part of one of the orifices is separated from at least part of the other orifice a distance substantially the same as the width of the sample holders and / or sample holder supports, or has a difference in length with said width of ± 25%.

[0038] According to another preferred embodiment of the second aspect of the invention, each blowing step is related to a blowing profile, each profile defining at least one parameter of the air flow selected from air flow rate, position or distance to the sample, and power applied by the pressure source, preferably wherein the protocol or dispensing protocol comprises a plurality of dispensing steps related each to at least one reagent and optionally to a blowing profile, more preferably said blowing profile determined based on the reagent associated to a dispensing step of the protocol, preferably the previous, current or following dispensing step.

[0039] According to another preferred embodiment of the second aspect of the invention, the automatic staining device further comprises an adjustable orientation mechanism configured to vary manually or automatically the angle at which air is blown with respect to the sample holder.

[0040] According to another preferred embodiment of the second aspect of the invention, the automatic staining device further comprises a protective element for the air blower protecting at least one orifice thereof, configured to protect the air blower from reagent splashes that could cause clogging or partial obstruction of the orifices.

[0041] According to another preferred embodiment of the second aspect of the invention, the air blowing unit is configured to perform multiple passes of air blowing in a given blowing step, each pass having at least one parameter that may differ from a previous pass.

[0042] According to another preferred embodiment of the second aspect of the invention, the automatic staining device further comprises a sensor arrangement for verifying air flow rate or pressure, wherein the processor is configured to adjust the air flow or speed and / or the position of the air blower, preferably with respect to the sample holder, during a dispensing or blowing step.

[0043] According to another preferred embodiment of the second aspect of the invention, the memory further comprises instructions that, when executed by the processor, cause the processor to select a specific blowing step, a blowing profile or to regulate one or more blowing parameters, based on the viscosity of the reagent just dispensed, the type of sample holder, information related to the next reagent dispensing step or on the characteristics of the biological sample. According to another preferred embodiment of the second aspect of the invention, the air blower is coupled to the robotic arm such that the robotic arm can move the air blower from a parking position to a working position, preferably at different distances from the sample holder, so as to adapt to different sections of the protocol or sample geometry.

[0044] In a third aspect of the invention, an automatic staining device for dispensing staining reagents over one or more biological samples is disclosed, wherein one or more reagents are delivered through a controlled pressure-driven flow. The automatic staining device comprises a pressure-driven fluid displacement mechanism; a memory and a processor in communication with said memory, the memory comprising information of at least one protocol, said protocol comprising information related to at least one reagent dispensing step and at least one spreading step; one or more sample holder supports suitable to support one or more sample holders; one or more reagent vials; a robotic arm comprising at least one dispensing probe, wherein the dispensing probe is fluidly connected to the pressure-driven fluid displacement mechanism. The automatic staining device further comprises a spreading unit, preferably comprised in the robotic arm, comprising a non-rotatable spreading element, wherein a longitudinal axis of said spreading element is or can be placed parallel to the surface of the one or more sample holders and / or the surface of the sample holder supports, and wherein the memory comprises instructions that, when executed by the processor, cause the device to perform a spreading step based on the previous reagent dispensing step, more preferably based on the viscosity of the reagent previously dispensed or depending in if the viscosity of the reagent dispensed is above or not a predetermined viscosity threshold.

[0045] According to a preferred embodiment of the third aspect of the invention, the spreading element (31) is coupled to the robotic arm via a mechanism that allows guided displacement along the vertical axis (Z axis) over a limited distance.

[0046] According to a preferred embodiment of the third aspect of the invention, the spreading unit further comprises a supporting element attached to the spreading element, and a supporting piece preferably attached to the robotic arm, wherein the spreading element is supported by the supporting element, and the spreading element can be displaced parallel to a longitudinal axis of the supporting element and / or in the direction perpendicular to the longitudinal axis of the spreading element, between a first configuration wherein the supporting piece supports the weight of the spreading element and optionally also of the supporting element, and a second configuration wherein the spreading element is not supported by the supporting piece, such as when it is supported by the sample holder and / or the sample holder support during a spreading step. According to another preferred embodiment of the third aspect of the invention, the spreading unit further comprises a supporting element attached to the spreading element, and a supporting piece preferably attached to the robotic arm, wherein the spreader supporting element can slide or traverse the supporting piece until a certain point allowing to move the spreading unit the length of the supporting piece or less.

[0047] According to another preferred embodiment of the third aspect of the invention, the spreading support element can tilt an angle <|), preferably between -10 and 10 degrees with respect to the supporting piece.

[0048] According to another preferred embodiment of the third aspect of the invention, the longitudinal length of the spreading element is substantially the same as the width of the sample holders and / or sample holder supports, or has a difference in length size with respect to said width of ± 50%, preferably ± 25%.

[0049] According to another preferred embodiment of the third aspect of the invention, the spreading element is cylindrical or a prism.

[0050] According to another preferred embodiment of the third aspect of the invention, the spreading element comprises a distal part, a proximal part, and a central part, wherein a diameter of the distal and proximal part is the same for both distal and proximal parts, and is bigger than a diameter of the central part, wherein the diameter of the distal and proximal parts is between 0.02 and 2 mm longer than the diameter of the central part, preferably is between 0.06 and 0.6 mm longer, more preferably is between 0.1 and 0.3 mm longer.

[0051] According to another preferred embodiment of the third aspect of the invention, the spreading element is made of a material selected from polymer, metal, ceramic, hybrid composite.

[0052] According to another preferred embodiment of the third aspect of the invention, the spreading element is configured to adapt its height (Z-axis) dynamically during spreading, thereby following the topography of the sample holder or sample, and / or is configured to vary its inclination angle with respect to the Z-axis to optimize reagent spreading.

[0053] According to another preferred embodiment of the third aspect of the invention, the spreading element is coupled to a force control mechanism configured to limit the pressure exerted on the sample holder during spreading, thereby preserving the integrity of fragile substrates or sensitive samples.

[0054] According to another preferred embodiment of the third aspect of the invention, the spreading element is configured to be raised from the sample holder after a spreading step, and displaced to a washing station. According to another preferred embodiment of the third aspect of the invention, the automatic staining device further comprises a washing receptacle suitable to fit the length of the spreading element.

[0055] According to another preferred embodiment of the third aspect of the invention, the washing receptacle comprises a cover or protective element configured to prevent splashing of cleaning solution outside the washing area or over the spreading element or a dispensing probe.

[0056] According to another preferred embodiment of the third aspect of the invention, the washing receptacle is configured to clean the spreading element by at least one of direct fluid dispensing, preferably from one or more dispensing orifices within the washing receptacle, immersion of the spreading element in a cleaning solution, or a combination thereof, preferably for a predetermined period of time based on the previous reagent dispensing steps.

[0057] According to another preferred embodiment of the third aspect of the invention, the automatic staining device further comprises a drying station for the spreading element, said drying station comprising an absorbent surface suitable to remove residual cleaning solution from the spreading element by contact, preferably wherein the spreading element is displaced over the absorbent surface in one or more passes.

[0058] According to a preferred embodiment of the second or third aspect of the invention, the system further comprises at least one camera, and wherein the memory comprises further instructions that, when executed by the processor, cause the processor to: receive one or more images captured by the at least one camera, the images comprising at least one sample or sample holder; determine the presence, size, and / or position of the sample or sample holder in the images using a trained machine learning model; and optionally, adapt at least one parameter of the blowing step and / or the spreading step based on the determined presence, size, and / or position of the sample or sample holder.

[0059] According to a fourth aspect of the invention, an automatic staining device for dispensing staining reagents over one or more biological samples is disclosed. The automatic staining device comprising: a pressure-driven fluid displacement mechanism; a memory and a processor in communication with said memory; one or more sample holder supports suitable to support one or more sample holders; one or more reagent vials; a robotic arm comprising at least one dispensing probe, wherein the dispensing probe is fluidly connected to the pressure-driven fluid displacement mechanism; one or more sensors selected from: accelerometers, gyroscopes, magnetometers, encoders, current sensors, temperature sensors, flow sensors, and capacitive sensors. The system characterized in that the memory comprises instructions that, when executed by the processor, cause the processor to: collect data from at least one or more of said sensors over a temporal window, thereby generating a data matrix of sensor values; andprovide said data matrix as input to at least one embedded artificial intelligence model configured to perform at least one of:

[0060] 1 . classification of the activity of the robotic structure,

[0061] 2. detection of anomalies in the robotic structure,

[0062] 3. detection of inclination or loss of horizontal stability,

[0063] 4. determination of the quality of the support structure,

[0064] 5. estimation of compressor or pump wear,

[0065] 6. detection of activation of hydraulic pumps,

[0066] 7. estimation of heater wear,

[0067] 8. detection of liquid level, wherein the output of the embedded artificial intelligence model is used to generate a diagnostic, warning, or control signal for the operation of the automatic staining device.

[0068] According to another preferred embodiment of the fourth aspect of the invention, the memory comprises a plurality of embedded artificial intelligence models, each configured to process a respective subset of sensor data and to output a respective diagnostic or classification result, wherein the processor is configured to execute said models in parallel or sequentially.

[0069] According to another preferred embodiment of the fourth aspect of the invention, at least one embedded artificial intelligence model is a neural network trained for N-class classification of the activity of the robotic structure, using as input data from at least one of: accelerometer, gyroscope, magnetometer, encoder, and current sensors.

[0070] According to another preferred embodiment of the fourth aspect of the invention, at least one embedded artificial intelligence model is an anomaly detection model configured to detect abnormal movement or operation of the robotic structure, using as input data from at least one of: accelerometer, gyroscope, magnetometer, encoder, and current sensors.

[0071] According to another preferred embodiment of the fourth aspect of the invention, at least one embedded artificial intelligence model is configured to detect inclination or loss of horizontal stability of the device, using as input data from at least one of: accelerometer, gyroscope, magnetometer, and encoder sensors, and to output a warning if the inclination exceeds a predetermined threshold.

[0072] According to another preferred embodiment of the fourth aspect of the invention, at least one embedded artificial intelligence model is a regression model configured to estimate the quality index of the support structure of the device, based on inertial sensor data. According to another preferred embodiment of the fourth aspect of the invention, at least one embedded artificial intelligence model is a regression model configured to estimate the wear of the compressor or hydraulic pumps, based on a comparison of estimated and actual usage time, and / or flow sensor data.

[0073] According to another preferred embodiment of the fourth aspect of the invention, at least one embedded artificial intelligence model is a regression model configured to estimate the wear of the reaction chamber heaters, based on a comparison of applied electrical power and measured thermal output, using as input data from temperature sensors and power measurement data.

[0074] According to another preferred embodiment of the fourth aspect of the invention, at least one embedded artificial intelligence model is a classification model configured to detect the activation state of hydraulic pumps, using as input data from accelerometer, gyroscope, and magnetometer sensors.

[0075] According to another preferred embodiment of the fourth aspect of the invention, at least one embedded artificial intelligence model is a classification model configured to detect the liquid level in a reagent vial, using as input data from a capacitive sensor and a Z-axis encoder, and optionally further comprising as input the Z-axis speed, Z-axis position, liquid identification, and / or a basic liquid detection algorithm.

[0076] According to a fifth aspect of the invention, an automatic staining device for dispensing staining reagents over one or more biological samples is disclosed, the automatic staining device comprising: a pressure-driven fluid displacement mechanism; a memory and a processor in communication with said memory; one or more sample holder supports (42) suitable to support one or more sample holders (41); one or more reagent vials (51); a robotic arm (10) comprising at least one dispensing probe (80), wherein the dispensing probe (80) is fluidly connected to the pressure-driven fluid displacement mechanism; at least one inertial sensor selected from: accelerometer, gyroscope, and magnetometer. Wherein the device is characterized in that the memory comprises instructions that, when executed by the processor, cause the processor to: collect data from the at least one inertial sensor during execution of a staining protocol; provide said data to an embedded artificial intelligence model trained to detect loss of horizontal stability or excessive inclination of the device or its support table; compare the output of the artificial intelligence model to a predetermined threshold; and if the output indicates a loss of stability or inclination exceeding the threshold, generate a warning signal and / or interrupt the staining protocol. According to another preferred embodiment of the fourth aspect of the invention, the warning signal is displayed on a user interface and / or transmitted to a remote monitoring system.

[0077] Brief description of the drawings

[0078] To enable a better understanding of the present disclosure, and to show how the present disclosure may be carried out, reference will now be made, by way of example only, to the accompanying schematic drawings, wherein:

[0079] Figure 1 shows a drawing of an automatic staining device according to one or more embodiments of the invention.

[0080] Figure 2 shows a drawing of part of the robotic arm of an automatic staining device according to one or more embodiments of the invention, wherein the robotic arm comprises or has attached a camera, a spreading unit and a wash nozzle or probe.

[0081] Figure 3 shows a drawing of a close up of the spreading unit, wherein the spreading element is shown as a horizontal cylinder, according to one or more embodiments of the invention.

[0082] Figure 4 shows a drawing of an even further close up of the spreading element of the spreading unit according to one or more embodiments of the invention.

[0083] Figures 5, 6 and 20 show a drawing of the part of the robotic arm wherein the spreading unit is attached, and below it is shown a sample holder support with a sample holder on top, according to one or more embodiments of the invention. Figure 5 shows the spreading unit being supported fully by the robotic arm, whereas in Figure 6 it can be appreciated that the spreading unit, in particular the spreading element and the supporting element has been vertically displaced with respect to the robotic arm when entering into contact with the sample holder, which now supports the weight of the spreading unit. Figure 20, on the other hand, show a tilt angle <|) of the spreading element and / or supporting element with respect to the axis perpendicular to the surface of the sample holder (i.e. , normal to the surface) when the device is sweeping or movind the spreading element, according to one or more embodiments of the invention.

[0084] Figure 7 discloses a diagram of the steps followed by the processing of the automatic staining device of the invention for a smart or intelligent dispensing, according to one or more embodiments of the invention. Figure 8 shows an example of sample holder recognition and image processing for label reading, according to one or more embodiments of the invention.

[0085] Figure 9 shows an example of the image of the lid of a reagent vial and image processing for label reading, according to one or more embodiments of the invention.

[0086] Figure 10 shows a sample holder comprising a grid of apertures, which can be used to calibrate the camera and the image processing, according to one or more embodiments of the invention.

[0087] Figure 11 shows an example of a methodology to determine if the position observed of a sample holder coincides with the expected position or the position of the sample holder support previously calibrated, according to one or more embodiments of the invention.

[0088] Figure 12 shows a drawing of a row of sample holders supports comprising each a sample holder with a label, and a signalling LED, and it is further shown a close up of one sample holder support, according to one or more embodiments of the invention.

[0089] Figure 13 shows a drawing of a sample holder rack, comprising 14 columns of sample holders, each one comprising 3 rows of sample holders, according to one or more embodiments of the invention.

[0090] Figure 14 shows a drawing of a reagent vial rack, comprising 7 columns of 8 rows each of supports for reagent vials, wherein the first and eight row are filled with reagent vials, according to one or more embodiments of the invention.

[0091] Figure 15 shows a drawing of a detail of the robotic arm that comprises the air blowing unit and the spreading unit, according to one or more embodiments of the invention.

[0092] Figure 16 shows a drawing of a close up of an air blower of an air blowing unit, according to one or more embodiments of the invention.

[0093] Figure 17 shows a drawing of a close up of the dispensing probe comprised in the robotic arm, according to one or more embodiments of the invention.

[0094] Figure 18 shows a close up of a reagent vial with the lid open according to one or more embodiments of the invention.

[0095] Figure 19 shows a washing station according to one or more embodiments of the invention.

[0096] Figure 20 show a drawing of the part of the robotic arm wherein the spreading unit is attached, and below it is shown a sample holder support with a sample holder on top, according to one or more embodiments of the invention. In figure 20 it can be appreciated a tilt angle <|) of the spreading element and / or supporting element with respect to the axis perpendicular to the surface of the sample holder (i.e., normal to the surface) when the device is sweeping or moving the spreading element, according to one or more embodiments of the invention.

[0097] Figure 21 shows a drawing of a close up of a robotic arm comprising a spreading unit and a blowing unit which can move relatively to each other at least in the vertical direction, according to one or more embodiments of the invention.

[0098] Description of the invention

[0099] Definitions

[0100] As used herein, “a” or “an” means “at least one” or “one or more.”

[0101] It is noted that the term “about”, if used herein, refers to + / - 30%, preferably + / - 20%, preferably + / - 15%, more preferably + / - 10%, of the indicated referred value. It may also refer to the exact value, that is, + / - 0% or + / - the standard measurement error. Also, any quantity, composition percentages and / or dimension mentioned herein is understood to present an error of + / - 30%, preferably + / - 20%, preferably + / - 15%, more preferably + / - 10%, more preferably + / - 5% or + / - 0% of the indicated value. It may also refer to the exact value, that is, + / - 0% or + / - the standard measurement error.

[0102] As used herein, the conjunctive term "and / or" between multiple recited elements is understood as encompassing both individual and combined options. For instance, where two elements are conjoined by "and / or", a first option refers to the applicability of the first element without the second. A second option refers to the applicability of the second element without the first. A third option refers to the applicability of the first and second elements together.

[0103] When used herein the term "comprising" can be substituted with the term "containing" or "including" or sometimes when used herein with the term "having".

[0104] When used herein "consisting of' excludes any element, step, or ingredient not specified in the claim element. When used herein, "consisting essentially of” does not exclude materials or steps that do not materially affect the basic and novel characteristics of the claim.

[0105] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as is commonly understood by one of ordinary skill in the art to which this invention belongs. All patents, applications, published applications and other publications referred to herein are incorporated by reference in their entirety. If a definition set forth in this section is contrary to or otherwise inconsistent with a definition set forth in applications, published applications and other publications that are herein incorporated by reference, the definition set forth in this section prevails over the definition that is incorporated herein by reference.

[0106] The term "staining reagents", in the context of the invention, refers to any chemical substances or mixtures used to stain biological samples, including but not limited to dyes, antibodies, enzymes, aqueous solutions or hybridization reagents. These reagents are applied to biological samples such as tissue sections, cell smears, or other biological materials in order to highlight or differentiate structures for observation under a microscope. Staining may include, but is not limited to, histological staining, immunoenzymatic staining, fluorescent staining and hybridization techniques, and is employed for diagnosing diseases, conducting research, and other scientific analyses.

[0107] The term “automatic staining device”, in the context of the invention, refers to any apparatus configured to automatically perform one or more steps of a protocol for the staining process of biological samples, including but not limited to the delivery, incubation, washing, or drying of reagents and / or fluids. In some embodiments, the automatic staining device can also blow air over the samples and / or apply a mechanical spreader over the reagent and / or sample holder. The device is adapted to handle staining reagents as defined herein, as well as other fluids such as buffers, rinsing solutions, or drying media, by means of mechanical, fluidic, or electronic components that operate under programmed or sensor-based control, preferably dispensed through pressure-driven fluid displacement mechanism such as fluid pumps. The automatic staining device may incorporate pumps, valves, robotic actuators, or similar systems to ensure precise and reproducible application of reagents or fluids over the biological samples positioned on suitable sample holders, regardless of the specific configuration or type of staining technique employed.

[0108] The term "pressure-driven pumped flow", in the context of the invention, refers to the mechanism by which reagents are transported and dispensed over biological samples, utilizing a pump to create a controlled flow of liquid. This flow is driven by pressure exerted by the pump, which may include, but is not limited to, piston-based pumps, syringe-type pumps, peristaltic pumps, diaphragm pumps, or any similar mechanism that displaces fluid. Unlike capillary flow, which relies on passive forces within narrow tubes, a pressure-driven pumped flow ensures consistent and directed movement of reagents, allowing for precise application and control.

[0109] The term "sample holder supports", in the context of the invention, refers to the structural components designed to support and / or hold sample holders during the staining process. These supports may include features such as orifices or grids that allow excess reagent to flow through, preventing fluid accumulation between the sample holders and the support structure. In some embodiments, the sample holder supports may also comprise additional components such as inclination sensors to monitor and adjust the position of the samples, or temperature control units to regulate the environment during staining.

[0110] The term "sample holders", in the context of the invention, refers to the structures used to contain and position biological samples during the staining process. These may include, but are not limited to, glass slides, well plates, or other substrates suitable for holding biological specimens such as tissue sections or cell smears. The sample holders preferably comprise a label or handwritten notations indicating information regarding the sample or the reagents or the staining procedure that needs to be applied over the sample comprised in the sample holder. For example, a staining process may comprise a protocol intended to identify cellular or subcellular structures and / or molecules / biomarkers by staining or colouring methods, such as, but not limited to, using colorimetric dies and / or fluorescence dies.

[0111] The term "biological sample", in the context of the invention, refers to any material of biological origin that is intended to be stained for diagnostic, research, or analytical purposes. This may include tissue sections, cell smears, cytology samples, or other biological materials that require staining for enhanced visualization. The biological samples are typically prepared on sample holders and may undergo various types of staining processes to highlight cellular structures, proteins, nucleic acids, or other biological components.

[0112] The term "reagent vials", in the context of the invention, refers to containers designed to hold the staining reagents used in the automatic staining process. These vials are preferably equipped with a lid that can be moved between an open and closed state, allowing the withdrawal or dispensing probe to access the reagent inside when in the open state, and preventing it in the closed state. In some embodiments, the system may detect the open or closed status of the lid to ensure that the process only begins when the lid is open, optionally providing an alert or prompt to the user otherwise. In other embodiments, the vials may be equipped with a membrane that allows the probe to pass through without the need to open the lid, therefore not needing the feature to detect the open and closed status and ensuring a continuous and automated process without manual intervention.

[0113] The term "air blower", in the context of the invention, refers to a device designed to direct airflow over a biological sample or sample holder, preferably to remove excess reagent either by drying it and / or by displacing it outside the boundaries of the sample holder. The air blower may comprise one or more orifices through which the air is blown, may comprise an angled air exit to direct the air towards the sample and sample holder at an angle in between the horizontal and vertical directions, more preferably between 30 and 60 degrees with respect to the vertical direction. The air blower is preferably shaped and configured to blow air over the entire width of the sample holder, more preferably in a substantially evenly way.

[0114] The term "air flow sensor", in the context of the invention, refers to a sensor that measures and allows to regulate the flow of air generated by the air blower. This sensor preferably allows for the detection of the quantity and / or speed of air being blown, and the controller or processor receiving the information from the air flow sensor can calibrate the distance of the ait blower with respect to the sample holder, and the duration of the airflow to determine the dryness or moisture level of the sample holder.

[0115] The term "mixing vials", in the context of the invention, refers to containers used for mixing two or more reagents prior to their application onto biological samples. These vials allow the preparation of custom or pre-programmed reagent combinations in an automated manner to ensure the correct formulation is dispensed during the staining process.

[0116] The term "fluid pump", in the context of the invention, refers to a mechanism used to create pressure-driven flow for the transport of reagents from the reagent vials to the dispensing probe and ultimately onto the biological samples. This pump may include various types of pumps, such as syringe pumps, peristaltic pumps, or diaphragm pumps, that enable precise control of reagent flow during withdrawal, mixing, and dispensing operations. A fluid pump is thus a pressure-driven fluid displacement mechanism.

[0117] The term "moisture level", in the context of the invention, refers to the amount of liquid remaining on the surface of the biological sample or sample holder after reagent application and subsequent removal by the air blower and / or the spreading element. An optimal moisture level allows the next reagent to spread easily over the sample surface, while a sample that is too dry can hinder the proper distribution of additional reagents, potentially affecting the quality of the staining process. The term may encompass states ranging from completely dry (no detectable liquid), through various intermediate states of partial wetness or residual dampness, to fully saturated or completely wet conditions. The term may encompass, for example, a thin residual film of liquid, visible droplets, a measurable percentage of water content by weight or volume, or a specific relative humidity at the surface, such as, but not limited to, 0% (dry), 10%, 50%, or 100% (fully wet), as determined by gravimetric, optical, electrical, or sensor-based methods, among other options.

[0118] The term "dispensing probe", in the context of the invention, refers to the component responsible for transferring reagents from the reagent vials to the sample holders. In some embodiments, the dispensing probe is capable of both withdrawing reagents from the vials and dispensing them over the samples, enabling a dual function in reagent handling, whereas in other embodiments may be fluidly connected to a withdrawal probe that carries on the function of withdrawing the reagents and providing them to the dispensing probe. The dispensing probe is preferably fluidly connected to the fluid pump which ensures accurate and controlled reagent application.

[0119] The term "trained machine learning model", in the context of the invention, refers to a computational model that has been trained using datasets of images or other relevant data to perform tasks such as identifying the presence, position, orientation, and size of sample holders and / or reagents, wherein the dataset of images preferably comprises labelled images used fortraining of the different possibilities that can be found (such as images of reagent vials open and closed, different positions and orientations of the sample holders, etc) and also test images to determine the accuracy of the model. The model or models can thus be trained to recognize specific labels or conditions, such as the open or closed state of reagent vials, based on the input provided from the device’s cameras. There may be different models for each task, or a model that is trained to carry out more than one task, and / or multimodal models that can receive different types of input at the same time, or different types of images.

[0120] The term "open or closed state of the lid", in the context of the invention, refers to the status of the lid covering the reagent vials, indicating whether the lid is in a position that allows access for the dispensing probe to withdraw reagents (open) or not (closed). The system may detect the open or closed state to either proceed with or pause the staining process. In some embodiments, the lid may be equipped with a membrane that permits the probe to pass through without needing to open the lid, therefore not needing the system to determine such state.

[0121] The term "supporting element", in the context of the invention, refers to a component of the spreading unit that supports the spreading element. The supporting element can be displaced relative to the spreading element when the spreading element is in contact with the sample holder. This displacement prevents excessive pressure from being applied to the sample holder, reducing the risk of damaging or breaking the sample holder during the reagent spreading process.

[0122] The term "spreading element", in the context of the invention, refers to the part of the spreading unit responsible for distributing reagents over the surface of the sample or sample holder. The spreading element is preferably cylindrical. More preferably, the spreading element comprises a central part having a smaller diameter than the distal and proximal parts, allowing the central part to spread an even and uniform layer of reagent, for example, around 0.1 mm, while the distal and proximal parts are suitable to rest on the edges of the sample holder. However, other configurations are possible as long as they have a similar functionality, such as a flange with a central part with a smaller length, a prism, or even a cylinder that has a bevel or a transverse section parallel to the longitudinal axis of the cylinder, or a symmetric normal cylinder.

[0123] The term "inclination sensors", in the context of the invention, refers to sensors that detect the angle or tilt of the sample holder supports, or the automatic staining device or its components with respect to the vertical or horizontal plane. These sensors may include, but are not limited to, accelerometers, tilt sensors, inclinometers, gyroscopes, inertial measurement units (IM Us), angle sensors, orientation sensors, or gravitational sensors. Inclination sensors ensure the proper alignment of the sample holders during the staining process, greatly improving consistency in reagent application.

[0124] The term “pressure-driven fluid displacement mechanism”, in the context of the invention, refers to any system or assembly configured to move or dispense fluids by applying a controlled pressure differential to the fluid, thereby causing its displacement through conduits, channels, or dispensing elements. The term may encompass mechanisms such as, but not limited to, positive displacement pumps, peristaltic pumps, piston pumps, diaphragm pumps, syringe pumps, gear pumps, pressure vessels, compressed air or gas-driven systems, or combinations thereof, and may operate using pneumatic, hydraulic, or mechanical actuation. The pressure applied is preferably generated by electric motors, however in some embodiments may be generated by manual actuation, compressed gas sources, or other suitable means, and may be regulated to achieve precise control of flow rate, volume, or dispensing timing for a wide range of fluid viscosities and reagent types.

[0125] The term “protocol”, in the context of the invention, refers to a set of instructions, parameters, or data sequences that define the order, timing, and conditions for dispensing one or more reagents or fluids, and in some embodiments may as well perform associated steps such as blowing, spreading, washing, and / or drying during an automated staining process. The protocol or dispensing protocol may encompass, for example, information specifying reagent types, dispensing volumes, flow rates, dispensing locations, sequence of application, duration of each step, air flow profiles, spreading parameters, drying times, and sample-specific adjustments, as well as conditional logic or adaptive steps based on sensor feedback or sample characteristics, such as, but not limited to, protocols for immunohistochemistry, cytology, in situ hybridization, or special stains for histological or cytological samples.

[0126] The term “blowing step”, in the context of the invention, refers to a process stage during which a flow of gas, for example air, nitrogen, or another inert or reactive gas, is directed over a sample, sample holder, or reagent layer to achieve an effect such as, but not limited to, removing excess reagent, spreading a reagent, partially or fully drying a surface, homogenising a liquid layer, or controlling the moisture level. The blowing step may encompass single or multiple passes, may be performed at variable flow rates, pressures, temperatures, or angles, and may be executed before, after, or between reagent dispensing or spreading steps. The gas may be delivered through one or more orifices, nozzles, air knives, or diffusers, and the step may be adapted based on parameters such as reagent viscosity, sample type, or protocol requirements.

[0127] The term “blowing profile”, in the context of the invention, refers to a set of one or more parameters that define the characteristics of an air blowing step applied to a sample or sample holder, where such parameters may encompass, for example, air flow rate or speed, air pressure, temperature, humidity, duration of blowing, speed of blowing, position or distance of the air blower relative to the sample or sample holder support, angle of air incidence, number of passes, sequence of passes, and power applied by the pressure source. The term may encompass profiles tailored for specific reagents, sample types, or protocol steps, such as, but not limited to, profiles for rapid drying, gentle moisture retention, even reagent spreading, or selective removal of excess liquid, and may be implemented as pre-set, user-defined, or dynamically adjusted parameter sets stored in memory or determined in real time by the device.

[0128] The term “reagent dispensing step”, in the context of the invention, refers to a discrete action or programmed event within a staining protocol in which a specific volume of a reagent is delivered to a biological sample or sample holder by the device. The term may encompass actions such as, but not limited to, dispensing staining solutions, washing buffers, fixatives, blocking agents, counterstains, or mounting media, and may involve delivery by pressure- driven flow, syringe pumps, peristaltic pumps, or other fluid displacement mechanisms. A reagent dispensing step can be defined by parameters such as reagent type, volume, flow rate, dispensing position, timing, and sequence relative to a dispensing step of the protocol, preferably a previous, current, or following reagent dispensing steps, and may be executed in combination with or independently from other protocol steps such as air blowing or mechanical spreading.

[0129] The term "information regarding the biological sample," in the context of the invention, refers to any data, metadata, or parameters that describe, identify, or characterize the biological sample to be processed by the device. The term may encompass, but is not limited to, sample type (for example, tissue section, cell smear, cytology specimen, or blood extension), sample origin (for example, human, animal, plant, or microorganism), sample dimensions or geometry, sample thickness, fixation method, staining history, sample identification codes, barcode or RFID data, sample age, storage conditions, sample preparation protocol, and any relevant morphological or physical properties that may influence reagent dispensing, spreading, or drying steps.

[0130] The term “washing receptacle”, in the context of the invention, refers to a container or vessel configured to receive, hold, and / or facilitate the cleaning of one or more components of the automatic staining device, such as, but not limited to, spreading elements, dispensing probes, or sample holders, preferable of a spreading element, by containing a cleaning solution or enabling direct fluid dispensing, immersion, or rinsing. The washing receptacle may encompass structures such as open or closed tanks, troughs, wells, basins, or chambers, and may be constructed from materials such as, but not limited to, stainless steel, glass, polymer, ceramic, or composite materials. The washing receptacle may further comprise features such as, but not limited to, integrated fluid inlets or outlets, covers or splash guards, racks or holders for positioning components, or drainage systems, and may be dimensioned or shaped to accommodate components of various sizes and geometries.

[0131] The term "force control mechanism", in the context of the invention, refers to any system, device, or arrangement configured to monitor, regulate, or limit the amount of force applied by a component, for example during a spreading, dispensing, or contact step, so as to prevent excessive pressure or damage to a target surface or object. The term may encompass mechanical, electromechanical, pneumatic, hydraulic, or electronic systems such as, but not limited to, load cells, strain gauges, force sensors, spring-based limiters, torque limiters, compliant actuators, servo-controlled motors with feedback, piezoelectric sensors, or softwarebased control algorithms that adjust actuator output based on real-time force measurements. The force control mechanism may operate in open-loop or closed-loop configurations and may be integrated into robotic arms, end effectors, or other moving parts of the device.

[0132] Description

[0133] The main challenges faced by current automatic staining devices, particularly those relying on capillarity-based systems, revolve around frequent maintenance due to reagent crystallization, cumbersome cleaning processes, and the difficulty in accessing internal components. Noncapillary alternatives, while attempting to resolve these issues, introduce new problems, such as the excessive sensibility to instabilities and inclination, increased reagent consumption, and the difficulty in handling viscous reagents. Current devices also often lack full automatization and precision in reagent dispensing, leading to wasted materials and increased risk of contamination, while still requiring significant user intervention for configuration and verification, which introduces the potential for human error. There is a need for a fully automated, smart staining device that eliminates the issues associated with capillarity, controls reagent usage efficiently, ensures uniform reagent application, and automates the entire process, significantly reducing user intervention and errors.

[0134] In this invention we disclose a device that addresses the aforementioned issues and successfully solves them.

[0135] Each embodiment disclosed herein is contemplated as being applicable to each of the other disclosed embodiments. Thus, all combinations of the various elements described herein are within the scope of the invention. It should also be understood that, unless clearly indicated to the contrary, in any methods claimed herein that include more than one step or act, the order of the steps or acts of the method is not necessarily limited to the order in which the steps or acts of the method are recited.

[0136] In a first aspect of the invention, an automatic staining device (1) for dispensing staining reagents or fluids over one or more biological samples is disclosed. Contrary to devices that use a capillary flow, in the device (1) of the invention one or more reagents or fluids are delivered through a controlled pressure-driven flow, since the automatic staining device (1) comprises a pressure-driven fluid displacement mechanism. The controlled pressure-driven flow enables precise and efficient reagent delivery, addressing the problems of reagent crystallization and the maintenance complexities seen in capillary-based systems.

[0137] In some embodiments, the pressure-driven fluid displacement mechanism may comprise various types of devices or fluid pumps that manage fluid flow through the application of pressure. This may comprise a motorized syringe, where a motor controls the movement of the syringe plunger, thereby regulating the flow rate and volume of the reagent being dispensed. This is particularly useful when dealing with viscous reagents or when precise reagent volumes are needed to avoid reagent wastage or over-application. Another possibility is the use of a peristaltic pump, in which fluid is propelled through flexible tubing by compressive rollers, enabling smooth, continuous flow without the risk of backflow and ensuring the reagent remains uncontaminated by the pump system itself. In some embodiments both types of pressure-driven fluid displacement mechanisms may be employed, for example for different reagents. A pneumatic pump may alternatively be used, wherein compressed air displaces the reagent from a reservoir towards the dispensing probe. Other options comprise a diaphragm pump, wherein the displacement of the reagent is achieved by the flexing of a diaphragm, creating the necessary pressure to move the fluid.

[0138] The controlled pressure-driven flow may be applicable to a wide variety of staining procedures, including but not limited to histological staining, immunoenzymatic staining, hybridization techniques, and / or cytological staining. For histological staining, the device may be used for procedures such as Hematoxylin and Eosin staining, Masson’s trichrome staining, or periodic acid-Schiff staining, which are commonly employed to visualize cellular structures in tissue sections. In the case of immunoenzymatic staining, the device may be configured to handle staining processes such as immunohistochemistry or enzyme-linked immunosorbent assays, where antigen-antibody reactions are used to visualize specific proteins or markers within cells or tissues. The device may further support hybridization techniques, such as in situ hybridization, where specific DNA or RNA sequences are detected in cells or tissues. Additionally, the device may be suitable for cytological staining, such as the Papanicolaou staining process used to stain cell smears and detect cellular abnormalities.

[0139] It is further noted that the device (1) may be configured to dispense reagents onto a variety of biological samples, including tissue sections, cell smears, and other biological materials. The pressure-driven fluid displacement mechanism may also be controlled by a processor to dynamically adjust flow rate, volume, and pressure based on the specific reagent, sample size, or staining protocol in use.

[0140] In some embodiments, the pressure control system may be equipped with sensors that monitor the pressure within the fluid displacement mechanism, ensuring precise control during the reagent dispensing process. Sensors such as pressure transducers or flow sensors may be employed to adjust pressure in real-time, preventing over-pressurization or clogging. Furthermore, the fluid displacement mechanism may be configured to handle reagents of varying viscosities, offering flexibility in the types of reagents that can be used during the staining procedure.

[0141] The automatic staining device (1) further comprises a processor in communication with a memory. The processor can manage and control various processes involved in the automatic staining procedure. In some embodiments, the processor may be configured to run machine learning algorithms or geometric algorithms to process data, such as images captured by the camera (20), to determine the orientation and placement of sample holders (41) and reagent vials (51). The memory may store staining protocols, image data, or other operational parameters, ensuring that the system can function autonomously and adjust the procedure based on real-time feedback. The processor may also be in communication with various sensors or external control systems, expanding its ability to regulate different parts of the device, such as the fluid displacement mechanism or air blower unit. In alternative embodiments, the processor may support remote connectivity, allowing external devices to upload protocols, monitor the staining process, or intervene manually if necessary. The automatic staining device (1) comprises in some embodiments at least one camera (20). The inclusion of at least one camera (20) is critical for enabling the automated detection of important elements such as the sample holders (41) and reagent vials (51). The camera (20) may capture images or video data, which in some embodiments are processed by the processor to determine whether sample holders (41) are correctly placed, whether reagent vials (51) are present and properly aligned, and / or whether vial lids (52) are in an open or closed state. In some embodiments, the camera (20) may be a high-resolution digital camera, a multi-spectral camera, or an infrared camera, depending on the type of analysis required. Alternatively, multiple cameras may be used, positioned at different angles to ensure full visibility of the working area. The camera (20) may also be equipped with optical zoom or autofocus capabilities to enhance its ability to capture fine details, such as small labels or barcodes on sample holders (41) or reagent vials (51).

[0142] The automatic staining device (1) further comprises one or more sample holder supports (42) suitable to support one or more sample holders (41), the sample holders (41) preferably comprising at least one biological sample.

[0143] The sample holder support (42) is designed to hold one or more sample holders (41), which may contain biological samples, and may comprise orifices, preferably in a grid like configuration, to let the excess reagent through in order to not compromise the stability or position of the sample holder (41), or may comprise grooves, for the same purpose. In some embodiments, the sample holders (41) may be glass slides, as commonly used in laboratory environments for histological or cytological analysis. These glass slides may be labelled, either with handwritten information or printed labels, which provide details about the type of staining procedure, sample identification, or other relevant data. The labels may also comprise barcodes or QR codes, enabling the camera (20) to automatically read and identify the correct staining protocol or reagents needed. In some embodiments the sample holder support (42) may be configured to support other types of sample holders (41) as well, including well plates, Petri dishes, or other container formats depending on the type of biological sample or staining procedure. In some embodiments, the support may be designed to adjust its size or configuration to accommodate different sample holder formats, or to support multiple holders simultaneously for high-throughput analysis. In some embodiments, the sample holder support (42) may also include temperature regulation features, such as heating or cooling elements, to ensure optimal conditions for the biological sample during the staining process.

[0144] The automatic staining device (1) further comprises one or more reagent vials (51). In some embodiments said vials (51) comprise a lid (52), wherein the lid (52) is movable between an open and a closed state. The reagent vials (51) contain staining reagents that can be dispensed over the biological samples. The reagent vials (51) comprise a lid (52) that is movable between an open and a closed state, preferably moved by the user, but in some embodiments the device may automatically open the lid (52) when needed to access the reagent. In some embodiments, the lid (52) may be motorized or spring-loaded, controlled by the processor to ensure precise timing and opening for reagent withdrawal. Alternatively, the lid (52) may be manually operable or involve mechanical systems such as a sliding or pivoting mechanism. Additionally, the reagent vials (51) may comprise an orifice or outlet port (54), which allows a dispensing probe to enter the vial (51) when the lid (52) is open, facilitating the withdrawal of the reagent without fully removing the lid (52). This configuration helps maintain the integrity of the reagent by minimizing exposure to air or contaminants. The design of the lid (52) may vary depending on the reagent's properties, with options such as screw caps, snap-fit lids, or hinged covers being possible alternatives.

[0145] In some embodiments, the reagent vials (51) may be designed to accommodate different reagent types, including liquid, viscous, or powdered reagents, preferably liquid. They may be made from various materials, such as glass, plastic, or other chemically inert substances, to prevent any adverse reactions with the reagents. The vials (51) may also feature transparent or translucent sections, allowing the user, the camera (20) or other sensors to detect the remaining reagent volume and trigger a refill or alert when levels are low. Alternatively, the vials (51) or the robotic arm (10) may have integrated sensors to monitor reagent levels directly, which can be communicated to the processor for inventory management or process control.

[0146] The automatic staining device (1) further comprises in some embodiments one air blowing unit comprising an air blower (60) and an air flow sensor, wherein the air blowing unit is configured to remove reagent from the sample and / or sample holders by blowing air over said reagent.

[0147] The air blowing unit comprises an air blower (60) and preferably also an air flow sensor, which together allow for the precise regulation of air flow over the reagent dispensed, the sample or sample holders (41). The air blowing unit may be configured to remove excess reagent by either drying the reagent or displacing it out of the sample holder by blowing air or other gases, such as nitrogen or carbon dioxide. The air flow sensor plays a role in monitoring the air velocity and adjusting it as necessary to ensure optimal conditions for removing excess reagent without disturbing the sample itself. In some embodiments, the air blower (60) may be positioned at different angles or may feature multiple air outlets to ensure an even distribution of air over the sample, allowing for uniform drying or displacement of the reagent. The air blower may also have adjustable power settings to accommodate different types of reagents or sample conditions, ensuring flexibility in various staining procedures. The automatic staining device (1) preferably comprises one or more mixing vials, which, for example, allows providing a dedicated space for mixing different reagents before they are applied to the biological sample. The mixing vials may be fluidly connected to the pressure- driven fluid displacement mechanism, allowing the reagents to be mixed automatically under controlled conditions, or be independent vials. These vials may be made of inert materials, such as glass or plastic, to prevent contamination or reaction with the reagents. In some embodiments, the mixing process may be controlled by the processor, allowing for precise ratios of different reagents to be mixed depending on the staining protocol. The mixing vials may be configured to handle reagents of various viscosities, and they may include sensors to monitor the mixing process, such as optical or viscosity sensors, to ensure homogeneity before dispensing the mixed reagents.

[0148] The automatic staining device (1) further comprise a robotic arm (10) comprising at least one dispensing probe (80), wherein the dispensing probe (80) is fluidly connected to the pressure- driven fluid displacement mechanism. The robotic arm (10) comprises at least one dispensing probe (80), wherein the dispensing probe (80) is fluidly connected to the pressure-driven fluid displacement mechanism. This connection allows the dispensing probe (80), in some embodiments, to withdraw a reagent from a reagent vial (51) and dispense the reagent into a sample, a sample holder (41), or a mixing vial. The robotic arm (10) may be configured to move with multiple degrees of freedom, ensuring precise placement of the dispensing probe (80) over the target area. In some embodiments, the robotic arm (10) may include additional joints, dispensing probes (80 or extendable segments to reach multiple sample holders (41) or reagent vials (51) in a high-throughput system. The movement of the robotic arm (10) can be controlled by the processor, which may use data from the camera (20) or sensors to ensure that the probe is correctly aligned with the reagent vials or sample holders.

[0149] In some embodiments the dispensing probe (80) may further be fluidly connected to at least one reagent withdrawal probe, to streamline the process of withdrawal and dispensing. In an alternative configuration, the dispensing probe (80) may be capable of withdrawing reagents from a reagent vial (51) and then dispensing the same or mixed reagents onto a sample, a sample holder (4!) or into a mixing vial. In some embodiments, the dispensing probe (80) may be interchangeable, or present interchangeable tips or nozzles that can be selected based on the viscosity of the reagent or the size of the dispensing area required. Alternatively, the device may include separate probes for withdrawal and dispensing, depending on the operational requirements or the staining protocols used.

[0150] The automatic staining device (1) may be configured to withdraw one or more reagents from the reagent vials (51), optionally mix two or more reagents in one or more mixing vials, and dispense the withdrawn or mixed reagents over one or more sample holders (41). This flexibility allows for a range of staining protocols to be conducted automatically, with minimal user intervention. In some embodiments, the processor may determine the required reagents for each sample based on data captured by the camera (20) such as the labels on the reagent vials (51) or the sample holders (41), such as the type of sample or the staining protocol associated with the sample. The system may also allow for dynamic adjustments to the mixing or dispensing process, based on real-time feedback from sensors monitoring the sample or reagent conditions.

[0151] Preferably, the automatic staining device (1) or the robotic arm (10) is configured to: withdraw one or more reagents from the one or more reagent vials (51); optionally, mix two or more reagents in the one or more mixing vials; and dispense the one or more withdrawn reagents or the mixed reagents over one or more sample holders (41). This is preferably done with the dispensing probe (80).

[0152] In some embodiments, the air blowing unit is further configured to blow air over the reagent only until a moisture level is left on the sample and / or sample holder (41) adequate to facilitate further reagent dispensing over said sample and / or sample holder (41), wherein the adequate moisture level is determined based on the flow of air sensed by the air flow sensor. This moisture level is of great importance to facilitate the proper dispensing and spreading of subsequent reagents over the sample.

[0153] In some embodiments, the adequate moisture level may be determined by the air flow sensor, which monitors the flow of air blown over the reagent and provides feedback to the processor. The system may be configured to previously calibrate this moisture level based on empirical and / or theoretical data or specific staining protocols. For example, the moisture level may be calculated as a function of the air flow sensed by the air flow sensor, and optionally, it may also depend on the time duration of the air blowing. For example, the longer the air is blown, the drier the sample becomes, allowing the processor to modulate the air flow and time accordingly. Additionally, some blowing flow speeds may be best suited for drying the sample, whereas higher ones can be best suited for disperse and distribute the reagent over the sample or sample holder (41).

[0154] The system may also take into consideration the distance between the air blower and the sample holder (41). In some embodiments, the distance may remain constant during the air blowing process, simplifying the calculation of the moisture level based on flow rate and time. However, in alternative embodiments, the distance between the blower and the sample holder may change dynamically, requiring the processor to adjust the air flow accordingly. For example, if the air blower is positioned farther from the sample holder, it may increase the air flow to compensate for the greater distance, while reducing air flow when closer to the sample to prevent over-drying.

[0155] Additionally, the air blower may move along the length of the sample holder (41) during the air blowing process, ensuring even distribution of air across the entire surface. In some embodiments the blower (60) may rotate or adjust its blowing power to compensate for differences in distance when blowing from the bottom to the top of the sample holder. The movement and rotation may be controlled by the processor to ensure that the reagent is properly dispersed across the sample, avoiding excessive drying or uneven reagent distribution.

[0156] In some embodiments, the air blower (60) may be positioned at an angle relative to the surface of the sample holder (41), or may have a shape or an exiting orifice or slit (64) that is oriented such that the air does not arrive perpendicular to the sample or sample holder (41), but with an angle, preferably the angle, with respect to the normal vector of the plane define by the sample holder (41) i.e., the vertical direction, is between 10° and 90°, preferably between 20 and 80°, more preferably between 30° and 70°, even more preferably between 35° and 60°, even more preferably between 35° and 55°, even more preferably around or equal to 45°. This angular configuration facilitates the dispersion of the reagent across the sample while drying it to the adequate moisture level. In some embodiments the angle may be adjustable depending on the reagent type, the surface area of the sample, or other factors specific to the staining protocol. For example, the blower may be positioned at a 30° to 45° angle with respect to the sample holder, ensuring that the reagent is spread evenly without completely drying out the sample.

[0157] The air blower may also comprise in some embodiments one or more orifices or slits (64), which enable even drying or blowing of air over the sample holder (41). In some embodiments, the blower may include one or more slits (64) or one or more orifices , for example with the width of the slit (64), or the distance between the most distal orifices with respect to each other selected to provide uniform air distribution over the sample holder (41), at least in the longitudinal direction of the sample holder (41) or over its width. Preferably, the width of the slit or the distance between the most distal orifices is similar to, or within a specific range of, the width of the sample holder (41). This range may be within ±50%, ±25%, or more preferably within ±15% or ±10% of the width of the sample holder, ensuring an even drying process across the entire surface and even distribution or displacement of reagent. This configuration reduces the risk of uneven moisture levels on different parts of the sample holder, which could otherwise interfere with the proper spreading of subsequent reagents. The system may also comprise in some embodiments ambient humidity or temperature sensors, which could affect the drying process, and can for example adjust the air blowing process accordingly. For instance, in a high-humidity environment, the blower (60) may need to operate for a longer period to achieve the desired moisture level, while in a dry environment, the blowing time may be reduced.

[0158] In some embodiments in which the air flow sensor is not comprised, then the functionality of blowing air over the reagent only until an adequate moisture level is left on the sample and / or sample holder (41) can still be carried out, with preconfigured and calibrated configurations, for example regarding the duration, distance and speed or power of the blowing.

[0159] In calculating the adequate moisture level, the processor may utilize in some embodiments machine learning algorithms that have been trained on various types of samples and reagents, allowing it to predict the optimal moisture level for each specific scenario. This dynamic adjustment ensures that the system can handle a wide range of staining procedures, from histological to cytological processes, while maintaining the integrity of the samples and minimizing reagent waste.

[0160] In a preferred embodiment of the automatic staining device (1) of the invention, the memory in communication with the processor comprises data representing a set of instructions that when executed by the processor cause the processor to: i'. receive one or more images taken by one of the at least one camera (20) comprising the image of one or more sample holders (41) and determine the presence, the position and / or the orientation of the sample holders (41) based on said images, wherein the determination of the position and / or orientation is made through geometric algorithms or through a first trained machine learning model, further wherein the set of instructions cause the processor to read one or more labels comprised in the image of the one or more sample holders (41) through the first or a second trained machine learning model, wherein said labels comprise at least data relating to a staining procedure reagents; ii'. receive one or more images taken by one of the at least one camera (20) comprising the image of one or more reagent vials (51), and determine the presence of the one or more reagent vials (51) and / or the open or closed state of the lid (52) of said reagent vials based on said images, wherein the determination of the presence of the one or more reagent vials and / or the open or closed state of the lid is made through the first, the second or a third trained machine learning model, preferably wherein the set of instructions cause the processor to read one or more labels comprised in the image of the one or more reagent vials (51) through the first, second, third or a fourth trained machine learning model, wherein said labels comprise at least data relating to a reagent, preferably to the reagent comprised in the corresponding vial (51).

[0161] It is noted that the processor may be configured to receive one or more images captured by the at least one camera (20). The camera images may include visual representations of one or more sample holders (41), and the processor is tasked with determining the presence, position, and / or orientation of these sample holders (41). The position and orientation may be determined using geometric algorithms that analyse key features in the image to calculate relative angles, distances, and alignments. For instance, geometric pattern recognition can compare the edges and corners of the sample holders (41) to predefined templates stored in the memory, ensuring the holders are correctly positioned for the staining procedure. Alternatively, this task may be performed by a trained machine learning model, which has been trained to recognize the orientation and placement of sample holders based on labelled training data. The machine learning model can generalize across different types and sizes of sample holders (41), offering flexibility in real-world applications.

[0162] In addition to detecting the presence and orientation of sample holders (41), the processor is preferably also configured to read labels present in the images. These labels may include human-readable information, such as the pathologist's name, laboratory ID, staining technique, date, or sample copy number. The human-readable portion may be processed through Optical Character Recognition (OCR), enabling the system to interpret text-based data directly from the label. In some cases, the labels may also include a DataMatrix code, which is a machine-readable identifier that stores compressed information, such as the sample ID or both the sample ID and staining technique. The processor may decode the DataMatrix using the same or a separate machine learning model trained to interpret such codes. This dual functionality allows the system to read labels from different sources, whether the label was generated by the staining device itself or by an external system. In scenarios where the label was created by an external system, and includes only the sample ID and technique, the system may automatically register the sample in the device's internal database and associate it with the appropriate staining protocol, reagents, and procedural data, ensuring seamless integration and automation.

[0163] In some embodiments, the processor is capable of receiving images taken by the camera (20) of the reagent vials (51). The processor may determine the presence of reagent vials (51) and assesses whether the lid (52) of each vial is in an open or closed state. This determination may again be made through geometric algorithms that recognize the shape, contours, and relative position of the vial and its lid, identifying whether the lid is open or closed based on its alignment or displacement. Alternatively, a trained machine learning model may be employed to detect more complex variations in the visual appearance of the vials, recognizing whether the lid (52) is fully opened, partially opened, or closed.

[0164] The system may further analyze labels present on the reagent vials (51). These labels, similar to the ones on the sample holders (41), may contain both human-readable data and encoded information in formats like DataMatrix. The human-readable portion may include details such as the reagent ID and an acronym indicating the reagent type. This information may be processed by the same OCR module that handles the labels on the sample holders (41). The encoded DataMatrix, on the other hand, contains more detailed information regarding the reagent vial (51), such as its ID, type, initial volume of reagent, expiration date, batch number, and other critical data for vial management. The system, through a trained machine learning model or a specialized decoding algorithm, extracts this information from the code and may automatically register or update the reagent vial (51) in the device's database, enabling the processor to monitor reagent usage, track reagent expiration, and ensure that the correct reagents are available and ready for the staining procedure.

[0165] In some embodiments, the determination of the presence, position, and orientation of the sample holders (41) and reagent vials (51), as well as the reading of labels, may be distributed among several machine learning models. For instance, a first machine learning model may be responsible for identifying the sample holders (41) and determining their orientation, while a second model focuses on label reading. A third model may handle the detection of the reagent vials (51) and the state of their lids (52), and a fourth model may focus on decoding complex label data, such as batch numbers, expiration dates, and volumes.

[0166] The system may also incorporate a feedback loop, wherein the processor uses the output of the machine learning models to make dynamic adjustments to the staining process. For example, based on the label information, the system may automatically retrieve the necessary reagents, adjust the volume of reagent to be dispensed based on the sample type, or modify the staining protocol depending on the reagent batch or expiration date. Furthermore, the system can flag issues such as missing reagent vials, incorrect sample placement, or misaligned sample holders, alerting the user or adjusting the procedure to correct the issue without manual intervention.

[0167] It is noted that various types of machine learning algorithms may be employed to achieve these objectives, and the models can be trained and fine-tuned to ensure reliable and accurate performance in diverse environments and with varying data inputs.

[0168] For the task of image analysis, including the determination of the presence, position, and orientation of the sample holders (41) and reagent vials (51), convolutional neural networks (CNNs) are preferably employed. CNNs are highly effective for image classification, object detection, and segmentation, all of which are necessary to distinguish between different components of the device. For example, a CNN can be trained to recognize the contours and edges of a sample holder (41) to determine its orientation and position on the support (42). Similarly, it can detect whether the lid (52) of a reagent vial (51) is open or closed by analysing the geometric and visual features of the vial’s shape and lid alignment.

[0169] The machine learning models may be trained using supervised learning techniques, where labelled datasets of images are provided. These images may depict various scenarios, such as sample holders (41) placed in different orientations, reagent vials (51) with lids (52) in both open and closed states, and labels with different text and barcode formats. The training data could include thousands of images annotated with the correct positions, orientations, lid states, and label contents. The CNN models can then learn from these labelled images by identifying patterns and features that are consistent with the correct annotations.

[0170] To train the models effectively, the data used fortraining and testing may need to be processed and augmented. For example, images can be pre-processed by normalizing the pixel values, adjusting brightness or contrast, and cropping or resizing them to a consistent format. Data augmentation techniques such as random rotations, translations, flips, and zooms can be applied to artificially increase the size and diversity of the training dataset. This helps the model generalize better and perform well under different lighting conditions, sample holder orientations, or camera angles, reducing the risk of overfitting to specific scenarios in the training data.

[0171] The labels read by the processor from the images may involve Optical Character Recognition (OCR) for human-readable text and DataMatrix decoding for machine-readable codes. OCR tasks can be handled by recurrent neural networks (RNNs), particularly long short-term memory (LSTM) networks, which are capable of sequential data processing. LSTMs are effective at recognizing and interpreting the sequence of characters in a label, even if the label is slightly distorted or partially obscured. The DataMatrix decoding may be handled by CNNs or other specialized decoding algorithms, which can identify and extract the compressed information from the barcode-like structure of the DataMatrix.

[0172] The machine learning models used for label reading (both OCR and DataMatrix decoding) may also be trained on labelled datasets. In the case of human-readable text, the dataset may contain various types of labels with different fonts, sizes, and layouts, annotated with the correct text sequences. For DataMatrix decoding, the training set may contain different DataMatrix codes, each paired with the correct decoded information. The system could be trained to handle different levels of image quality, ensuring that the models can successfully read labels even under suboptimal conditions, such as low resolution or poor lighting.

[0173] The machine learning models may need to be periodically retrained or fine-tuned as new data becomes available, especially if the staining device (1) is deployed in environments with different sample types, labelling systems, or reagent vials. Transfer learning techniques may be employed to fine-tune the models on new datasets without the need for large-scale retraining from scratch. This involves starting with a pre-trained model (on a general dataset of images or text) and fine-tuning it on a smaller, task-specific dataset. Transfer learning significantly reduces the amount of data and computational resources required to achieve accurate performance in new contexts.

[0174] The data used for training and testing may be obtained from various sources, such as preexisting databases of labelled biological sample images, internal databases of images captured during the staining process, or data shared between laboratories using similar systems. The datasets may be enlarged through collaboration with other research institutions or medical facilities, where anonymized data from real-world use cases can be incorporated. Data from different environments (e.g., different lighting conditions, equipment variations) ensures that the models are robust and adaptable to a wide range of practical situations.

[0175] For more complex image recognition tasks, the system may use deep learning models that combine CNNs with LSTMs, forming hybrid architectures. For instance, the CNN could be used to extract spatial features from the image, while the LSTM processes the sequential information (such as the layout of labels, barcodes, or vial arrangements). This combination is especially useful in tasks like reading long text labels or interpreting sequences of reagent vials positioned in a specific order.

[0176] The system may also integrate reinforcement learning algorithms to improve its decisionmaking process. For example, reinforcement learning could be used to optimize the placement of the dispensing probe (870) based on feedback from the staining results. The system could "learn" which adjustments lead to better staining outcomes by continuously analysing images of completed staining processes and correlating them with the procedural adjustments made during reagent dispensing.

[0177] The performance of the machine learning models may be continuously monitored using metrics like accuracy, precision, recall, and F1 score during testing. The dataset may be split into training, validation, and testing sets to ensure that the models are evaluated on unseen data, preventing overfitting. Cross-validation techniques can also be employed to ensure that the model’s performance is consistent across different subsets of the data. It is noted that the dispensing by the dispensing probe (80) comprised in the robotic arm (10), or in the automatic staining device (1), which is performed via a pressure-driven fluid displacement, depends on the determination of the presence, position and / or orientation of the sample holder (41) and the reading of the one or more labels of the sample holders (41), previously processed, and from the determination of the presence and open or closed state of the lid of one or more reagent vials (51) and optionally from the reading of the one or more labels from the reagent vials (51).

[0178] The precise control of this dispensing process is dependent on several critical determinations made by the device, ensuring that the reagent is accurately applied to the sample in the correct position, quantity, and at the appropriate time.

[0179] In some embodiments a relevant factor is the determination of the presence, position, and / or orientation of the sample holder (41). This determination is made based on the images captured by the at least one camera (20) and processed by the processor using either geometric algorithms or machine learning models. These models are capable of recognizing the position and / or orientation of the sample holder (41) on the support (42), ensuring that the dispensing probe (80) aligns correctly with the sample holder (41) to avoid misapplication of the reagent. If the sample holder (41) is misaligned or improperly placed, the system may adjust the position of the robotic arm (10) or, preferably, provide feedback to the user to correct the placement before dispensing begins.

[0180] Another relevant factor for some embodiments is the reading of one or more labels on the sample holders (41). These labels, which may include human-readable text or encoded information such as DataMatrix codes, contain vital data about the staining procedure. The data read from the labels may include the specific staining technique required, the type of sample, and any special handling instructions. This information can be used by the processor to select the correct reagent and determine the optimal amount to dispense. If the label includes information about the type of sample or staining technique, the device can adjust the pressure-driven fluid displacement system to match the reagent’s required volume and flow rate. The label reading process may involve OCR for text and specialized decoding algorithms for barcodes or DataMatrix codes, ensuring the system can interpret both human-readable and machine-readable data.

[0181] Furthermore, in some embodiments the dispensing process also relies on the determination of the presence and open or closed state of the lid (52) of one or more reagent vials (51). The camera (20) captures images of the reagent vials (51), and the processor analyzes these images to detect whether the vials are correctly placed and whether their lids (52) are open or closed. This alllows that the robotic arm (10) only attempts to withdraw reagent from vials that are available and properly prepared for dispensing. If the lid (52) is detected as closed, the system may either automatically open the lid (52) if the device is configured to do so, or it may issue a prompt for manual intervention. This step prevents errors such as attempting to withdraw reagent from a sealed vial, which could result in system malfunction or contamination.

[0182] Optionally, the system may also perform the reading of one or more labels on the reagent vials (51). These labels may contain information about the reagent, including the reagent type, volume, expiration date, and batch number. This data may be processed by the machine learning models to ensure that the correct reagent is selected for the staining procedure, and that it is within its valid usage period. The reagent vial labels may also include DataMatrix codes or other encoded information that can be decoded to automatically update the system’s inventory or validate the availability of the required reagent for the staining process.

[0183] The integration of these determinations, regarding the sample holder’s (41 ) position and labels, as well as the reagent vial’s (51) presence, lid state, and labels, facilitates that the pressure- driven fluid displacement system operates efficiently and accurately. The device preferably automatically coordinates these various elements, minimizing the risk of reagent waste, improper staining, or sample contamination. By ensuring that all components are in the correct state before dispensing begins, the device enhances the reliability and automation of the staining process, reducing the need for manual oversight.

[0184] Advantageously, the automatic staining device (1) described herein overcomes the limitations inherent in staining devices that rely on capillarity for reagent distribution. The use of a pressure-driven fluid displacement system, controlled by a processor and supported by advanced Al functionalities, enables precise and efficient reagent dispensing, even in situations where capillary action would be insufficient or problematic. This device eliminates the common issues related to reagent crystallization, uneven reagent distribution, and complex maintenance associated with capillary-based systems. In some embodiments, by automating the detection of sample holders (41), reagent vials (51), and their respective states, the device achieves full automation of the staining process, significantly reducing the need for manual intervention and increasing throughput and accuracy.

[0185] The integration of machine learning models in some embodiments allows for the real-time processing of images captured by the camera (20), ensuring the correct positioning and orientation of sample holders (41), reading of labels, and verification of reagent vials (51) and their lids (52). This results in a highly adaptive system capable of handling a wide range of samples, reagents, and staining protocols with minimal user input. The device automatically adjusts its operations based on the data it receives, ensuring optimal reagent usage and minimizing waste. In some embodiments of the device (1) the air blowing unit plays an useful role, since is configured to blow air over the reagent until an adequate moisture level remains on the sample holder (41), facilitating further reagent processing. This precise control of moisture is essential in preventing the drying out of the sample, which would hinder the distribution of subsequent reagents. The ability to maintain the sample at the perfect moisture level can be calculated in some embodiments based on real-time data from the air flow sensor, with the processor adjusting the air flow, duration, and angle of the blower to achieve optimal conditions. This controlled drying and dispersing of the reagent evenly across the surface of the sample holder

[0186] (41) allows for consistent, high-quality staining results.

[0187] Furthermore, the air blowing unit, which may present different orifices or slits, is preferably configured or shaped such that the air is evenly dispersed across the sample, addressing another common issue with non-capillary systems, uneven reagent spread. In some embodiments the system may adjust the air blower’s position and power, which coupled with the use of mechanical pumps to control reagent flow, offers unparalleled precision in reagent distribution, achieving uniform coverage across the sample while maintaining the appropriate moisture levels. This leads to improved staining quality, reduced reagent consumption, and a more reliable staining process overall.

[0188] According to a preferred embodiment of the automatic staining device (1) of the invention, the automatic staining device (1) further comprises a spreading unit (30) comprising a supporting element (32) and a spreading element (31), the spreading unit (30) preferably comprised in the robotic arm (10), wherein the longitudinal axis of said spreading element (31) is or can be placed parallel to the surface of the one or more sample holders (41) and / or the surface of the sample holder supports (42), and wherein the spreading element is supported by a supporting element (32) that can be displaced relative to the spreading element (31) in the direction perpendicular to the longitudinal axis of the spreading element (31) and parallel to the longitudinal axis of the supporting element (32), between a first configuration wherein the supporting element (32) supports the weight spreading element (31), and a second configuration wherein the spreading element (31) is not supported by the supporting element (32), such as when is supported by the sample holder (41) and / or the sample holder support

[0189] (42).

[0190] In some embodiments, the spreading element (31) or the entire spreading unit (30) may be configured to move vertically with respect to the robotic arm (10). This vertical movement is controlled within specific distance limits to prevent unintentional damage to the sample holder (41), particularly when dealing with fragile materials such as glass slides. The vertical tolerance may allow the spreading element (31) to make light contact with the surface of the sample or sample holder (41) without exerting excessive pressure that could cause breakage, as the only force exerted over the sample holder would be its weight would thanks to this vertical movement or tolerance of the spreading element (31) with respect to the robotic arm.

[0191] Additionally, the spreading unit (30) allows for smart, targeted dispensing and distribution of reagents over the portion of the sample holder (41) that contains the biological sample. By positioning the spreading element (31) precisely over the sample area, the device ensures that the reagent is applied efficiently, reducing waste and avoiding unnecessary coverage of areas outside the sample. This targeted approach is particularly beneficial for minimizing the use of expensive or hazardous reagents and for ensuring uniform application directly over the regions of interest on the sample holder (41).

[0192] The spreading element (31) may adopt various shapes, such as a cylinder, flange, or prism, and may rotate or remain stationary depending on the staining protocol. Additionally, the spreading element (31) may have hydrophobic properties to prevent sticking and facilitate even distribution of the reagent across the surface.

[0193] Advantageously, the integration of the spreading unit (30) into the robotic arm (10) allows for mechanical control over the distribution of the reagent across the sample surface, enabling precise application, especially in cases where viscosity or surface tension would otherwise hinder even spreading. The ability to move vertically with controlled tolerance prevents damage to fragile sample holders (41), while ensuring that the spreading element (31) makes sufficient contact to distribute the reagent effectively. Furthermore, the targeted dispensing capability of the spreading unit (30) allows for precise reagent application only to the sample areas, optimizing reagent usage and improving the overall quality and efficiency of the staining process.

[0194] According to a preferred embodiment of the automatic staining device (1), it further comprises one or more inclination sensors to detect the inclination of the one or more sample holders with respect to the vertical direction.

[0195] The term "vertical direction" may refer to what is commonly understood as the absolute vertical direction or be determined relative to the gravitational pull, additionally it may be understood as perpendicular to the surface of the sample holder. This inclination detection ensures that the sample holders (41) are properly aligned for the staining process, reducing the risk of uneven reagent distribution due to improper positioning.

[0196] The inclination sensors may include various types of devices, such as accelerometers, tilt sensors, inclinometers, gyroscopes, or orientation sensors. In some embodiments, an inertial measurement unit (IMU) is used, which combines both a three-axis accelerometer and a three- axis gyroscope. The IMU continuously measures both the linear acceleration and angular velocity of the sample holders (41), providing real-time data on their orientation. The inclusion of both accelerometers and gyroscopes ensures that the system can account for dynamic movements, making it suitable for environments where the device may be subject to vibration or slight shifts.

[0197] To mitigate the accumulation of errors typically associated with successive calculations, such as integration from gyroscope data, the system may implement a sensor fusion algorithm. This algorithm combines the angle data from the accelerometer and the angular velocity data from the gyroscope, and uses a statistical confidence model to calculate a more accurate and reliable inclination angle. The result is a robust real-time determination of the sample holder's (41) inclination, with the system updating the angle up to 20 times per second, ensuring precise alignment adjustments during the staining process.

[0198] In other embodiments, alternative sensors may include an angle sensor, which can directly measure the angular position of the sample holders (41) relative to a reference axis, or a gravitational sensor that specifically detects the direction of the gravitational pull to calculate inclination. These alternatives provide flexibility in the design and operation of the device, allowing different sensor configurations to be employed based on the desired level of precision and real-time feedback.

[0199] Advantageously, the inclusion of inclination sensors ensures that the automatic staining device (1) can accurately detect and correct the alignment of the sample holders (41) before and during the staining process. This feature is particularly beneficial in non-capillary systems, where uneven inclination could lead to improper reagent distribution or pooling on one side of the sample holder (41). By using a combination of accelerometer and gyroscope data processed through a sensor fusion algorithm, the system achieves a high level of precision, reducing errors and ensuring consistent staining results. Furthermore, the ability to detect and adjust inclination in real time adds to the overall automation of the device, minimizing the need for manual alignment and ensuring optimal staining conditions for each sample.

[0200] According to a preferred embodiment of the automatic staining device (1) of the invention, the sample holder supports (42) comprises one or more orifices, preferably a grid of at least 6 orifices to let the excess reagent flow through instead of letting the reagent fluid accumulate between the sample holder (41) and the sample holder support (42), preferably comprising at least 9 orifices, more preferably comprising a grid of at least 3x6 orifices, even more preferably comprising a grid of at least 3x12 orifices, allowing sufficient drainage for different types of reagents and sample sizes. In addition to providing a path for excess reagent flow, the grid of orifices may also serve a secondary purpose in aiding the calibration of the camera (20) and facilitating the identification of the correct position of the sample holder (41) on the support (42). The grid pattern can create a reference structure that the camera (20) can easily detect, allowing the processor to calculate the sample holder's (41) position and orientation with precision. The regularity of the grid offers distinct visual markers that help the system distinguish between different areas of the sample holder support (42), ensuring that the sample holder (41) is placed accurately for optimal staining results.

[0201] The design of the orifices and the grid itself can be adjusted depending on the specific staining protocols, reagents used, and the size of the sample holders (41). For example, the grid pattern may be expanded or reduced, and the size of the orifices may be adapted to account for the viscosity of the reagents. In some embodiments, the grid may be made of materials that enhance durability, chemical resistance, or cleaning efficiency.

[0202] Advantageously, the inclusion of a grid of orifices in the sample holder support (42) not only prevents the accumulation of excess reagent but also enhances the device’s overall automation and precision. By allowing reagent to flow through the support, the device (1) helps preventing the sample holder (41) to be inclined or be displaced, which is critical when dispensing. Also, it may ensure consistent reagent application and avoids potential interference caused by pooling. The grid also improves the system's ability to calibrate the camera (20) and detect the precise placement of the sample holder (41), contributing to the accuracy of the staining process. This design feature, combined with the flexibility to adjust the number and size of orifices, ensures that the automatic staining device (1) can accommodate a wide range of staining protocols and sample holder configurations, improving both the quality and efficiency of the staining procedure.

[0203] According to a preferred embodiment of the automatic staining device (1) of the invention, wherein the set of instructions, when executed by the processor, further cause the processor to: iii'. receive one or more images taken by one of the at least one camera (20) comprising the image of one or more sample holders (41), which can be the same or different as the images received in step (i'), provide said one or more images to the first, the second, the third or a fourth trained machine learning model, and determine the position and size of the sample in the one or more sample holders (41) based on the output received from the machine learning model; wherein the dispensing, and / or the mixing or withdrawal, by the automatic staining device (1) further depends on the determination of the position and size of the sample in the one or more sample holders from step (iii’).

[0204] The determination of the sample's position and size is crucial for optimizing the reagent usage, especially when working with smaller or irregularly shaped samples. By analyzing the images through the machine learning models, the processor can intelligently adjust the reagent volume and the specific areas where the reagents should be applied. For example, the system may apply more reagent to a larger sample area while avoiding unnecessary application over empty portions of the sample holder (41).

[0205] The trained machine learning models may be designed to recognize various sample types, including tissue sections, cell smears, or other biological specimens. These models may have been trained on a large dataset of labeled images, allowing them to generalize across different staining scenarios, sample holder formats, and sample sizes. Additionally, the models can detect the edges of the sample, determine the boundaries between different regions of interest, and provide output that guides the device's actions during the staining process.

[0206] Advantageously, this embodiment of the automatic staining device (1) significantly enhances the precision and efficiency of reagent dispensing. By usingn machine learning models to determine the exact position and size of the sample in the sample holders (41), the device (1) allows that the reagents are applied only where needed, minimizing reagent waste and reducing the risk of over-application. This intelligent system allows for targeted reagent application, even on small or irregularly shaped samples, ensuring consistent staining quality across all samples. Moreover, the real-time adjustment of the dispensing process based on the sample's size and position further enhances the device's flexibility and automation, enabling it to handle diverse staining protocols and sample types with minimal user intervention.

[0207] According to a preferred embodiment of the automatic staining device (1) of the invention, the automatic staining device (1) is configured to dispense an amount of reagent proportional to the size of the sample determined in step (iii’), and is further configured to dispense said reagent in a position of the sample holder (41) depending on the position of the sample determined in step (iii’). This proportional dispensing ensures that the volume of reagent used is precisely matched to the sample's dimensions, optimizing reagent usage and ensuring that only the necessary amount is applied to the sample.

[0208] The configuration allows the automatic staining device (1) to dynamically adjust the reagent volume according to the size of the sample. For smaller samples, the device may dispense a smaller volume of reagent, preventing excess reagent from accumulating on the sample holder (41) and potentially affecting the staining results. For larger samples, the reagent volume is increased accordingly, ensuring thorough and consistent coverage of the entire sample. Additionally, the device dispenses the reagent at the exact location where the sample is positioned, ensuring that the reagent is applied directly over the sample and not in areas of the sample holder (41) that do not contain biological material.

[0209] This precision in both the volume and location of reagent dispensing can be achieved through the integration of the machine learning models that determine the sample’s size and position, combined with the pressure-driven fluid displacement system that controls the reagent flow.

[0210] Advantageously, the ability to dispense a reagent volume proportional to the size of the sample, and in the exact position where the sample is located, greatly improves the efficiency and precision of the staining process in some embodiments. This feature minimizes reagent waste, particularly when using expensive or hazardous reagents, and ensures that the sample is treated evenly, regardless of its size or location on the sample holder (41). Furthermore, this proportional and targeted dispensing helps maintain consistent staining quality across different samples and protocols, contributing to the overall accuracy and automation of the staining device. By adapting the reagent application in real-time, the device achieves superior results in both small and large sample staining applications.

[0211] According to a preferred embodiment of the automatic staining device (1) of the invention, the set of instructions when executed by the processor further cause the processor to: iv'. receive one or more images taken by one of the at least one camera (20) comprising the image of one or more mixing vials, provide said one or more images to the first, the second, the third, the fourth or a fifth trained machine learning model, and determine the presence and position of the one or more mixing vials based on the output received from the machine learning model; wherein the mixing by the automatic staining device (1) depends on the determination in step (iv’) of the presence and position of the one or more mixing vials.

[0212] This, in some embodiments, ensures that the device verifies the correct placement and orientation of the mixing vials before initiating any reagent mixing processes, thereby preventing errors such as missing vials or incorrect vial positioning, which could affect the homogeneity of the reagent mixture or cause operational errors.

[0213] The machine learning models utilized for this task can be trained to detect various vial types, sizes, and positions within the device. These models can recognize mixing vials under different lighting conditions, with varying degrees of occlusion, or in different positions relative to the camera (20). By ensuring that the vials are correctly detected and positioned, the system can precisely control the mixing process, adjusting the timing, volume, and speed of reagent mixing as required by the staining protocol.

[0214] Advantageously, the integration of machine learning models to verify the presence and position of mixing vials enhances the device’s overall automation and reliability. By ensuring that the mixing vials are properly identified and positioned before reagent mixing begins, the device minimizes the risk of procedural errors, such as mixing reagents in an absent or misaligned vial. This level of precision is particularly important when mixing reagents that must be handled with care or in specific proportions, as it guarantees consistent and homogeneous results. Additionally, the use of trained machine learning models to detect vial presence in real-time enables the system to adapt dynamically to different vial configurations, further contributing to the flexibility and robustness of the automatic staining device (1).

[0215] According to a preferred embodiment of the automatic staining device (1) of the invention, longitudinal length of the spreading element is substantially the same as the width (45) of the sample holders (41) and / or sample holder supports (42), or has a difference in length with said width of ± 25%.

[0216] By matching or approximating the width (45) of the sample holder (41), the spreading element (31) can operate effectively across the entire surface without needing excessive movement or repositioning. This proportional sizing reduces the likelihood of missed areas or uneven reagent distribution, which could negatively impact the staining quality. The allowed tolerance of ± 25% ensures flexibility in adapting the spreading element (31) to various sample holder (41) sizes, allowing the device to accommodate different staining procedures and sample formats without requiring significant hardware modifications.

[0217] In some embodiments, the spreading element (31) may feature additional functionalities, such as rotation, adjustable pressure, or specialized surface properties (e.g., hydrophobicity), which work in conjunction with the proportional sizing to optimize the spreading and distribution of reagents.

[0218] Advantageously, the alignment of the spreading element’s (31) length with the width (45) of the sample holders (41) allows that the reagent is applied evenly across the entire sample area, reducing the risk of reagent pooling or insufficient coverage. The proportional length of the spreading element (31) allows for more efficient and quicker reagent application, reducing the time required for each staining procedure. Furthermore, the flexibility provided by the ± 25% tolerance accommodates a variety of sample holder formats and sizes, enhancing the versatility of the automatic staining device (1) and allowing it to function effectively in different laboratory settings and for a range of staining protocols.

[0219] According to a preferred embodiment of the automatic staining device (1) of the invention, the spreading element (31) is cylindrical and comprises a distal part (311), a proximal part (312), and a central part (313), wherein the diameter (D1) of the distal (311) and proximal part (312) is the same for both distal (311) and proximal (312) parts, and is bigger than the diameter (D2) of the central part (313), wherein the diameter (D1) of the distal (311) and proximal (312) parts is between 0.02 and 2 mm longer than the diameter (D2) of the central part (313), preferably is between 0.06 and 0.6 mm longer, more preferably is between 0.1 and 0.3 mm longer.

[0220] This design allows the spreading element (31) to make uniform contact with the sample holder (41) while minimizing excess pressure on the central part (313), which could otherwise lead to uneven reagent distribution or damage to the sample holder (41). The cylindrical shape, with a slight variation in diameter between the central part (313) and the distal and proximal parts (311 , 312), creates a stable spreading action that ensures even reagent application across the entire surface.

[0221] The specific dimensions of the diameter difference allow for precise control over the spreading process, particularly in delicate staining procedures where the reagent must be distributed uniformly and gently. This design helps prevent reagent pooling or incomplete coverage, both of which could negatively affect the staining results.

[0222] Advantageously, the cylindrical design of the spreading element (31), with its differentiated diameters between the distal / proximal parts (311 , 312) and the central part (313), provides precise and even spreading of reagents across the sample holder (41). The controlled diameter variation ensures optimal surface contact without exerting excessive pressure, preventing damage to the sample holder (41) and contributing to high-quality, consistent staining results. Additionally, the range of dimensional tolerance in the diameter difference (D1 and D2) accommodates various reagent viscosities and sample holder surface properties, enhancing the flexibility and reliability of the automatic staining device (1) across different staining protocols and sample types.

[0223] According to a preferred embodiment of the automatic staining device (1) of the invention, the air blower (60) has a shape and orientation adequate to evenly spread the reagents over the sample and / or sample holder (41), preferably such that the air is blowed at an angle between 10 to 80 degrees with respect to the normal to the surface of the sample holder, more preferably between 25 and 65 degrees, even more preferably between 35 and 55. This angled airflow allows for the reagent to be spread uniformly across the surface of the sample without causing excessive drying or pooling in specific areas.

[0224] The orientation and shape of the air blower (60) are relevant in controlling the dispersion of the reagent, particularly in cases where precise application is required. Blowing air at an angle allows the airflow to gently push the reagent over the sample’s surface, ensuring even coverage while preventing the reagent from accumulating in one area. The design of the air blower (60) may include specialized nozzles or slits that direct the airflow in a consistent and controlled manner, minimizing turbulence and promoting uniform spreading.

[0225] The the angle of the air blower (60) may be adjusted in some embodiments based on the type of reagent being used, the size and shape of the sample holder (41), and the specific staining protocol. In some embodiments, the air blower (60) may be adjustable, allowing the angle of the air to be dynamically modified during the staining process based on real-time feedback from the processor or sensors monitoring the reagent distribution.

[0226] Advantageously, the design and orientation of the air blower (60) enhance the effectiveness of reagent distribution, ensuring that the reagent is spread evenly across the sample holder (41) without over-drying or causing excess accumulation in localized areas. By ensuring uniform reagent application, the device improves the quality of staining, reduces reagent waste, and minimizes the need for manual adjustments, thus contributing to the full automation and precision of the automatic staining device (1).

[0227] According to a preferred embodiment of the automatic staining device (1) of the invention, the air blower (60) comprises one or more orifices (64) to blow the air, and: a. the longitudinal axis (62) of the air blower has a length (61) substantially the same as the width (45) of the sample holders (41) and / or sample holder supports (42), or has a difference in length with said width of ± 25%; and / or b. if the air blower comprises only one orifice, said orifice is elongated in the direction of the longitudinal axis (62) of the air blower (60), wherein the maximum length of said orifice is substantially the same as the width (45) of the sample holders (41) and / or sample holder supports (42), or has a difference in length with said width of ± 25%; and / or c. if the air blower comprises more than one orifice, said orifices are placed along the longitudinal axis (62) of the air blower (60), wherein the two more distanced orifices with respect to each other are separated such that at least part of one of the orifices is separated from at least part of the other orifice a distance substantially the same as the width (45) of the sample holders (41) and / or sample holder supports (42), or has a difference in length with said width of ± 25%.

[0228] This design ensures that the air blower (60) effectively covers the entire width of the sample holder (41), whether it uses a single elongated orifice or multiple orifices arranged along its longitudinal axis. The precise control over the width and positioning of the orifices allows for uniform air distribution across the surface, improving the overall quality and consistency of the reagent application.

[0229] Advantageously, the inclusion of orifices (64) aligned with the width (45) of the sample holders (41) ensures that the air blower (60) delivers uniform airflow across the entire sample, reducing the risk of uneven reagent drying or pooling. The flexibility of the air blower’s (60) design in different embodiments or comprised at the same time in one embodiment — whether with a single elongated orifice or multiple orifices — allows it to adapt to various sample holder sizes or blowing speeds or powers while maintaining optimal air distribution. This configuration significantly enhances the control over reagent spreading, further contributing to the overall automation and precision of the staining process in the automatic staining device (1). The proportional length of the air blower (60) or the spacing of multiple orifices also ensures that the device can handle a variety of sample holder formats, making it highly adaptable to different laboratory requirements.

[0230] According to a preferred embodiment of the automatic staining device (1) of the invention, one of the at least one cameras (20) is comprised in the robotic arm (10), and the central axis of the lens of said camera presents an inclination angle (21) with respect to the normal axis of the plane of the sample holders and / or sample holder supports of between 10 and 80 degrees, preferably between 25 and 65 degrees, more preferably between 30 and 60 degrees, even more preferably of around 45 degrees.

[0231] This inclined configuration of the camera (20) is preferably designed to minimize reflection when capturing images of the sample holders (41), which are often made of glass slides. Glass surfaces can produce significant glare or reflections when light hits them at certain angles, particularly if the camera lens is positioned perpendicular to the glass surface. By setting the camera (20) at an inclination angle (21), the system avoids direct reflections from the glass, ensuring clearer, more accurate images of the sample and any associated labels or reagents.

[0232] The inclined camera (20) thus enhances the image quality, making it easier for the system to accurately detect the position, orientation, and size of the sample, as well as any reagent distribution or other relevant features on the glass slide. This improved image quality is especially important for the machine learning models used in the device, which rely on clear and detailed visual data to perform their tasks effectively.

[0233] Advantageously, the angled camera setup significantly reduces glare and reflections from the glass surfaces of the sample holders (41), resulting in more reliable and higher-quality images. This ensures that the system can accurately interpret the sample and reagents, improving the precision of the automated staining process. The ability to avoid reflections also allows for more consistent imaging across various lighting conditions, making the device more adaptable and reliable. This configuration improves the overall efficiency and automation of the device, ensuring consistent and accurate results during the staining procedure while minimizing errors due to visual distortions caused by reflections.

[0234] According to a preferred embodiment of the automatic staining device (1) of the invention, one of the one or more inclination sensors is configured to detect the inclination of the automatic staining device (1) with respect to the vertical direction, preferably wherein said inclination is detected as a function of time or compared with previous measurements to determine the stability of the automatic staining device. The term "vertical direction" may refer to the absolute vertical axis or be determined relative to gravitational pull.

[0235] This stability detection is very important to ensure that the sample holders (41) remain securely in place and that the robotic arm (10) can accurately position and dispense reagents without disruption, and for the reagents to be evenly dispersed in the sample holders (41), whereas if there is an inclination this would be impossible. Additionally, any instability in the device could cause the sample holders (41) to shift, which would negatively impact the precise application of reagents and compromise the overall staining process.

[0236] In some embodiments, the inclination sensor, such as an accelerometer, may continuously or punctually measure the gravitational forces acting on the device. The data can be sent to the processor, where a convolutional neural network (CNN) trained on stability patterns preferably processes the information. This CNN model, embedded within the processor, has preferably been trained using data from both stable and unstable scenarios, allowing it to detect, for example, three conditions: stable, unstable, and an intermediate "gray zone" where there is a warning of potential instability, but not yet a need to halt operations.

[0237] By detecting the inclination over time and comparing the data with previous measurements, the system can determine if the automatic staining device (1) is stable, ensuring that the sample holders (41) remain unmoved and the robotic arm (10) performs its tasks without error. If the device detects instability, it can issue warnings or even halt the staining process to prevent reagent misplacement or damage to the samples. Advantageously, the ability to monitor and detect the stability of the automatic staining device (1) ensures that the device operates with precision and reliability. The real-time analysis of inclination, combined with the trained CNN, allows the system to proactively identify issues before they impact the staining process. By ensuring that the device remains stable, the automatic staining device (1) prevents errors such as reagent misplacement, sample holder shifts, or robotic arm inaccuracies, all of which could lead to compromised staining quality. This stability monitoring is a key feature in maintaining the overall efficiency and accuracy of the device, especially in environments where vibrations or external forces might otherwise cause instability.

[0238] According to a preferred embodiment of the automatic staining device (1) of the invention, the device (1 ) further comprises a washing receptacle (79) suitable to fit the length of the spreading element. he receptacle may be made from materials that are resistant to chemical wear and compatible with the reagents used in the staining process. Its design ensures that it securely holds the spreading element (31) during washing, minimizing the risk of damage to the element or inadequate cleaning.

[0239] Advantageously, the inclusion of a washing receptacle (79) sized to fit the spreading element (31) ensures that the device can maintain a high level of hygiene and cleanliness, which is important for preventing contamination between samples or reagents. This feature allows for efficient cleaning of the spreading element (31) without manual intervention, thus contributing to the overall automation and reliability of the device. By ensuring that the spreading element (31) is properly cleaned and ready for subsequent use, the washing receptacle (79) enhances the operational efficiency and quality of the staining process, providing consistent results across multiple staining procedures.

[0240] Figures

[0241] To enable a better understanding of the present disclosure, reference will now be made, by way of example only, to the accompanying schematic drawings. It is important to note that the following schematic drawings are provided solely as exemplary, non-limiting potential embodiments of the invention. Accordingly, the drawings are to be regarded as illustrative instead of restrictive, serving merely to aid in the explanation of the invention's principles and potential applications.

[0242] The figures provided are merely representational and may not be drawn to scale. Certain proportions thereof may be exaggerated, while others may be minimized. The figures are intended to illustrate various implementations of the invention that can be understood and appropriately carried out by those of ordinary skill in the art. Commonly designated elements among the various figures refer to common or equivalent elements in the depicted embodiments. The figures are not intended to be exhaustive or to limit the invention to the precise form disclosed. It should be understood that the invention can be practiced with modification and alteration, and that the invention be limited only by the claims and the equivalents thereof.

[0243] Figure 1 shows a possible embodiment of the automatic staining device (1) of the invention for carrying out treatments of histological staining, immunoenzymatic staining, and / or hybridisation on tissue sections or cell smears. In detail, the device (1) comprises a housing and a cover or lid that can be opened, exposing the staining or hybridisation area comprising the sample holder supports (42) and the sample holders (41), and also the rack (50) with the support (53) for the reagent vials (51), in a horizontal plane, under which a waste tray is located. This allows for an easy access and, if necessary, an easy maintenance. The area is segmented into compartments which are adapted to house sample holders (41) and which act as reaction chambers. A robotic arm (10) carrying an image capture camera (20), preferably high-precision camera, is shown. In a plane below than the staining or hybridisation area, a first reagent area / structure is shown that houses a plurality of common reagent reservoirs. Next to the staining or hybridisation area, a second reagent area / structure is shown which houses a plurality of specific reservoirs (51), as well as a mixing unit and a drying unit.

[0244] The device (1) of Figure 1 can comprise an LED lamp inside the cabin that serves to illuminate the entire work area at the required times (loading of doors, calibration of equipment, etc.), as well as LED lighting placed in the Z axes so that when capturing the image of a sample holder (41), the tissue can be detected without reflection problems. Both lamps can be turned off during the performance of the series to ensure that the technique is not affected by any light. The device (1) of Figure 1 can comprise a humidity and ambient temperature sensor of the cabin that is at the top, a sensor for opening the hood and the locks that prevent it from opening during a series for reasons of safety of the technician and to ensure the correct lighting of the interior during the processes of detection of fabric, reading of codes, etc., as well as a LED strip of the chassis that indicates to the user what state the instrument is in, if it is in process, finished, paused, request to pause or in error.

[0245] The first reagent area / structure is shown housing a plurality of common reagent reservoirs . The area is provided with a housing with a plurality of common reagent reservoirs. A common reagent reservoir may comprise at its top, a reservoir plug, and a reagent outlet orifice or bore. At its bottom, the reservoir has a stabilising projection of the common reagent reservoir. Also the means of identifying the reagents and their place of placement in the housing may be comprised in the first area / structure that houses the common reagents, and may further comprise level sensors for each reagent reservoir.

[0246] Figures 12 and 13 show in detail the staining or hybridisation area comprising a rack with a plurality of columns or elongated supports that house, such as 14, in a possible embodiment, and for example three sample holder supports (42) each column, which in turn can house one sample holder (41) each. The column or elongated support, is preferably horizontal, can be inserted or removed from the area independently of the other units. The compartment is shown with a light indicator (44), preferably an LED. The surface of the sample holder support (42) may comprise a plurality or a grid of orifices to let excess reagent to be disposed of, or, may be provided with grooves for the expulsion of excess residues on its upper part, which facilitate the cleaning of the reagents used. The sample holder support, in its lower part, is preferably provided with a heating system / thermal blanket and / or a vibration system to facilitate staining or hybridisation reactions. Preferably, a waste tray that collects waste from the sample holder supports (42) is placed below the staining or hybrydisation area, and can be composed of two parts, an upper part in the form of a grid and a lower part provided with an outlet orifice.

[0247] Figure 14 show in detail a possible embodiment of the support for several removable reagent vials rack (53), which, in its upper part, houses a plurality of specific reagent vials (51), while in its lower part it is provided with a thermostatted heater. Shown in figure 14 are seven of such racks (53), wherein each rack comprises up to 8 reagent vials (51) according to one or more exemplary embodiments shown in figure 14. Each channel of the thermostatted heater can have an optical sensor of the presence of the rack, in such a way that it allows knowing whether or not a rack has been introduced to read only the positions of the vials of the racks that have been detected. The support is segmented into a plurality of racks provided with a gripping portion, the rack housing the specific reagent reservoirs (51), in the embodiment shown, 8 reagent vials (51). The rack can be inserted or removed from the area independently of the rest of the racks by means of the gripping portion. A specific reagent vial (51) is shown in figures 18 and 9 in detail, in which the cap or lid (52) is opened, and it can be seen an outlet port (54) to allow the probe to aspirate specific reagents. Figure 9 further shows an example of image processing for automatic label recognition, wherein the label is spatially transformed to be more readable.

[0248] Figure 19 shows in detail a possible embodiment of the mixing and drying unit (70) housing a washing station (75) of the one or more dispensing probes, preferably of the specific reagent dispensing probe, an automatic reagent mixing unit (77), tubes for the automatic mixing of reagents (76), a drying station (78) of the spreading element (31), and a washing station or receptacle (79) of the spreading element (32). In the rack of specific vials or reservoirs (51), a plurality (n) of vials or reservoirs (51) can be placed, which can be aspirated and dispensed directly into the slides (41) or can be dispensed into the mixing tubes (76) to make mixtures of different specific reagents, since these mixing reagents have several hours of stability and if they were not mixed in situ, they could not be used in the treatments. Additionally, having the possibility of mixing reagents according to the protocols implemented by computer, gives versatility to the apparatus-method-system, since it will not need to have as many reagent racks as needed for the different treatments, allowing from a limited number of reagents, by combination between them, to expand in number and the type of treatments to be applied to the samples.

[0249] Illustratively, different types of mixing reagents are described below, in particular, reagents that when mixed have a stability of between 30 minutes to 24 hours. A planner comprised in the automatic staining device (1), who preferably controls the device (1) of the invention, depending on when he needs the mixing reagent and its stability, will look for the best time to perform the mixing automatically. Preferably, it will carry out the process when the robot is stopped (waiting for incubations, for example) so as not to extend the total time of the series. The mixtures may be of a plurality (n= 2, 3, 4, etc.) of reagents. It should be noted that there is a minimum mixing volume that is limited by the mixing ratio and the minimum dispensing volume of the system. For example, there is a mixture of 2 reagents wherein 25 pl of one is dispensed for each 1000 pl of the other. As the system cannot dispense less than 25 pl, the minimum mixture would be 1025 pl. On the other hand, in a protocol for a slide, 300 pl are used. Once the two reagents are dispensed into the mixing tube, a homogenisation of the mixture is performed. The way to do this is by aspirating and dispensing the volume of the mixed reagent in one of the tubes (26) (in the form of agitation). In a particular embodiment, the mixing tubes (26) are not cooled.

[0250] A possible embodiment of the robotic arm (10) has means for a displacement in the XYZ axes and is provided with a high precision image capture camera (20), disposed at an angle of 45 degrees (see figure 2). The camera is disposed associated with the displacement means in the Z axis, adjacent to the dispensing probe (80), which is shown in detail in figure 17, wherein it is further shown a control element for the probe, and the spreading unit (30), so that the dispensing of reagents, their spreading and the optical evaluation of the sample can be carried out with the same device. The robotic device can be provided with common reagent dispensing comb and air blow drying system (36). The configuration of the invention, integrating a plurality of components in the same element, allows a reduction in the time and energy consumption of the protocol in use.

[0251] According to some embodiments, a common reagent dispensing comb is integrated in the displacement means for the Z axis of the robotic device (10). Preferably, the comb comprises 6 dispensing probes.

[0252] Figure 15 shows in detail the configuration of the spreading unit (30), adjacent to the air blow unit (60), integrated in the displacement means (Z) of the robotic device (10), which allow a reduction in the consumption of time and energy of the protocol in use. Figure 3 shows the spreading element (31) provided with two side portions, referred to as distal and proximal part indistinctly, (312 and 311), and a central part (313), wherein the side portions have the same diameter (D1), which is longer than the diameter (D2) of the central portion. This allows the side portions to rest on the sample holders (41), and when the spreading element (31) combs, or rolls or passes along the length of the sample holder (41) to spread the reagent, there is left over the sample and the sample holder (41) a thin and even layer of reagent, with a thickness approximate to half of the difference of D1 and D2 in length. Preferably the longitudinal length (L2) of the side portions (311 and 312) is much smaller, preferably below a fourth of the longitudinal length (L1) of the spreading element (31). Figure 16 shows in detail the air blowing unit (60), provided with a slit (64) which preferably has a length equal or similar to the width of the sample holders (41) or +- 25% said with. In some embodiments it may have orifices placed along an axis (62) which is preferably parallel to the surface of the sample holders (41).

[0253] It is noted that a filter for gas removal can be integrated into the apparatus of the invention.

[0254] A possible configuration of components integrated into the displacement means (Z) may comprise, among others, a probe, chamber, comb, blowing system, and electronic component. Figure 33 shows a possible configuration of components integrated in the displacement means (Z) comprising, among others, a probe, roller, comb provided with six dispensing probes, and a blowing system. A possible configuration of components integrated into the displacement means (Z) may comprise, among others, a probe, roller, blowing system, and a temperature monitoring system comprising an infrared (IR) sensor, electronic temperature control board, and connection pins of the electronic board for the IR sensor. These configurations allow a reduction (improvement) in the time and energy consumption of the protocol in use.

[0255] The features described in relation to the apparatus (1) can be integrated and are interchangeable with the components of the system of the invention.

[0256] Figures 5 and 6 show a part of the robotic arm (10) comprising the spreading unit (30) and the air blowing unit (60), which are placed over the proximal end of the sample holder support (42) which supports a sample holder (41) or slide (41), according to one or more embodiments of the invention. It can be appreciated in figure 5 how the spreading unit (30) rests on the supporting piece (33) of the robotic arm (10), therefore the robotic arm (10) holds all the spreading unit (30) weight. However, in figure 6 it can be appreciated how, once the robotic arm has approached vertically the sample holder (41) to situate the spreading unit (30) over said sample holder (41), prior to beginning the spreading of the reagents, the spreading unit

[0257] (30) now has been displaced with respect to the supporting piece (33) of the robotic arm, and the spreading unit (30) rests now over the sample holder (41) due to the spreading element

[0258] (31), or, in particular, the side parts (311 and 312) of the spreading element (31), resting on the sample holder (41), therefore the force applied to the sample holder (41) is only the weight of the spreading unit (30), which is enough to guarantee a correct spreading, but not excessive as to compromise the integrity of the sample holder (41), which typically is made of a thin layer of glass. IT is observed in figure 6, how the spreader support element (32) can slide or traverse the supporting piece (33) of the robotic arm until certain point allowing to move the spreading unit (30) the length of the supporting piece (33) or less, in the vertical axis with respect to the robotic arm, and has a stopper on top to prevent from sliding out.

[0259] Figures 8 to 10 show examples of image processing for reading sample holder (41) labels, reagent vials (51) labels in the case of figures 8 and 9 respectively, and for using the sample holder support (42), and / or the orifice grid, for calibrating the camera in the case of figure 10. Figure 11 show an example of internal processing to determine if the position or orientation of the calculated edges (top, bottom and lateral ones, for example) of a slide or sample holder (41) is in poor, good or excellent agreement with the previously calibrated position or orientation of the calculated edges of the sample holder support (42).

[0260] Figure 7 shows an example of the workflow that can be followed for a smart dispensing, wherein once the camera is calibrated, an image or more of the reagent vials (51) and of the sample holder or slide (41) is taken, then said one or more images are processed to determine the status of the sample holder (41), such us for example the positioning or orientation, or the positioning and size of the sample comprised on the sample holder (41) and of the status of the reagent vials (51), such as for example the presence of vials (51) and open or closed status, as well as reading labels in one or both of the sample holder (41) and the reagent vial (51) to obtain information of the type of reagent or staining procedure that has to be followed, and this processed information is taken in consideration to provide a smart dispensation, preferably a targeted dispensing with respect to the sample, is provided or instead it is paused, and for example a message or alert is prompted to the user, until the status of the sample holder (41) or the reagent vials (51) is the expected or correct one. Methods and examples

[0261] Camera Calibration

[0262] The automatic staining device (1) may employ computer vision techniques to automate the detection, analysis, and interaction with objects such as sample holders (41) and / or reagent vials (51). In some embodiments, the device performs camera calibration using an algebraic method without the use of artificial intelligence. This calibration process preferably involve capturing images wherein the camera's contrast is maximized and the gain is minimized, enhancing the perception of calibration patterns such as beacons with reduced noise. The characteristic points of the beacon or sample holder support (42), such as grids, circles, or squares, are detected within the image. Once these characteristic points are identified, the camera's intrinsic parameters, including but not limited to distortion coefficients and focal length, may be calibrated. The relative position of the camera with respect to the reference frame of the robotic arm is then computed, utilizing intrinsic and extrinsic calibration methods to determine the camera's internal properties and its spatial relationship within the robotic environment. Techniques such as the SolvePnP algorithm and calibration functions are preferably applied to compute these parameters, allowing accurate mapping between image coordinates and the robotic system's coordinate system.

[0263] Detection of Labels and Patterns

[0264] In some embodiments, the device incorporates methods for detecting labels on, for example, sample holders (41) and / or reagent bottles (51), and / or patterns in sample holder supports (42), which are critical for identifying and tracking objects during the staining process. Edge detection techniques, such as the Canny edge detection algorithm, may be utilized to identify the boundaries of labels and beacons within the images. Following edge detection, line detection methods like the Hough Line Transform can be applied to detect straight lines representing, for example, the edges of labels. The detected lines may then be classified into categories, including but not limited to top, bottom, left, and right edges, to identify the four corners of a label or beacon, wherein the sample holder support may be regarded as a beacon, or the pattern of the orifices in the sample holder support may be utilized instead of a label or beacon for recognition. Clustering techniques such as K-means clustering can be employed for this classification, which may involve machine learning to organize the lines geometrically.

[0265] Masking and thresholding techniques are preferably employed to isolate regions of interest, enhancing detection accuracy by highlighting relevant areas and suppressing background noise. An iterative detection process may be implemented, wherein the system continuously adjusts thresholds and detection parameters until the correct bounding box or polygon covering the label is identified. If the label meets certain size criteria and four defining lines around the label can be extracted, the system may conclude that a label is present; otherwise, the process may halt and an error may be logged.

[0266] Calibration and Alignment

[0267] To ensure accurate positioning and manipulation of samples, the device may execute precise alignment between the camera and the robotic system. Coordinate assignment is performed by associating detected image points, such as the corners of sample holders, with real-world coordinates. This allows for accurate mapping between the image plane and the physical workspace of the robotic arm. Error calculation methods can be employed to compute error vectors representing any misalignments between the detected positions and the expected positions. Recalibration may be conducted by projecting two-dimensional image points back into three-dimensional space to correct alignment errors, ensuring that the system maintains high accuracy in sample detection and manipulation over time.

[0268] Affine transformations can be utilized to manipulate images through operations such as scaling, rotation, and translation. These transformations preserve parallelism and ratios of distances, allowing the device to adjust images for better alignment with detected objects or to normalize images for further processing. Affine transformations facilitate consistent analysis despite variations in image orientation or scale, which is essential for accurate detection and positioning of sample holders and labels.

[0269] Additionally, homographic, or projective, transformations can be applied to correct perspective distortions in images, particularly when objects are viewed at an angle. By mapping points from one plane to another, the device can rectify images to present objects, such as labels or beacons, in a frontal view. This correction enhances the accuracy of subsequent image processing tasks by providing a consistent viewpoint, which is crucial for tasks like barcode recognition and precise robotic manipulation.

[0270] Label or Barcode Reading and Validation

[0271] Label or barcode reading and validation may be performed to identify samples and associate them with corresponding data. In some preferred embodiments, the device employs deep learning models for the recognition of barcodes and labels on sample holders (41) and / or reagent bottles (51). Optical Character Recognition (OCR) techniques are preferably utilized to extract textual information from images, even when barcodes or text are partially obscured or of low quality. Machine learning models, such as Long Short-Term Memory (LSTM) networks or convolutional neural networks, may be implemented to enhance accuracy in text recognition, and / or to allow recognition of hand-written text. These models are preferably trained using datasets comprising various barcode and label images, allowing the system to generalize across different formats and conditions. Perspective correction using methods like WarpPerspective() may be applied to adjust the label's perspective, and image sharpening techniques such as applying a convolution filter may enhance the clarity of the barcode and text. The extracted code may be validated against a list of expected sample identifiers for the current batch, and if a match is found, the device proceeds; otherwise, an error may be logged.

[0272] Detection of Reagent Vials and Status Recognition

[0273] In some embodiments Deep learning models are employed for the detection of reagent vials (51) and the determination of their status, such as whether a vial is present and whether it is open or closed. Convolutional neural networks are preferably utilized to analyse images captured by the device's cameras, identifying the presence and state of vials with high accuracy. The models may be trained using image augmentation techniques to increase the available dataset, including but not limited to Gaussian smoothing, sharpening, and random flips, to enhance robustness against variations in imaging conditions.

[0274] In some embodiments, the network architecture may be based on MobileNetV2 for feature extraction, followed by fully connected layers for classification. Activation functions such as the softmax function can be used in the final layer to output class probabilities. The loss function utilized is preferably cross-entropy loss, optimized using algorithms like Stochastic Gradient Descent (SGD) with progressively adjusted learning rates over multiple epochs. Preprocessing techniques may be applied to the images, including dynamic and static data augmentation. Dynamic augmentation involves applying random transformations to image attributes such as brightness, contrast, and saturation during training, enhancing the model's ability to generalize under varying conditions. Static augmentation may include Gaussian smoothing, image sharpening, and cropping.

[0275] A batch size may be selected based on computational resources and dataset size, and validation strategies, such as k-fold cross-validation, are preferably employed to ensure model reliability and generalization. This implementation allows the device to accurately detect vials and their statuses, facilitating automated reagent handling.

[0276] Detection of Tissue Stains via Image Segmentation Techniques

[0277] In some embodiments, the automatic staining device (1) processes images of biological samples to detect tissue stains and prepare for targeted reagent dispensing. Segmentation methods can be applied to separate stained tissue regions from the background. Topological methods, such as Topological Blob Merging and Refinement (TBMR), are preferably utilized to identify connected regions representing tissue.

[0278] In some embodiments geometric filtering is performed to eliminate regions that do not match expected size or shape characteristics, such as regions with high eccentricity or irregular shapes. Clustering techniques, such as K-means clustering, can be employed to classify pixels based on intensity or colour, further refining the segmentation and enhancing the accuracy of tissue detection.

[0279] Once tissue regions are detected, the device preferably approximates these regions into polygons for analysis and processing. Algorithms like the Approximate Polygonal Curve (ApproxPolyDP) method can be used to represent the contours of tissue regions with simplified polygonal shapes. This approximation reduces computational complexity and facilitates the calculation of dispensing strategies by providing a manageable representation of the tissue areas.

[0280] In cases where detected tissue areas are negligible or below a predefined threshold, the device may assume that the entire sample holder (41) is covered with tissue to prevent missing any areas during staining. Perspective adjustments may be applied to align the polygons accurately with the sample holder's coordinate system.

[0281] Image Fusion

[0282] According to some embodiments, the device captures multiple images of the sample holders (41), which are fused to enhance the visibility of relevant regions, such as tissue samples. Image fusion may involve combining regions from images captured under different conditions, such as varying contrast or exposure settings. The fusion process selects key areas from each image and merges them to create a composite image that retains the most informative content from both. Three-dimensional calibration information from the calibration module can be integrated into the image fusion process to ensure proper alignment and accuracy. This integration corrects any distortions or misalignments due to camera angle or positioning, ensuring that the fused image accurately represents the tissue regions on the sample holder. Calibration data also aids in mapping the fused image to real-world coordinates, which is essential for precise reagent dispensing.

[0283] Smart Reagent Dispensing

[0284] The automatic staining device (1) of the invention preferably optimizes reagent dispensing by calculating the optimal positions and volumes for reagent droplets over the detected tissue regions. In some embodiments, the system calculates maximal dispensing zones based on the size and distribution of tissue regions. Maximal dispensing zones are determined by analysing the spatial distribution of tissue regions and calculating positions that maximize coverage with minimal reagent usage. Geometric algorithms may be utilized to define these zones, considering factors such as the size, shape, and proximity of tissue areas.

[0285] Strategies may be implemented to merge adjacent dispensing zones when possible, reducing the number of dispensing actions required. If neighbouring droplets can be merged without exceeding maximum allowed volumes, the system fuses these droplets into a single larger drop. The device preferably evaluates whether merging zones results in efficient coverage without exceeding maximum allowable reagent volumes per dispensing action.

[0286] Drop locations may be adjusted to centre over tissue regions, and the volume of each drop may be optimized to ensure adequate coverage without excess reagent usage. Volume minimization techniques adjust the volume of reagent dispensed based on the area of tissue to be covered, optimizing reagent usage and minimizing waste.

[0287] For example, the system may first calculate individual droplets to cover the tissue region and then merge adjacent droplets if fusion results in a reduced total volume. The resulting droplets can then be centred over the largest tissue areas, ensuring full coverage while optimizing reagent usage. This intelligent dispensing mechanism may achieve significant reductions in reagent usage, possibly around 46%, by optimizing dispensing zones and minimizing wastage. The device may thus adjust dispensing parameters dynamically based on factors such as tissue size, shape, and location.

[0288] In a second aspect of the invention, an automatic staining device (1) for dispensing staining reagents or fluids over one or more biological samples is disclosed, wherein one or more reagents are delivered through a controlled pressure-driven flow. The automatic staining device (1) comprises a pressure-driven fluid displacement mechanism, such as a fluid pump; a memory and a processor in communication with said memory, wherein the memory preferably comprises at least one of: information of at least one protocol, said protocol comprising information related to at least one reagent dispensing step and at least one blowing step; and information related to the one or more biological samples. The device further comprises one or more sample holder supports (42) suitable to support one or more sample holders (41); one or more reagent vials (51); an air blowing unit comprising an air blower (60) and, preferably, an air flow sensor; and a robotic arm (10) comprising at least one dispensing probe (80), wherein the dispensing probe (80) is fluidly connected to the pressure-driven fluid displacement mechanism, preferably wherein the air blowing unit is also comprised in the robotic arm (10). The memory comprises instructions that, when executed by the processor, cause the processor, during a blowing step, to regulate the flow or speed of air in the air blowing unit or the time duration of air blowing, the robotic arm displacement or speed during the air blowing step and / or to regulate the distance of the air blower (60) to the sample or sample holder (41) during the blowing step, preferably based on information stored in the memory regarding a dispensing step of the protocol, preferably a previous, current or following reagent dispensing step and / or information regarding the biological sample.

[0289] It is further noted that the memory may comprise any suitable data storage medium, such as, but not limited to, solid-state drives, hard disk drives, flash memory, or cloud-based storage, and may be volatile or non-volatile. The processor may comprise any type of microprocessor, microcontroller, field-programmable gate array (FPGA), or application-specific integrated circuit (ASIC), and may be configured to execute instructions stored in the memory. In some embodiments, the processor may also function as a controlling unit, or may be in communication with a separate controlling unit, such as a programmable logic controller (PLC), industrial PC, or distributed control system, to coordinate the operation of the device components, including the pressure-driven fluid displacement mechanism, air blowing unit, robotic arm, and sensors.

[0290] In some embodiments, the memory may store information related to one or more protocols, which may comprise a sequence of reagent dispensing steps and, optionally, one or more blowing steps. It is noted that a protocol in some embodiments may be defined as a set of instructions or parameters that specify the order, timing, volume, and type of reagents to be dispensed, as well as the conditions and parameters for any associated air blowing or spreading steps. The information related to reagent dispensing steps may include, for example, the identity of the reagent, the volume or flow rate to be dispensed, the location and duration of application, and the sequence relative to other steps. The information related to blowing steps may include, for example, the timing, duration, air flow rate, temperature, humidity, and spatial parameters such as the distance and orientation of the air blower relative to the sample or sample holder or the speed of movement of the air blowing unit and / or the speed of movement of the robotic arm. Additionally, information related to blowing steps may include different regions of the sample holder over which the blowing will be carried out with different parameters or blowing profiles, said parameters being for example, but not limited to, the timing, duration, air flow rate, temperature, humidity, and spatial parameters such as the distance and orientation of the air blower relative to the sample or sample holder or the speed of movement of the air blowing unit and / or the speed of movement of the robotic arm.

[0291] It is noted that the protocol does not require a blowing step after each reagent dispensing step, and the inclusion of blowing steps is optional and protocol-dependent. In some embodiments, certain protocols may not require any blowing steps at all, while others may include one or more blowing steps at specific points in the protocol, such as after the application of particular reagents, before the application of a subsequent reagent, or at the end of the staining process. The device is configured to create or accept and execute protocols with or without blowing steps, providing flexibility and adaptability to different staining requirements. In some embodiments, the protocols may be input manually by a user, for example, via a graphical user interface, touchscreen, or external computer, or may be selected from a library of pre-defined protocols by name or identifier. Examples of protocols may include, but are not limited to, hematoxylin and eosin (H&E) staining, immunohistochemistry (IHC), periodic acid-Schiff (PAS) staining, or in situ hybridization, each comprising a specific sequence of reagent dispensing and optional blowing or spreading steps. In some embodiments, the device may be configured to automatically select or modify protocols based on the type of biological sample, the reagents available, or user preferences, for example, by means of artificial intelligence or rule-based algorithms.

[0292] It is further noted that the information stored in the memory may also comprise information related to the one or more biological samples, such as, but not limited to, sample type, tissue origin, thickness, fixation status, or previous processing history. This information may be used by the processor to adapt the protocol, for example, by adjusting reagent volumes, flow rates, or blowing parameters to optimise staining quality for different sample types. In some embodiments, the device may comprise a camera, such as, but not limited to, a digital CMOS or CCD camera, which may be used to identify samples, for example, by reading barcodes, QR codes, or to image analysis by using artificial intelligence algorithms to recognise sample features, sample holders, or sample holder supports. The camera may also be used for quality control, monitoring reagent application, or verifying the position and orientation of the samples and / or sample holders.

[0293] It is noted that the air blowing unit may comprise one or more air blowers (60), which may be of any suitable type, such as, but not limited to, centrifugal fans, axial fans, diaphragm pumps, piezoelectric blowers, or compressed air nozzles. The air blower (60) in some embodiments is configured to deliver air at controlled flow rates and / or pressures, and / or in some embodiments, temperatures, and may comprise one or more orifices or nozzles, for example dimensioned to match the width or geometry of the sample holder (41) or sample holder support (42), thereby ensuring even and uniform reagent spreading, removing or drying. The air blowing unit may further comprise an air flow sensor, such as, but not limited to, a thermal mass flow sensor, differential pressure sensor, or hot-wire anemometer, which may be used to monitor and regulate the flow of air in real time. The air flow may be measured in units such as litres per minute (L / min), cubic centimetres per second (cm3 / s), or standard cubic feet per minute (SCFM), and in some embodiments may be adjusted dynamically by the processor or controlling unit based on protocol requirements or sensor feedback.

[0294] In some embodiments, the air blowing unit may be integrated into the robotic arm (10), allowing for precise and programmable positioning of the air blower (60) relative to the sample or sample holder (41). The robotic arm (10) may comprise any suitable actuation mechanism, such as, but not limited to, electric motors, pneumatic actuators, or hydraulic actuators, and may be configured for multi-axis movement, for example, in Cartesian, polar, or articulated configurations. The robotic arm (10) may further comprise at least one dispensing probe (80), which may be fluidly connected to the pressure-driven fluid displacement mechanism by means of flexible tubing, rigid conduits, or integrated microfluidic channels. The dispensing probe (80) may be of any suitable material, such as stainless steel, polymer, or ceramic, and may comprise single or multiple channels for dispensing different reagents. In some embodiments, the air blowing unit may further comprise an anti-splatter nozzle or protection, to prevent reagent to reach the opening of the air blower (60) and crystallize or solidify therein, thus obstructing it, for example this anti-splatter nozzle or protection may comprise a flap that blocks the path of fluid droplets from the sample holder or wash probes to the position of the air blower (60).

[0295] It is noted that during a blowing step, in some embodiments the memory may comprise instructions that, when executed by the processor, cause the processor to regulate the flow of air in the air blowing unit and / or to regulate the distance of the air blower (60) to the sample or sample holder (41). The regulation of air flow may be achieved by adjusting the speed of the air blower, opening or closing valves, or modulating the duty cycle of piezoelectric elements, among other options. The distance of the air blower (60) to the sample or sample holder (41) may be regulated by moving the robotic arm (10), adjusting telescopic or sliding mounts, or by means of automated height adjustment mechanisms. The distance may be measured in millimetres or centimetres, for example, and may be adjusted between, for example, 1 mm and 100 mm, among other options, depending on the desired spreading or drying effect.

[0296] It is further noted that in some embodiments the regulation of air flow and / or distance during a blowing step may be based on information stored in the memory regarding a dispensing step of the protocol, preferably a previous, current, or following reagent dispensing step. In this context, a previous reagent dispensing step may refer to the most recent reagent application prior to the blowing step, a current reagent dispensing step may refer to a reagent application occurring simultaneously or in close temporal proximity to the blowing step, and a following reagent dispensing step may refer to the next scheduled reagent application in the protocol. The parameters of the blowing step, such as air flow rate, duration, or distance, may be in some embodiments adapted based on the characteristics of the reagent applied in the previous step (for example, its viscosity, volatility, or required drying time), the requirements of the current step (for example, simultaneous spreading of a co-applied reagent), or the needs of the following step (for example, ensuring optimal moisture level for the next reagent). In some embodiments, the regulation may also take into account information regarding the biological sample, such as its type, thickness, or sensitivity to drying, to prevent over-drying or sample damage.

[0297] In some embodiments, the device may further comprise additional sensors, such as inclination sensors, accelerometers, or gyroscopes, to monitor the horizontal positioning and stability of the sample holders (41), sample holder supports (42), or the device itself. These sensors may provide feedback to the processor or controlling unit, allowing, for example, providing the user a warning if the sample holders (41) are not horizontal and / or to stop the process.

[0298] Advantageously, the automatic staining device (1) provides a highly flexible and adaptive platform for the automated staining of biological samples, enabling precise and reproducible reagent application and spreading through pressure-driven dispensing and programmable air blowing. The integration of memory-stored protocols, sensor-based process control, and robotic actuation allows for the execution of complex and customisable staining workflows, minimising reagent waste, reducing maintenance requirements, and ensuring optimal staining quality across a wide range of sample types and protocols. The capability to regulate air flow and distance based, for example, on protocol and sample information, allows to achieve the optimal degree of sample or reagent moisture (or dryness), which enables dispensing further reagents that need certain level of moisture to properly spread over the sample, and / or to achieve a level of dryness or moisture adequate for the sample or protocol, addressing the limitations of non-capillary systems. Additionally, in some embodiments the inclusion of sample identification features, such as a camera and Al-based recognition, further enhances automation and traceability, making the device ideally suited for modern histology and pathology laboratories seeking reliability, efficiency, and standardisation.

[0299] According to a preferred embodiment of the second aspect of the invention, the flow of air in the air blowing unit and / or the distance of the air blower (60) to the sample is regulated based on a dispensing step of the protocol, preferably the previous, current or next reagent dispensing step of the protocol.

[0300] It is noted that in some embodiments, the regulation of the flow of air in the air blowing unit and / or the distance of the air blower (60) to the sample may be dynamically determined by the processor or controlling unit in response to information associated with a dispensing step of the protocol, preferably the previous, current, or next reagent dispensing step as defined in the protocol. The previous reagent dispensing step may refer to the most recently completed reagent application, the current step may refer to a reagent application occurring simultaneously or in close temporal proximity to the blowing step, and the next step may refer to the subsequent scheduled reagent application. The regulation may comprise, for example, adjusting the air flow rate, speed, pressure, temperature, or duration, preferably the air speed, as well as modifying the spatial positioning of the air blower (60) relative to the sample or sample holder (41), such as by varying the vertical or horizontal distance, angle, or orientation. These adjustments may be based on parameters such as the type of reagent dispensed, its physical or chemical properties (for example, viscosity, volatility, or drying time), the required spreading or drying effect, or the compatibility with subsequent reagents. The air flow may be measured in units such as litres per minute (L / min), cubic centimetres per second (cm3 / s), or standard cubic feet per minute (SCFM), and the distance may be measured in millimetres or centimetres, for example, between 1 mm and 100 mm, among other options.

[0301] In some embodiments, the regulation may be implemented by means of pre-defined rules or algorithms stored in the memory, or may be determined in real time based on sensor feedback, such as from air flow sensors, pressure sensors, or cameras. The processor may access the protocol to retrieve information about the sequence and characteristics of reagent dispensing steps, and may use this information to anticipate or respond to the requirements of each step. For example, after dispensing a reagent with high viscosity, the air flow or air speed may be increased or the distance decreased to ensure uniform spreading, whereas for a volatile reagent, a lower air flow or greater distance may be selected to prevent excessive evaporation. The regulation may also take into account the timing and coordination of dispensing and blowing steps, allowing for simultaneous or sequential operation as required by the protocol.

[0302] Advantageously, this approach enables precise and adaptive control of reagent spreading and drying, optimising staining quality and process efficiency for a wide range of protocols and reagent types.

[0303] According to another preferred embodiment of the second aspect of the invention, the air blowing unit is configured to remove a reagent from the sample and / or sample holders by blowing air over said reagent; and / or blow air over the reagent only until a moisture level is left on the sample and / or sample holder (41) adequate to facilitate further reagent dispensing or reagent spreading over said sample and / or sample holder (41).

[0304] It is noted that in some embodiments, the air blowing unit may be configured to remove a reagent from the surface of the sample and / or sample holder (41) by directing a controlled flow of air over the reagent, thereby facilitating the removal of excess reagent, unbound staining solution, or wash buffer. The removal may be achieved by adjusting parameters such as air flow rate or air speed, pressure, temperature, and duration, preferably by adjusting air speed, which may be selected based on the type and viscosity of the reagent, the geometry of the sample holder (41), and the requirements of the staining protocol. The air flow may be delivered as a continuous stream, pulsed bursts, or oscillating patterns, and may be directed at various angles or distances to optimise removal efficiency. In some embodiments, the air blowing unit may comprise interchangeable nozzles or diffusers to adapt the air stream profile to different sample formats, such as slides, wells, or cassettes. In some embodiments the extent of reagent removal may be monitored by sensors, such as optical sensors, humidity sensors, or weight sensors, which may provide feedback to the processor for real-time adjustment of the blowing parameters.

[0305] It is further noted that, alternatively or additionally, the air blowing unit may be configured to blow air over the reagent only until a specific moisture level remains on the sample and / or sample holder (41), wherein the residual moisture is adequate to facilitate subsequent reagent dispensing or spreading. The target moisture level may be determined based on the requirements of the next protocol step, for example, to ensure optimal reagent mixing, spreading, or reaction kinetics. The moisture level may be measured by integrated humidity sensors, capacitive sensors, or by indirect methods such as monitoring the evaporation rate or surface reflectance. In some embodiments, the device may allow the user to pre-set or automatically determine the desired moisture threshold, and the air blowing unit may terminate or modulate the air flow accordingly. The system may also be configured to, in some embodiments, adapt the blowing parameters dynamically in response to environmental conditions, such as ambient humidity or temperature, to maintain consistent results across different laboratory settings.

[0306] Advantageously, this configuration enables precise control over reagent removal and residual moisture, thereby improving staining consistency and facilitating optimal conditions for subsequent reagent application or spreading. This approach minimises the risk of over-drying or incomplete reagent removal, supporting high-quality and reproducible sample processing.

[0307] According to another preferred embodiment of the second aspect of the invention, the air blower (60) has a shape and orientation adequate to evenly spread the reagents over the sample and / or sample holder (41), preferably such that the air is blowed at an angle between 10 to 80 degrees with respect to the normal to the surface of the sample holder, more preferably between 25 and 65 degrees, even more preferably between 35 and 55.

[0308] It is noted that in some embodiments, the air blower (60) may comprise a geometry and nozzle configuration specifically selected or adjustable to promote uniform distribution of air across the surface of the sample and / or sample holder (41). The shape of the air blower (60) may include, for example, linear slot nozzles, air knives, circular or elliptical outlets, multi-orifice arrays, or diffusers, among other options, each designed to generate a laminar or turbulent air flow profile suitable for even reagent spreading. The orientation of the air blower (60) relative to the sample holder surface may be fixed or dynamically adjustable, for instance, by means of servo motors, stepper motors, or manual adjustment mechanisms, to achieve a desired angle of incidence. In some embodiments, the angle at which air is delivered may be set between, for example, 10 and 80 degrees with respect to the normal to the surface of the sample holder, as measured by a digital inclinometer, optical encoder, or mechanical protractor, and may be further refined to preferred subranges such as between 25 and 65 degrees, or more preferably between 35 and 55 degrees, among other options. The selection of angle may be determined based on the type of reagent, viscosity, sample geometry, or protocol requirements, and may be stored as part of the protocol in the device memory. It is also noted that the air blower (60) thus in some embodiments may remove fluid or reagent from the sample or sample holder (41) and / or spread the fluid or reagent over the sample or sample holder (41), for example during total or partial removal of fluid or reagent it could also be spreading said fluid or reagent.

[0309] Alternatively, the air blower (60) may comprise interchangeable or modular nozzle attachments to accommodate different sample formats or reagent types, allowing for rapid reconfiguration of the air flow pattern and angle. In some embodiments, the air blower (60) may be integrated with sensors, such as flow visualization cameras or surface wetness detectors, to provide feedback on the uniformity of reagent spreading and enable real-time adjustment of the blower orientation or output. The air flow speed and distribution may be characterized by techniques such as particle image velocimetry (PIV), hot-wire anemometry, or computational fluid dynamics (CFD) simulations, and may be specified in units such as metres per second (m / s) or litres per minute (L / min). The system may further comprise software algorithms to optimize the combination of blower shape, orientation, and air flow parameters for different staining protocols or sample types.

[0310] Advantageously, this configuration ensures that reagents are spread evenly and reproducibly over the sample surface, minimizing artefacts and improving staining consistency across a wide range of sample holders and reagent types.

[0311] According to another preferred embodiment of the second aspect of the invention, the air blower (60) comprises one or more orifices or slits (64) to blow the air, and: a. the longitudinal axis (62) of the air blower has a length (61) substantially the same as the width (45) of the sample holders (41) and / or sample holder supports (42), or has a difference in length with said width of ± 25%; and / or b. if the air blower comprises only one orifice, said orifice is elongated in the direction of the longitudinal axis (62) of the air blower (60), wherein the maximum length of said orifice is substantially the same as the width (45) of the sample holders (41) and / or sample holder supports (42), or has a difference in length with said width of ± 25%; and / or c. if the air blower comprises more than one orifice, said orifices are placed along the longitudinal axis (62) of the air blower (60), wherein the two more distanced orifices with respect to each other are separated such that at least part of one of the orifices is separated from at least part of the other orifice a distance substantially the same as the width (45) of the sample holders (41) and / or sample holder supports (42), or has a difference in length with said width of ± 25%.

[0312] It is noted that in some embodiments, the air blower (60) may comprise one or more orifices (64) configured to direct air flow, wherein the arrangement and geometry of the orifices are selected to optimise coverage of the sample holders (41) and / or sample holder supports (42). The longitudinal axis (62) of the air blower (60) may, in some embodiments, have a length (61) that is substantially the same as the width (45) of the sample holders (41) and / or sample holder supports (42), or may differ from said width by up to ± 25%, as determined, for example, by direct measurement using calipers or optical methods, with the width (45) and length (61) expressed in millimetres or centimetres, among other options. This configuration may be achieved by selecting or manufacturing air blowers (60) with a range of lengths, for example, between 20 mm and 30 mm, or by providing adjustable or modular blower housings to accommodate different sample formats. In some embodiments, the air blower (60) may comprise a single orifice (64) that is elongated in the direction of the longitudinal axis (62), wherein the maximum length of the orifice is substantially the same as the width (45) of the sample holders (41) and / or sample holder supports (42), or may differ by up to ± 25%. The elongated orifice may have a slot-like geometry, for example, with a width between 0.1 mm and 10 mm and a length between 10 mm and 50 mm, preferably between 20 mm and 30 mm, among other options, and may be fabricated by extrusion, milling, injection or additive manufacturing.

[0313] Alternatively, in embodiments where the air blower (60) comprises more than one orifice (64), the orifices may be distributed along the longitudinal axis (62) such that the two most distanced orifices are separated by a distance substantially the same as the width (45) of the sample holders (41) and / or sample holder supports (42), or may differ by up to ± 25%. The orifices (64) may be circular, elliptical, rectangular, or of other shapes, and may have diameters or widths between, for example, 1 mm and 10 mm, among other options. The placement of the orifices (64) may be determined by the geometry of the sample holder (41), the desired air flow profile, or the requirements of the staining protocol, and may be fixed or adjustable by means of sliding mounts or interchangeable nozzle plates. In some embodiments, the air blower (60) may comprise a combination of elongated and discrete orifices, or may allow for selective activation of individual orifices to tailor the air flow pattern to specific sample formats.

[0314] Advantageously, this configuration enables uniform and efficient delivery of air across the entire width of the sample holders (41) and / or sample holder supports (42), thereby promoting even reagent spreading or drying and improving the reproducibility and quality of the staining process.

[0315] According to another preferred embodiment of the second aspect of the invention, each blowing step is related to a blowing profile, each profile defining at least one parameter of the air flow selected from: air flow rate, speed, position or distance to the sample, speed of the “sweeping” movement of the robotic arm (10) or air blowing unit (60), and power applied by the pressure source, preferably wherein the protocol comprises a plurality of dispensing steps related each to at least one reagent or fluid and optionally to a blowing profile, more preferably said blowing profile determined based on a reagent associated to a reagent dispensing step of the protocol, preferably the previous, current or following dispensing step.

[0316] It is noted that in some embodiments, each blowing step may be associated with a specific blowing profile, wherein the blowing profile comprises at least one parameter of the air flow, such as, but not limited to, air speed or flow rate, position or distance to the sample, robotic arm speed, power applied by the pressure source, and travel path or segment of the sample holder. The air flow rate may be specified in units such as litres per minute (L / min), cubic centimetres per second (cm3 / s), or standard cubic feet per minute (SCFM), and may be measured by integrated flow sensors or calculated based on blower specifications. The position or distance to the sample may be defined in millimetres or centimetres, for example, between 1 mm and 100 mm, and may be adjusted by the robotic arm or by automated height adjustment mechanisms. The power applied by the pressure source may be expressed in watts (W), percentage of maximum output, or as a pressure value in pascals (Pa), bars, or psi, and may be regulated by electronic controllers or variable speed drives. In some embodiments, the blowing profile may further comprise additional parameters, such as air temperature, humidity, duration of the blowing step, or the angle of air incidence, among other options, to provide a more comprehensive control of the air flow characteristics.

[0317] In further embodiments, the blowing profile may be defined as a sequence of stages or segments along the travel path of the sample holder (porta), wherein each stage corresponds to a specific percentage or portion of the total travel distance. For example, the blowing profile may specify that from 0% to 70% of the travel distance, a first set of air flow parameters is applied, while from 70% to 100% of the travel distance, a second set of parameters is used. Each stage or segment may be associated with distinct configurations of parameters, such as blowing height (distance from the air outlet to the sample), compressor power, and the speed of movement of the robotic arm. This allows for dynamic and localised adjustment of the air flow characteristics along the sample holder’s path, optimising the effect of the blowing step according to the specific requirements of each region or process phase. The travel path and its segmentation may be user-defined or automatically determined by the system based on protocol requirements or sample characteristics.

[0318] It is further noted that the protocol may comprise a plurality of dispensing steps, each related to at least one reagent and, optionally, to a blowing profile. The association between dispensing steps and blowing profiles may be defined in the protocol memory, allowing for customisation of air flow parameters for each reagent or process stage. In some embodiments, the blowing profile may be determined based on the reagent associated with a dispensing step of the protocol, preferably the previous, current, or following dispensing step, taking into account factors such as reagent viscosity, volatility, required drying or spreading effect, or compatibility with subsequent reagents. The determination of the blowing profile may be implemented by pre-defined rules, user input, or by automated algorithms that select optimal parameters based on stored reagent properties or historical process data. Alternatively, insome embodiments the system may allow for real-time adjustment of the blowing profile in response to sensor feedback or environmental conditions, allowing consistent and reproducible results across different protocols and sample types.

[0319] Additionally, the blowing profile may include the selection of a specific travel path or segment of the sample holder, with the possibility to edit and assign different parameter values (such as blowing height, compressor power, robotic arm speed, and travel distance) to each segment. This enables the creation of complex, multi-stage blowing profiles tailored to the geometry of the sample holder, the distribution of reagents, or the desired process outcome. For instance, the system may be configured so that during the initial portion of the travel (e.g., 0-70%), a gentle air flow at a higher distance is applied to avoid disturbing freshly dispensed reagents, while in the final portion (e.g., 70-100%), a stronger and closer air flow is used to ensure complete drying or spreading. The parameters for each segment can be individually programmed or selected from pre-set profiles, providing maximum flexibility and process control.

[0320] Advantageously, the use of individualised blowing profiles for each blowing step enables precise and adaptive control of reagent spreading and drying, optimising staining quality and process efficiency for a wide variety of reagents and protocol requirements. This approach further enhances the flexibility and customisability of the automatic staining device (1), supporting complex and highly tailored staining workflows. According to another preferred embodiment of the second aspect of the invention, the automatic staining device (1) further comprises an adjustable orientation mechanism configured to vary manually or automatically the angle at which air is blown with respect to the sample holder (41)

[0321] It is noted that in some embodiments, the automatic staining device (1) may comprise an adjustable orientation mechanism that is configured to allow manual variation of the angle at which air is blown with respect to the sample holder (41). The adjustable orientation mechanism may comprise, for example, a pivoting mount, a rotatable joint, a sliding bracket, a ball-and-socket joint, or a rack-and-pinion system, among other options, which may be actuated by the user to set the desired angle. The mechanism may include graduated markings, detents, or locking features to facilitate precise and repeatable adjustment, and may allow for angular variation over a range, for example, from 0 degrees (parallel to the sample holder surface) to 90 degrees (perpendicular to the sample holder surface), or any subrange thereof, as measured by a mechanical protractor, digital inclinometer, or visual reference. In some embodiments, the adjustable orientation mechanism may be integrated into the air blower (60) housing or the mounting interface with the robotic arm (10), and may be constructed from materials such as stainless steel, aluminium, engineering plastics, polymers, or composites to ensure durability and chemical resistance. The manual adjustment may be performed by means of a knob, lever, screw, or other user-operable element, and may be designed to require minimal force or tools for operation.

[0322] Alternatively, the adjustable orientation mechanism may be configured to allow for quickrelease or tool-less adjustment, enabling rapid reconfiguration of the air blowing angle between different protocol steps or sample formats. In some embodiments, the mechanism may be combined with visual or tactile feedback to confirm the selected angle, or may include a memory function to return to previously used positions. The range and resolution of angular adjustment may be selected based on the requirements of the staining protocols, the geometry of the sample holders (41), and the desired air flow characteristics, and may be specified in degrees or radians, for example, with increments of 0.5 degree, 1 degree, or other suitable values.

[0323] Advantageously, the inclusion of a manually adjustable orientation mechanism provides the user with direct and flexible control over the air blowing angle, facilitating optimal reagent spreading or drying for a wide variety of sample holders and staining protocols. According to another preferred embodiment of the second aspect of the invention, the automatic staining device (1) further comprises a protective element for protecting at least one orifice (64) or the air blower (60), configured to protect the air blower (60) from reagent splashes that could cause clogging or partial obstruction of the orifices. Said protective element may be around or partially around the air blower (60), for example a flap. In some embodiments, a protective element may be positioned in a dispensing probe for preventing reagent or fluid from said dispensing probe from reaching the air blower (60)

[0324] It is noted that in some embodiments, the automatic staining device (1) may comprise a protective element associated with the air blower (60), wherein said protective element is configured to shield at least one orifice (64) of the air blower (60) from reagent splashes. The protective element may comprise, for example, a mesh, grille, perforated plate, baffle, shield, or cover, among other options, and may be fabricated from materials such as stainless steel, chemically resistant polymers, glass, or coated metals. The geometry and placement of the protective element may be selected to ensure that it intercepts droplets or splashes of reagent that could otherwise reach and accumulate at the orifices (64), thereby reducing the risk of clogging or partial obstruction. In some embodiments, the protective element may be fixed or removable, and may be positioned directly in front of, around, or integrated with the orifices (64), with a spacing that allows for unimpeded air flow while providing effective protection. The dimensions of the protective element, such as mesh size or aperture diameter, may be selected based on the expected droplet size and air flow requirements, for example, mesh openings between 0.1 mm and 5 mm, among other options.

[0325] Alternatively, the protective element may comprise a hydrophobic or oleophobic coating to further prevent reagent adhesion, or may be designed with self-cleaning or replaceable features to facilitate maintenance. In some embodiments, the protective element may be transparent or translucent to allow visual inspection of the orifices (64), or may include integrated sensors to detect accumulation of reagent or blockages. The protective element may be attached to the air blower (60) by means of clips, screws, magnets, or snap-fit mechanisms, and may be designed for easy removal and cleaning. The configuration of the protective element may be adapted to different air blower (60) geometries, orifices(64) arrangements, and sample holder formats, providing flexibility for a range of device embodiments. In some embodiments, the protective element may be attached to any element of the robotic arm, for example to a washing probe.

[0326] Advantageously, the inclusion of a protective element for the air blower (60) reduces the risk of reagent-induced clogging or obstruction of the orifices (64), thereby supporting reliable and consistent air flow and minimising maintenance requirements. According to another preferred embodiment of the second aspect of the invention, the air blowing unit is configured to perform multiple passes of air blowing in a given blowing step, each pass having at least one parameter that may differ from a previous pass.

[0327] It is noted that in some embodiments, the air blowing unit may be configured to execute multiple passes of air blowing within a single blowing step, wherein each pass may be defined by at least one parameter that differs from a previous pass. The parameters that may vary between passes include, but are not limited to, air flow rate, air pressure, temperature, humidity, duration, angle of incidence, distance to the sample or sample holder, and spatial position of the air blower. For example, a first pass may be performed at a higher air flow rate to remove excess reagent or at a closer distance of the air blowing unit with respect to the sample holder, followed by a second pass at a lower flow rate, a bigger distance with respect to the sample holder or at a different angle to achieve uniform spreading or controlled drying. The number of passes may be pre-set or dynamically determined by the processor based on protocol requirements, sensor feedback, or user input, and each pass may be individually programmed or selected from a set of predefined profiles. The variation in parameters between passes may be implemented by adjusting blower speed, valve positions, actuator settings, or by repositioning the robotic arm, among other options. The sequence and characteristics of each pass may be stored in the memory as part of the blowing step definition, and may be adapted to the type of reagent, sample geometry, or environmental conditions.

[0328] Alternatively, the air blowing unit may comprise automated or manual controls to allow the user to define or modify the parameters of each pass in real time, or may employ feedback from integrated sensors, such as air flow sensors, humidity sensors, or cameras, to optimise the outcome of each pass. In some embodiments, the system may be configured to perform a variable number of passes, for example, between one and ten, among other options, with each pass tailored to achieve a specific effect, such as initial reagent removal, intermediate spreading, and final moisture adjustment. The parameters for each pass may be measured in standard units, such as litres per minute for air flow, or millimetres for distance, and may be recorded for process traceability and quality control.

[0329] Advantageously, the ability to perform multiple passes of air blowing with variable parameters within a single blowing step enables fine-tuned control over reagent removal, spreading, and drying, thereby enhancing staining consistency and process flexibility.

[0330] According to another preferred embodiment of the second aspect of the invention, the automatic staining device (1) further comprises a sensor arrangement for verifying air flow rate or pressure, wherein preferably the processor is configured to adjust the air blowing flow or speed and / or the position of the air blower (60) during a dispensing or blowing step. For example, to ensure a target partial or total drying or moisture level is achieved on the sample holder (41).

[0331] It is noted that in some embodiments, the automatic staining device (1) may comprise a sensor arrangement configured to verify air flow rate or pressure during operation. The sensor arrangement may include, for example, thermal mass flow sensors, differential pressure sensors, piezoresistive sensors, hot-wire anemometers, or capacitive pressure transducers, among other options, and may be positioned within the air blowing unit, along the air conduit, or proximate to the air blower (60) outlet. The air flow rate may be measured in units such as litres per minute (L / min), cubic centimetres per second (cm3 / s), or standard cubic feet per minute (SCFM), while pressure may be measured in pascals (Pa), bars, or pounds per square inch (psi). In some embodiments, the sensor arrangement may comprise a single sensor or a combination of multiple sensors to provide redundant or complementary measurements, and may be interfaced with the processor or a dedicated controlling unit for real-time data acquisition and analysis.

[0332] It is further noted that the processor, which in some embodiments may also function as the controlling unit, may be configured in some embodiments to adjust the compressor output, the air flow, air speed and / or the position of the air blower (60) during a dispensing or blowing step based on feedback from the sensor arrangement. The adjustment of compressor output or the air flow or speed may involve modulating the power supplied to the compressor, varying the duty cycle, or controlling valve positions to achieve the desired air flow rate or pressure. The position of the air blower (60) may be adjusted by actuating the robotic arm (10) or an associated positioning mechanism, thereby varying the distance, angle, or orientation relative to the sample holder (41). The processor may execute control algorithms or closed-loop feedback routines to ensure that a target partial or total drying level is achieved on the sample holder (41), wherein the drying level may be determined by pre-set protocol parameters, user input, or real-time sensor data, such as residual moisture measurements or surface reflectance.

[0333] Advantageously, the inclusion of a sensor arrangement for air flow or pressure verification, combined with processor-controlled adjustment of compressor output (air blown flow or speed) and air blower (60) position, enables precise and adaptive achievement of target drying levels, thereby improving process reliability and staining quality.

[0334] According to another preferred embodiment of the second aspect of the invention, the memory further comprises instructions that, when executed by the processor, cause the processor to select a specific blowing step, a blowing profile or to regulate one or more blowing parameters, based on the viscosity of the reagent just dispensed, the type of sample holder (41), information related to a reagent dispensing step, preferably the previous, current or the next step, or on the characteristics of the biological sample.

[0335] It is noted that in some embodiments, the memory may comprise instructions that, when executed by the processor, enable the automatic selection of a specific blowing step, the assignment of a blowing profile, or the regulation of one or more blowing parameters, wherein such selection or regulation is based on at least one of the following: the viscosity of the reagent just dispensed, the type of sample holder (41), information related to a reagent dispensing step, preferably the previous, current or the next step, or the characteristics of the biological sample. The viscosity of the reagent may be determined by direct measurement, for example, using a viscometer or rheometer, and may be expressed in units such as millipascal- seconds (mPa s) or centipoise (cP), or may be retrieved from a reagent database stored in the memory. The type of sample holder (41) may include, but is not limited to, glass slides, multiwell plates, coverslips, tissue cassettes, or microarray chips, and may be identified by the processor through user input, barcode or RFID scanning, or image analysis. Information related to a reagent dispensing step, preferably the previous, current or the next step, may comprise the identity, volume, or physical properties of the subsequent reagent, as well as the timing and sequence of application, all of which may be stored in the protocol. The characteristics of the biological sample may include parameters such as tissue type, thickness, fixation status, or previous processing history, and may be input manually, retrieved from a laboratory information system, or determined by integrated sensors or imaging systems.

[0336] In some embodiments, the instructions in the memory may implement decision rules, look-up tables, or machine learning algorithms to determine the optimal blowing step, profile, or parameter settings based on the aforementioned factors. For example, a reagent with high viscosity may trigger the selection of a blowing step with increased air flow rate or reduced distance to the sample holder (41) to ensure uniform spreading, while a fragile sample holder type may prompt the use of lower air pressure or a more diffuse air stream to prevent sample displacement. The regulation of blowing parameters may include, but is not limited to, air flow rate (in L / min or cm3 / s), air pressure (in Pa or bar), ambient temperature (in °C), duration (in seconds), angle of incidence (in degrees), air speed (in m / s), and / or distance to the sample (in mm or cm).

[0337] Advantageously, this configuration enables the automatic adaptation of blowing steps and parameters to the specific requirements of each reagent, sample holder, protocol stage, and biological sample, thereby optimising reagent spreading, drying, and overall staining quality.

[0338] According to another preferred embodiment of the second aspect of the invention, the air blower (60) is coupled to the robotic arm (10) such that the robotic arm (10) can move the air blower (60) from a parking position to a working position, preferably at different distances from the sample holder (41), so as to adapt to different sections of the dispensing protocol or sample geometry.

[0339] It is noted that in some embodiments, the air blower (60) may be mechanically or functionally coupled to the robotic arm (10) in a manner that enables the robotic arm (10) to reposition the air blower (60) between at least a parking position and a working position. The coupling may be achieved by means of mounting brackets, quick-release connectors, magnetic couplings, or integrated actuator interfaces, among other options, and may allow for secure attachment while permitting controlled movement. The parking position may be defined as a location where the air blower (60) is retracted, stowed, or otherwise positioned away from the sample holder (41) to avoid interference during non-blowing steps, maintenance, or sample loading. The working position may be defined as a location where the air blower (60) is optimally positioned to direct air flow over the sample holder (41) for reagent spreading, drying, or removal. In some embodiments, the robotic arm (10) may be programmed to move the air blower (60) to a plurality of working positions at different distances from the sample holder (41), for example, between 1 mm and 100 mm, as measured by integrated position encoders, laser rangefinders, or mechanical stops, among other options. The selection of distance and position may be determined based on the requirements of different sections of the protocol, the geometry of the sample holder (41), or the type of reagent being processed.

[0340] Alternatively, the system may allow for dynamic adjustment of the air blower (60) position in real time, for example, to accommodate variations in sample height, orientation, or to target specific regions of the sample holder (41). The movement of the air blower (60) by the robotic arm (10) may be coordinated with other device operations, such as reagent dispensing, washin, or sensor measurements, and may be controlled by pre-defined protocols, user input, or automated algorithms. In some embodiments, the parking and working positions may be user-configurable, and the transition between positions may be performed smoothly to minimise vibration or disturbance to the sample. The coupling mechanism may further comprise features to enable electrical or pneumatic connections, sensor integration, or quick maintenance access, supporting a wide range of device configurations and operational scenarios.

[0341] Advantageously, this arrangement allows for automated and flexible positioning of the air blower (60), enabling optimal adaptation to different protocol steps and sample geometries while minimising interference and maximising process efficiency.

[0342] In a third aspect of the invention, an automatic staining device (1) for dispensing staining reagents over one or more biological samples is disclosed, wherein one or more reagents are delivered through a controlled pressure-driven flow. The automatic staining device (1) comprises a pressure-driven fluid displacement mechanism; a memory and a processor in communication with said memory, the memory comprising information of at least one protocol, said protocol comprising information related to at least one reagent dispensing step and at least one spreading step; one or more sample holder supports (42) suitable to support one or more sample holders (41); one or more reagent vials (51); and a robotic arm (10) comprising at least one dispensing probe (80), wherein the dispensing probe (80) is fluidly connected to the pressure-driven fluid displacement mechanism. The automatic staining device (10) further comprises a spreading unit (30), preferably comprised in the robotic arm (10), comprising a spreading element (31), preferably a non-rotatable spreading element (31), wherein a longitudinal axis of said spreading element (31) is or can be placed parallel to the surface of the one or more sample holders (41) and / or the surface of the sample holder supports (42), and wherein the memory comprises instructions that, when executed by the processor, cause the device (1) to perform a spreading step based on the previous reagent dispensing step, more preferably based on the viscosity of the reagent previously dispensed, or more preferably depending in if the viscosity of the reagent dispensed is above or not a predetermined viscosity threshold whose value is preferably stored in the memory. The previous dispensed reagent is referred to as to the reagent that the spreading element (30) is going to spread, or act upon, because said reagent has already been dispensed, and thus that reagent’s viscosity is relevant to determine if spreading or not (if doing a spreading pass or activate the spreading unit or not), and / or in modifying spreading parameters such as number of passes or spreading actions, force, distance to the sample holder, and / or speed, among other possible parameters..

[0343] For example, a more viscous reagent may require a different spreading force, speed, or number of passes compared to a less viscous reagent, in order to achieve uniform coverage and avoid artifacts such as streaks or incomplete spreading. A less viscous reagent, on the other hand, may not necessitate using the spreader, thus saving in time and efficiency. This is particularly important in automated staining protocols where multiple reagents with varying properties are applied in sequence, and where optimal spreading of each reagent is critical for the quality and reproducibility of the staining result.

[0344] It is noted that in some embodiments, the spreading unit (30) may comprise a spreading element (31), preferably a non-rotatable spreading element (31), which may be constructed from materials such as, but not limited to, polytetrafluoroethylene (PTFE), silicone, stainless steel, glass, or chemically resistant polymers, among other options. The non-rotatable spreading element (31) may have a variety of shapes, such as a straight bar, blade, spatula, squeegee, or pad, and may be configured with a flat, curved, or contoured surface to optimise contact with the sample holder (41) or sample holder support (42). The longitudinal axis of the spreading element (31) may be defined as the axis running along the greatest dimension of the element, for example, the length of a bar or blade, and may be positioned or repositioned by the robotic arm (10) to be parallel to the surface of the sample holders (41) and / or sample holder supports (42), as determined by mechanical alignment, position encoders, or visual feedback. In some embodiments, the spreading element (31) may be fixed in orientation or may be adjustable to accommodate different sample geometries or holder formats, with the parallelism measured in degrees or by direct distance measurement, for example, within ±2 degrees or ±1 mm, among other options.

[0345] Alternatively, the spreading unit (30) may comprise a spreading element (31) that is not strictly non-rotatable, and in some embodiments, the spreading element (31) may be configured as a rotatable roller, a pad, a brush, or any other spreading element capable of distributing reagents over the sample surface. The spreading element (31) may be actuated by the robotic arm (10) to move linearly, oscillate, or follow a programmed path across the sample holder (41), with the spreading force, speed, and contact pressure controlled by the processor based on protocol requirements or sensor feedback. The spreading step may be performed after a reagent dispensing step, and the memory may comprise instructions to adapt the spreading parameters, such as speed, pressure, or number of passes, based on the viscosity of the reagent previously dispensed, which may be measured in millipascal-seconds (mPa s) or centipoise (cP), or retrieved from a reagent database. The spreading step may be further adapted based on the type of sample holder (41), the geometry of the sample, or the requirements of the staining protocol, and may be coordinated with other device operations for optimal process integration.

[0346] Advantageously, the inclusion of a spreading unit (30) with a non-rotatable or otherwise adaptable spreading element (31) enables precise and uniform distribution of reagents over the sample surface, particularly when spreading parameters are dynamically adjusted based on reagent viscosity, thereby improving staining consistency and quality.

[0347] According to a preferred embodiment of the third aspect of the invention, the spreading element (31) is coupled to the robotic arm via a mechanism that allows guided displacement along the vertical axis (Z axis) over a limited distance.

[0348] Therefore, In some embodiments, the spreading element (31) is configured to move relative to the sample holder along a direction substantially perpendicular to the plane defined by the sample holder, typically referred to as the vertical axis or Z axis (for example the normal direction to the surface of the sample holder or normal to the surface of the earth, or it could also be defined as the gravitational axis). The spreading element (31) is preferably mounted to the robotic arm or to the spreading unit (30) in such a manner that it is permitted to move along this axis over a limited distance or within a predetermined range of motion. This range may be defined, for example, as between 0.1 cm and 5 cm, preferably between 0.5 cm and 3 cm, and more preferably between 0.5 cm and 2 cm, depending on the specific application and sample holder geometry.

[0349] The movement of the spreading element (31) along the vertical axis may be described as a guided or constrained displacement, such that the spreading element (31) is not completely free to move along the Z axis, but rather is limited to a defined interval or play. Additionally, in some embodiments such guided movement may be provided by means that have certain tolerance or excess in the range of motion, so as to also allow a certain tilt during a “sweeping” motion of the spreader (for example able to tilt an angle <|), such as between -10 and 10°). This limited vertical movement may be relative to the robotic arm, to the spreading unit (30), or to the sample holder itself, depending on the mounting configuration. In some embodiments, the spreading element (31) is supported by the robotic arm when in a raised position, and, when lowered onto the sample holder, is permitted to move vertically within the allowed range independently of the robotic arm, so as to not break or damage the sample or sample holder and / or to adapt to its morphology. This arrangement allows the spreading element (31) to follow minor variations in the height or flatness of the sample holder, reducing the risk of breakage or excessive force during the spreading operation.

[0350] Alternatively, in some embodiments the spreading element (31) may be coupled to the robotic arm via a sliding or telescopic connection, or by means of a mechanism that provides a degree of vertical play, allowing the spreading element (31) to move along the Z axis within the defined range. In some embodiments, the spreading element (31) is mounted with a limited decoupling of movement along the vertical axis, such that, when in contact with the sample holder, it is able to move vertically within the predetermined range relative to the robotic arm or spreading unit (30).

[0351] The vertical movement may also be described as a limited or guided displacement, a constrained range, a defined interval, or a predetermined range of motion, all referring to the same concept of allowing the spreading element (31) to move along the Z axis only within a certain distance, rather than being completely fixed or completely free.

[0352] During operation, the robotic arm may perform a sweeping or swiping motion, moving the spreading element (31) horizontally along or across the surface of the sample holder. When the spreading element (31) is in contact with the sample holder, the robotic arm primarily provides the horizontal movement, while the spreading element (31) is permitted to move vertically within the allowed range to accommodate surface irregularities or to avoid applying excessive force. In this way, the spreading element (31) can effectively spread the reagent over the sample holder without risking damage to the holder or the sample.

[0353] In some embodiments, due to mechanical tolerances or intentional design, the spreading element (31) may also be permitted to tilt or pivot within a limited range during the sweeping motion. This tilting along an angle <|) may occur passively, as a result of the play in the mounting mechanism, or may be actively controlled, and can further assist in adapting to the surface of the sample holder or in achieving a more uniform spreading of the reagent.

[0354] Advantageously, the above configurations provide a robust and adaptable spreading mechanism, allowing the spreading element (31) to accommodate variations in sample holder geometry and reagent volume, while maintaining precise control over the spreading process and minimising the risk of sample holder breakage.

[0355] According to a preferred embodiment of the third aspect of the invention, the spreading element (31) alone or in combination with the supporting element (32), weights between 1 and 50 grams, preferably between 5 and 20 grams, even more preferably around 15 grams. Advantageously, this weight enables a proper spreading of the reagent or fluid without applying external force, just with the forced produced by the mass of the spreading element (31) or of the spreading element (31) plus the supporting element (32).

[0356] According to a preferred embodiment of the third aspect of the invention, the spreading unit

[0357] (30) further comprises a supporting element (32) attached to the spreading element (31), and a supporting piece (33) preferably attached to the robotic arm, wherein the spreading element

[0358] (31) is supported by the supporting element (32), and the spreading element (31) can be displaced parallel to a longitudinal axis of the supporting element (32) and / or in the direction perpendicular to the longitudinal axis of the spreading element (31), between a first configuration wherein the supporting piece (33) supports the weight of the spreading element (31), and a second configuration wherein the spreading element (31) is not supported by the supporting piece (33), such as when is supported by the sample holder (41) and / or the sample holder support (42) during a spreading step.

[0359] It is noted that in some embodiments, the spreading unit (30) may comprise a supporting element (32) that is mechanically attached to the spreading element (31), wherein the supporting element (32) may be configured as a bar, rail, rod, frame, or plate, fabricated from materials such as, but not limited to, stainless steel, aluminium, engineering plastics, or composites, among other options. The supporting piece (33) may be attached to the robotic arm (10) by means of fasteners, clamps, magnetic couplings, or quick-release mechanisms, and may be constructed to provide stable support for the spreading element (31) when in a resting or non-operational position. In some embodiments, the spreading element (31) may be supported by the supporting element (32) in such a way that it can be displaced either parallel to the longitudinal axis of the supporting element (32) or in a direction perpendicular to the longitudinal axis of the spreading element (31), for example, by means of linear bearings, sliding guides, telescopic joints, or articulated linkages. The displacement may be actuated manually, by the robotic arm (10), or by integrated actuators such as stepper motors or pneumatic cylinders, and may be measured in millimetres or centimetres, for example, with a typical range of travel between 1 mm and 200 mm, among other options.

[0360] In some embodiments, the system may be configured such that the spreading element (31) transitions between a first configuration, wherein the supporting piece (33) supports the weight of the spreading element (31), and a second configuration, wherein the spreading element (31) is not supported by the supporting piece (33) but instead is supported by the sample holder (41) and / or the sample holder support (42) during a spreading step. The transition between configurations may be achieved by vertical or horizontal displacement, pivoting, or by a combination of translational and rotational movements, and may be controlled by the robotic arm (10) or by gravity-assisted mechanisms. In some embodiments, the supporting element (32) and the spreading element (31) may be integrated as a single piece, such that the supporting function is inherent to the geometry of the spreading element (31) itself, for example, in the form of a monolithic blade, bar, or pad. Alternatively, the supporting element (32) may be detachable or interchangeable to accommodate different spreading elements (31) or to adapt to various sample holder formats. The system may further comprise sensors, such as force sensors or position encoders, to detect the configuration of the spreading element (31) and to ensure proper contact with the sample holder (41) during operation.

[0361] Advantageously, this arrangement allows for controlled and repeatable engagement and disengagement of the spreading element (31) with the sample holder (41), facilitating precise reagent spreading while minimising wear and supporting flexible adaptation to different sample geometries.

[0362] According to another preferred embodiment of the third aspect of the invention, the spreading unit (30) further comprises a supporting element (32) attached to the spreading element (31), and a supporting piece (33) preferably attached to the robotic arm, wherein the spreader supporting element (32) can slide or traverse the supporting piece (33) until certain point allowing to move the spreading unit (30) the length of the supporting piece (33) or less.

[0363] It is noted that in some embodiments, the supporting element (32) may be configured to be mechanically attached to the spreading element (31) and to be capable of sliding or traversing along the supporting piece (33), which is preferably fixed to the robotic arm (10). The sliding or traversing movement may be enabled by means of linear bearings, grooves, rails, telescopic guides, or low-friction bushings, among other options, and may be actuated manually, by gravity, or by the robotic arm (10) itself. The range of movement may extend up to the full length of the supporting piece (33) or may be limited to a partial length, for example, by mechanical stops, detents, or software-defined travel limits, with the displacement measured in millimetres or centimetres, such as, but not limited to, a range between 2 mm and 300 mm, among other options. The supporting piece (33) may be constructed from materials such as stainless steel, aluminium, polymers, or composites, and may have a cross-sectional geometry selected from, but not limited to, rectangular, circular, or T-shaped profiles to optimise rigidity and ease of sliding. In some embodiments, the supporting element (32) may comprise features such as handles, locking mechanisms, or position indicators to facilitate precise positioning and secure retention during operation.

[0364] Alternatively, the sliding or traversing mechanism may be implemented by means of a rack- and-pinion system, magnetic tracks, or motorised actuators, allowing for automated or programmable movement of the spreading unit (30) along the supporting piece (33). The system may further comprise sensors, such as position encoders, limit switches, or optical detectors, to monitor the position of the supporting element (32) relative to the supporting piece (33) and to provide feedback to the processor or controlling unit. The ability to move the spreading unit (30) the length of the supporting piece (33) or less enables flexible adaptation to different sample holder sizes, geometries, or protocol requirements, and may facilitate multiposition spreading, sequential processing of multiple samples, or retraction of the spreading element (31) for cleaning or maintenance.

[0365] Advantageously, this configuration allows for controlled and adjustable positioning of the spreading unit (30) along the supporting piece (33), thereby enhancing the versatility and operational range of the device for various sample formats and spreading tasks.

[0366] According to another preferred embodiment of the third aspect of the invention, the spreading support element (32) can tilt between -10 and 10 degrees with respect to the supporting piece (33) or the normal direction to the surface of the sample holder, or is configured to be able to tilt or passively tilt between -10 and 10 degrees with respect to the supporting piece (33), or the normal direction to the surface of the sample holder, so as to, for example, adapt to the topography of the sample.

[0367] It is noted that in some embodiments, the supporting element (32) may be capable of tilting relative to the supporting piece (33) within a range of, for example, -10 to 10 degrees, as measured by a digital inclinometer, mechanical protractor, or angular sensor, with the angle defined with respect to the longitudinal axis of the supporting piece (33). This tilting may occur passively, for example, due to a designed tolerance or clearance between the interface of the supporting element (32) and the supporting piece (33), such as a loose fit, slotted guide, or flexible coupling, allowing the supporting element (32) to respond to variations in the surface topology of the sample holder (41) or sample holder support (42) during a spreading step. The passive tilting may be facilitated by gravity, the weight of the spreading element (31), or the resistance encountered during movement over the sample, and may be further enabled by the use of low-friction materials, bushings, or compliant joints at the interface. In some embodiments, the tilting range may be adjustable or limited by mechanical stops, detents, or elastic elements, and may be selected to optimise adaptation to sample irregularities while maintaining sufficient control over the spreading process.

[0368] Alternatively, the tilting of the supporting element (32) may be actively controlled by integrated actuators, such as miniature servos or piezoelectric elements, or may be monitored by angular sensors to provide feedback to the processor or controlling unit. The ability to tilt within the specified angular range may be particularly advantageous for accommodating uneven, warped, or non-planar sample surfaces, ensuring consistent contact and reagent distribution. The tilting angle may be specified in degrees or radians, and may be recorded for process traceability or quality control. In some embodiments, the system may allow for user adjustment of the tilting tolerance or range, or may provide interchangeable supporting elements (32) with different tilting characteristics to suit various sample types or protocol requirements.

[0369] Advantageously, the capability of the supporting element (32) to tilt with respect to the supporting piece (33) enables improved adaptation to the surface topology of the sample, thereby promoting uniform reagent spreading and reducing the risk of incomplete coverage or sample damage.

[0370] According to another preferred embodiment of the third aspect of the invention, the longitudinal length of the spreading element is substantially the same as the width (45) of the sample holders (41) and / or sample holder supports (42), or has a difference in length size with respect to said width of ± 50%, preferably ± 25%.

[0371] It is noted that in some embodiments, the spreading element may comprise a longitudinal length that is selected to match, or be within a defined tolerance of, the width (45) of the sample holders (41) and / or sample holder supports (42). The term "substantially the same" may be interpreted as a length that is equal to the width (45) within a margin of, for example, ± 50%, and more preferably within ± 25%, as determined by direct measurement using calipers, rulers, or optical measurement systems, with the dimensions expressed in millimetres or centimetres, among other options. For instance, if the width (45) of a sample holder (41) is 25 mm, the spreading element may have a longitudinal length between 20 mm and 50 mm, or more preferably between 25 mm and 40 mm, among other possible configurations. The spreading element may be fabricated in a range of standard or custom lengths to accommodate different sample holder formats, or may be provided as an adjustable or modular component, such as a telescopic bar, an extendable blade, or a system of interlocking segments, to allow adaptation to various widths. In some embodiments, the spreading element may be interchangeable, allowing the user or the device to select the most appropriate length for a given sample holder (41) or support (42), or may comprise markings or guides to facilitate alignment with the sample width.

[0372] Alternatively, the spreading element may be designed with a length intentionally shorter or longer than the width (45) of the sample holder (41) or support (42), within the specified tolerance, to achieve specific spreading effects, such as partial coverage, edge-to-edge contact, or controlled overlap. The selection of the spreading element length may be based on the requirements of the staining protocol, the geometry of the sample, or the desired uniformity of reagent distribution. The length of the spreading element may be specified in the device memory as part of the protocol parameters, or may be detected automatically by integrated sensors or by image analysis. In some embodiments, the spreading element may be constructed from materials such as stainless steel, PTFE, silicone, polymers, or other chemically resistant materials, and may have a cross-sectional profile selected to optimise contact and spreading efficiency for the given length.

[0373] Advantageously, configuring the spreading element to have a longitudinal length substantially matching the width of the sample holder (41) or support (42) enables uniform and efficient reagent distribution across the entire sample surface. This approach minimises the risk of incomplete coverage or reagent pooling at the edges, thereby supporting consistent staining quality.

[0374] According to another preferred embodiment of the third aspect of the invention, the spreading element (31) is cylindrical or a prism.

[0375] It is noted that in some embodiments, the spreading element (31) may comprise a cylindrical geometry, wherein the cross-section is circular and the longitudinal axis extends along the length of the cylinder, or may alternatively comprise a prismatic geometry, such as, but not limited to, a rectangular, square, triangular, hexagonal, or other polygonal cross-section. The cylindrical spreading element (31) may be solid or hollow, and may be fabricated from materials such as stainless steel, PTFE, silicone, glass, or engineering plastics, among other options, with diameters ranging, for example, from 2 mm to 30 mm, and lengths selected according to the width of the sample holder (41) or sample holder support (42). In the case of a prismatic spreading element (31), the cross-sectional dimensions may be, for example, between 2 mm and 30 mm for each side, and the length may be adapted as previously described. The surface of the spreading element (31) may be smooth, textured, or coated to optimise reagent spreading, and the element may be fixed or detachable for cleaning or replacement. In some embodiments, the choice between cylindrical and prismatic geometries may be determined by the type of reagent, the desired spreading profile, or the characteristics of the sample holder

[0376] (41), and the device may be configured to accept interchangeable spreading elements (31) of different shapes and sizes.

[0377] Advantageously, the use of a cylindrical or prismatic spreading element (31), preferably an uniform cylinder, provides consistent and uniform contact with the sample surface, facilitating even reagent distribution and supporting reliable staining results.

[0378] According to another preferred embodiment of the third aspect of the invention, the spreading element (31) comprises a distal part (311), a proximal part (312), and a central part (313), wherein a diameter (D1) of the distal (311) and proximal part (312) is the same for both distal (311) and proximal (312) parts, and is bigger than a diameter (D2) of the central part (313), wherein the diameter (D1) of the distal (311) and proximal (312) parts is between 0.02 and 2 mm longer than the diameter (D2) of the central part (313), preferably is between 0.06 and 0.6 mm longer, more preferably is between 0.1 and 0.3 mm longer.

[0379] It is noted that in some embodiments, the spreading element (31) may comprise a segmented geometry including a distal part (311), a proximal part (312), and a central part (313), wherein the distal (311) and proximal (312) parts each have a diameter (D1) that is equal for both ends and greater than the diameter (D2) of the central part (313). The difference in diameter (D1 minus D2) may be selected, for example, between 0.02 mm and 2 mm, as measured by calipers, micrometers, or optical measurement systems, and may be further refined to preferred subranges such as between 0.06 mm and 0.6 mm, or more preferably between 0.1 mm and 0.3 mm, among other options. The overall length and diameters of the spreading element (31) may be adapted to the width of the sample holder (41) or sample holder support

[0380] (42), and the transition between the distal / proximal parts and the central part may be abrupt or gradual, for example, by means of a step, taper, or fillet. The spreading element (31) may be fabricated from materials such as stainless steel, PTFE, silicone, glass, or engineering plastics, and the segmented geometry may be produced by machining, molding, extrusion, or additive manufacturing. In some embodiments, the distal (311) and proximal (312) parts may serve as end caps, collars, or reinforcement zones, and the central part (313) may be configured with a reduced diameter to optimise contact pressure, reagent flow, or spreading dynamics.

[0381] Alternatively, the segmented geometry may be implemented in spreading elements (31) of various cross-sectional shapes, such as cylindrical, prismatic, or elliptical, and the difference in diameter may be selected based on the viscosity of the reagent, the desired spreading profile, or the mechanical properties required for the application. The distal (311) and proximal (312) parts may be symmetrical or asymmetrical, and may include additional features such as textured surfaces, coatings, or integrated sensors. The central part (313) may be designed to minimise reagent retention or to facilitate cleaning, and the overall configuration may be adapted for use with interchangeable or modular spreading units (30). The diameters (D1 and D2) may be specified in millimetres, and the tolerances may be selected to ensure consistent performance across different device embodiments.

[0382] Advantageously, this segmented configuration of the spreading element (31) enables improved control of reagent distribution and minimises edge effects, supporting uniform spreading and reducing the risk of reagent pooling or incomplete coverage at the sample boundaries.

[0383] According to another preferred embodiment of the third aspect of the invention, the spreading element (31) is made of a material selected from: polymer, metal, ceramic, hybrid composite.

[0384] It is noted that in some embodiments, the spreading element (31) may comprise a material selected from, but not limited to, polymers, metals, ceramics, or hybrid composites. Examples of suitable polymers include, for instance, polytetrafluoroethylene (PTFE), polyether ether ketone (PEEK), polypropylene, polycarbonate, or silicone, among other options. Suitable metals may comprise stainless steel, aluminium, titanium, or nickel-based alloys, among others. Ceramics may include alumina, zirconia, silicon nitride, or glass-ceramic materials, among other alternatives. Hybrid composites may be formed by combining two or more of the aforementioned materials, such as polymer-ceramic composites, metal-polymer laminates, or fibre-reinforced polymers, among other options. In some embodiments, the material of the spreading element (31) may be selected to minimise adhesion of reagents and facilitate cleaning, for example, by employing low surface energy polymers, non-stick coatings, or surface treatments such as electropolishing, passivation, or plasma modification. Alternatively, the spreading element (31) may be fabricated from a functionally equivalent material that provides comparable chemical resistance, mechanical strength, and surface properties suitable for repeated contact with biological reagents and sample holders.

[0385] Advantageously, the selection of the spreading element (31) material from a broad range of polymers, metals, ceramics, or hybrid composites enables adaptation to diverse reagent chemistries and cleaning protocols, supporting durability and minimising reagent carryover.

[0386] According to another preferred embodiment of the third aspect of the invention, the spreading element (31) is configured to adapt its height (Z-axis) dynamically during spreading, thereby following the topography of the sample holder (41) or sample, and / or is configured to vary its inclination angle with respect to the Z-axis to optimize reagent spreading.

[0387] It is noted that in some embodiments, the spreading element (31) may be configured to passively or actively adapt its height along the Z-axis during a spreading step, thereby enabling the spreading element (31) to follow the surface topography of the sample holder (41) or the sample itself. Passive adaptation may be achieved, for example, by mounting the spreading element (31) on a compliant or spring-loaded mechanism, a floating carriage, a low-friction linear bearing, or a counterweighted support, which allows the spreading element (31) to move vertically within a defined range, such as, but not limited to, 0.1 mm to 10 mm, in response to variations in sample height or unevenness. Alternatively, active adaptation may be implemented by integrating electrical actuators, such as stepper motors, linear actuators, piezoelectric elements, or voice coil actuators, which are controlled by the processor or a dedicated control unit to dynamically adjust the Z-axis position of the spreading element (31) in real time. In some embodiments, the system may further comprise one or more sensors, such as contact sensors, force sensors, optical distance sensors, or laser triangulation sensors, to detect the position or force exerted by the spreading element (31) and to provide feedback for closed-loop control of the Z-axis movement. The adaptation of height may be continuous or stepwise, and may be specified in micrometres or millimetres, among other options, depending on the required precision and the topographical features of the sample holder (41) or sample.

[0388] It is further noted that, in addition to or instead of Z-axis adaptation, the spreading element (31) may be configured to vary its inclination angle with respect to the Z-axis, thereby optimising the contact profile and spreading efficiency for different sample geometries or reagent properties. The inclination angle may be adjusted passively, for example, by means of a gimbal mount, a compliant joint, or a tilting mechanism with a defined angular range, such as between -15 and +15 degrees, or actively by means of miniature servos, rotary actuators, or piezoelectric tilt elements, which may be controlled based on pre-set protocol parameters or real-time sensor feedback. The inclination angle may be measured by angular encoders, inclinometers, or potentiometers, and may be specified in degrees or radians. In some embodiments, the system may allow for simultaneous or independent adjustment of both Z- axis position and inclination angle, enabling the spreading element (31) to maintain optimal contact and spreading performance across complex or irregular sample surfaces. The configuration may be user-adjustable, automatically determined by the device, or programmable as part of the spreading protocol. Advantageously, this arrangement enables the spreading element (31) to maintain consistent contact and optimal spreading performance over samples with variable topography, thereby improving reagent distribution and staining uniformity.

[0389] According to another preferred embodiment of the third aspect of the invention, the spreading element (31) is coupled to a force control mechanism configured to limit the pressure exerted on the sample holder (41) during spreading, thereby preserving the integrity of fragile substrates or sensitive samples.

[0390] It is noted that in some embodiments, the spreading element (31) may be operatively coupled to a force control mechanism, which may comprise, for example, a spring-loaded mount, a compliant or elastic suspension, a magnetic or pneumatic buffer, or an active force feedback system utilising load cells, piezoresistive sensors, or strain gauges. The force control mechanism may be configured to limit the maximum pressure or force exerted by the spreading element (31) on the sample holder (41) during the spreading operation, with the force threshold selectable or adjustable according to the fragility of the substrate or the sensitivity of the biological sample. The pressure may be measured in units such as newtons (N), grams-force (gf), or pascals (Pa), and the mechanism may be designed to maintain the applied force within a defined range, for example, between 0.01 N and 5 N, among other options. In some embodiments, the force control mechanism may be passive, relying on mechanical compliance or pre-set spring constants, or may be active, with real-time feedback to the processor or controlling unit to dynamically modulate the position or movement of the spreading element (31) in response to measured force values. The system may further comprise user-adjustable settings, automatic detection of sample type, or protocol-based force profiles to optimise protection for a variety of sample holders (41), including, but not limited to, glass slides, polymer films, coated substrates, or microarray chips.

[0391] Alternatively, the force control mechanism may be integrated into the robotic arm (10) or the supporting structure of the spreading unit (30), and may include features such as force-limiting clutches, torque-limiting couplings, or electronically controlled actuators with programmable force limits. The mechanism may be configured to disengage, retract, or halt the spreading element (31) if the measured force exceeds a predetermined threshold, thereby preventing damage to fragile or sensitive samples. In some embodiments, the force control mechanism may also provide data logging or alert functions to record force events or notify the user of excessive pressure, supporting quality assurance and traceability. The configuration may be adapted to different spreading element (31) geometries, materials, and operational modes, and may be compatible with both manual and automated spreading protocols. Advantageously, the inclusion of a force control mechanism coupled to the spreading element (31) ensures that the pressure applied during spreading is limited to safe levels, thereby preserving the integrity of fragile substrates and sensitive biological samples. This feature supports reliable and reproducible reagent distribution without compromising sample quality.

[0392] According to another preferred embodiment of the third aspect of the invention, the spreading element (31) is configured to be raised from the sample holder (41) after a spreading step, and displaced to a washing station.

[0393] It is noted that in some embodiments, the spreading element (31) may be operatively configured to be raised from the surface of the sample holder (41) following completion of a spreading step, for example, by actuation of the robotic arm (10), a dedicated lifting mechanism, or a combination thereof. The raising action may be achieved by vertical displacement along the Z-axis, with the height of retraction selectable or programmable, for instance, between 1 mm and 100 mm, as measured by position encoders, limit switches, or optical sensors. Once raised, the spreading element (31) may be displaced laterally, linearly, or along a programmed path to a designated washing station, which may be located within the operational envelope of the robotic arm (10) or as a fixed module within the device. The displacement may be performed automatically under processor control or, in some embodiments, may be user-initiated. The washing station may comprise features such as one or more fluid jets, ultrasonic baths, spray nozzles, or immersion tanks, and may be configured to deliver cleaning solutions, rinsing fluids, or drying air to the spreading element (31), with the cleaning process monitored by sensors such as flow meters, optical detectors, or conductivity sensors.

[0394] Alternatively, the raising and displacement of the spreading element (31 ) to the washing station may be selectively performed depending on the requirements of the protocol, for example, only after certain spreading steps or based on information regarding the previous, current, or next reagent dispensing step. The control logic for determining when to initiate the washing sequence may be stored in the device memory and executed by the processor, allowing for protocol-dependent or reagent-specific cleaning cycles. The washing station may be designed to accommodate spreading elements (31) of various geometries and materials, and may include features for automated drying, inspection, or replacement of the spreading element (31) as required by the protocol or maintenance schedule.

[0395] Advantageously, this configuration enables automated and protocol-adaptive cleaning of the spreading element (31), thereby reducing cross-contamination between reagents and supporting consistent staining quality. According to another preferred embodiment of the third aspect of the invention, the automatic staining device (1) further comprises a washing receptacle (79) suitable to fit the length of the spreading element.

[0396] It is noted that in some embodiments, the washing receptacle (79) may be dimensioned to accommodate the full longitudinal length of the spreading element (31), with an internal length equal to or greater than the length of the spreading element, for example, between 20 mm and 300 mm, among other options, as measured by calipers or rulers. The washing receptacle (79) may comprise a trough, channel, or container fabricated from materials such as, but not limited to, stainless steel, polypropylene, PTFE, glass, or other chemically resistant materials, and may be configured with an open or closed top, sloped or flat bottom, and optional drainage or overflow features. In some embodiments, the receptacle (79) may be fixed or removable, and may be positioned within the operational range of the robotic arm (10) to facilitate automated transfer of the spreading element (31) for cleaning. The receptacle (79) may further comprise integrated fluid inlets or spray nozzles for delivery of washing solutions, rinsing fluids, or drying air, and may be equipped with sensors to monitor fluid level, temperature, or contamination.

[0397] Alternatively, the washing receptacle (79) may be designed to accommodate spreading elements (31) of varying lengths by means of adjustable end stops, telescopic walls, or modular inserts, thereby supporting compatibility with different device configurations or sample holder formats. The receptacle (79) may also include features such as removable racks, perforated trays, or magnetic holders to secure the spreading element (31) during washing, and may be configured for manual or automated cleaning cycles. In some embodiments, the receptacle (79) may be integrated with waste collection systems, filtration units, or recirculation pumps to manage cleaning fluids efficiently and minimise maintenance.

[0398] Advantageously, the inclusion of a washing receptacle (79) suitable to fit the length of the spreading element (31) enables thorough and efficient cleaning of the spreading element, thereby reducing cross-contamination and supporting consistent staining quality.

[0399] According to another preferred embodiment of the third aspect of the invention, the washing receptacle (79) comprises a cover or protective element configured to prevent splashing of cleaning solution outside the washing area or over the spreading element (31) or a dispensing probe or another element of the device..

[0400] It is noted that in some embodiments, the washing receptacle (79) may comprise a cover or protective element, which may be configured as a lid, shield, baffle, splash guard, or enclosure, among other options, and may be fabricated from materials such as, but not limited to, transparent or opaque polymers, stainless steel, glass, or chemically resistant composites. The cover or protective element may be fixed, hinged, sliding, removable, or flexible, and may be dimensioned to fully or partially enclose the washing area of the receptacle (79), thereby preventing the egress of cleaning solution droplets or splashes during washing operations. In some embodiments, the cover may include apertures, seals, or flexible membranes to allow the entry and exit of the spreading element (31) while maintaining an effective barrier against splashing. The geometry and placement of the cover or protective element may be selected to ensure that any cleaning solution projected during washing is contained within the receptacle (79), and to minimise the risk of droplets reaching the upper part of the spreading element, the robotic arm, or any adjacent device components.

[0401] Alternatively, the cover or protective element may comprise features such as angled surfaces, drip edges, or integrated drainage channels to direct any incidental splashes back into the washing receptacle (79), and may be designed to facilitate easy cleaning, inspection, or replacement. In some embodiments, the cover may be transparent to allow visual monitoring of the washing process, or may include integrated sensors to detect the presence of the spreading element (31) or the accumulation of cleaning solution. The configuration of the cover or protective element may be adapted to accommodate spreading elements (31) of different lengths or geometries, and may be compatible with automated or manual washing protocols. The objective of the cover or protective element is also to prevent cleaning solution from splashing onto the upper part of the spreading element or any other nearby element, which could subsequently drip onto a sample holder during the staining sequence.

[0402] Advantageously, the inclusion of a cover or protective element on the washing receptacle (79) effectively prevents the escape of cleaning solution splashes, thereby reducing the risk of contamination or unwanted dripping onto sample holders during subsequent processing steps.

[0403] According to another preferred embodiment of the third aspect of the invention, the washing receptacle (79) is configured to clean the spreading element (31) by at least one of: direct fluid dispensing, preferably from one or more dispensing orifices within the washing receptable (79), immersion of the spreading element (31) in a cleaning solution, or a combination thereof, preferably for a predetermined period of time based on the previous reagent dispensing steps.

[0404] It is noted that in some embodiments, the washing receptacle (79) may be configured to clean the spreading element (31) by means of direct fluid dispensing, which may be achieved through one or more dispensing orifices integrated within the washing receptacle (79). The orifices may be arranged along the length or at specific positions of the receptacle (79) to ensure comprehensive coverage of the spreading element (31), and may have diameters, for example, between 0.2 mm and 3 mm, among other options. The fluid dispensing may be controlled by a pump, pressurised reservoir, or gravity feed, and the flow rate may be regulated in units such as millilitres per minute (ml / min) or litres per hour (L / h), as measured by flow sensors or timers. Alternatively, or in addition, the washing receptacle (79) may be configured to allow immersion of the spreading element (31) in a cleaning solution, wherein the solution may be introduced and drained automatically or manually, and the immersion depth may be selected to ensure that the entire working surface of the spreading element (31) is submerged. The cleaning process may be performed for a predetermined period of time, for example, between 5 seconds and 5 minutes, among other options, with the duration determined based on information from previous reagent dispensing steps, such as the type or viscosity of the reagent previously applied.

[0405] In some embodiments, the washing receptacle (79) may further comprise automated features such as fluid level sensors, temperature sensors, or contamination detectors to monitor the cleaning process and ensure optimal performance. The draining of used cleaning solution may be facilitated by integrated valves, pumps, or gravity outlets, and the receptacle (79) may be configured to execute cleaning cycles automatically under processor control, based on protocol requirements or sensor feedback. The system may allow for the selection of different cleaning solutions, such as water, detergents, solvents, or disinfectants, and may be programmed to adapt the cleaning parameters according to the nature of the reagents used in previous steps. The receptacle (79) may also include agitation mechanisms, such as ultrasonic transducers or mechanical shakers, to enhance cleaning efficiency, and may be designed for easy maintenance and replacement of consumables.

[0406] Advantageously, this configuration enables thorough and protocol-adaptive cleaning of the spreading element (31), supporting reliable operation and minimising cross-contamination between staining steps.

[0407] According to another preferred embodiment of the third aspect of the invention, the automatic staining device (1) further comprises a drying station for the spreading element (31), said drying station comprising an absorbent surface suitable to remove residual cleaning solution from the spreading element (31) by contact, preferably wherein the spreading element (31) is displaced over the absorbent surface in one or more passes.

[0408] It is noted that in some embodiments, the drying station may comprise an absorbent surface configured to remove residual cleaning solution from the spreading element (31) by direct contact. The absorbent surface may be fabricated from materials such as, but not limited to, cellulose-based pads, microfiber cloths, nonwoven synthetic fabrics, polyurethane foams, or superabsorbent polymer composites, among other options. The absorbent surface may be arranged as a flat pad, a contoured strip, a roller, or a series of spaced elements, and may be replaceable or washable to maintain hygiene and performance. In some embodiments, the spreading element (31) may be displaced over the absorbent surface in one or more passes, either linearly, rotationally, or in a programmed path, with the number of passes, speed, and contact pressure determined by the processor, user input, or protocol requirements. The displacement may be achieved by actuation of the robotic arm (10), a dedicated drive mechanism, or by gravity-assisted movement, and the contact force may be controlled to optimise drying efficiency while minimising wear on the spreading element (31) or the absorbent surface. The drying station may be positioned adjacent to or integrated with the washing receptacle (79), and may include features such as guides, ramps, or alignment fixtures to ensure consistent contact between the spreading element (31) and the absorbent surface.

[0409] Alternatively, the absorbent surface may be configured as a multi-layered or zoned structure, with different materials or textures selected to maximise fluid uptake and minimise residue. The absorbent surface may be treated with hydrophilic, antimicrobial, or non-stick coatings to enhance performance and durability, and may be monitored by sensors, such as moisture sensors or optical detectors, to determine saturation level and prompt replacement or maintenance. In some embodiments, the drying station may be designed to accommodate spreading elements (31) of various geometries and lengths, and may be compatible with both manual and automated cleaning and drying cycles. The system may further allow for the adjustment of drying parameters, such as the number of passes or the dwell time on the absorbent surface, based on the type of cleaning solution used, the geometry of the spreading element (31), or the requirements of the subsequent staining protocol step.

[0410] Advantageously, the inclusion of a drying station with an absorbent surface enables efficient removal of residual cleaning solution from the spreading element (31), thereby reducing the risk of cross-contamination and ensuring optimal reagent application in subsequent staining cycles.

[0411] According to a fourth aspect of the invention, an automatic staining device for dispensing staining reagents over one or more biological samples is disclosed. The automatic staining device comprising: a pressure-driven fluid displacement mechanism; a memory and a processor in communication with said memory; one or more sample holder supports suitable to support one or more sample holders; one or more reagent vials; a robotic arm comprising at least one dispensing probe, wherein the dispensing probe is fluidly connected to the pressure-driven fluid displacement mechanism; one or more sensors selected from: accelerometers, gyroscopes, magnetometers, encoders, current sensors, temperature sensors, flow sensors, and capacitive sensors. The system characterized in that the memory comprises instructions that, when executed by the processor, cause the processor to: collect data from at least one or more of said sensors over a temporal window, thereby generating a data matrix of sensor values; and provide said data matrix as input to at least one embedded artificial intelligence model configured to perform at least one of:

[0412] - classification of the activity of the robotic structure, detection of anomalies in the robotic functionality, detection of anomalies in the robotic structure,

[0413] - detection of inclination or loss of horizontal stability,

[0414] - determination of the quality of the support structure, estimation of air blowing unit wear or or air blowing compressor wear,

[0415] - detection of activation of hydraulic pumps,

[0416] - estimation of heater wear,

[0417] - detection of liquid level, wherein the output of the embedded artificial intelligence model is used to generate a diagnostic, warning, or control signal for the operation of the automatic staining device.

[0418] According to another preferred embodiment of the fourth aspect of the invention, the memory comprises a plurality of embedded artificial intelligence models, each configured to process a respective subset of sensor data and to output a respective diagnostic or classification result, wherein the processor is configured to execute said models in parallel or sequentially.

[0419] According to another preferred embodiment of the fourth aspect of the invention, at least one embedded artificial intelligence model is a neural network trained for N-class classification of the activity of the robotic structure, using as input data from at least one of: accelerometer, gyroscope, magnetometer, encoder, and current sensors.

[0420] According to another preferred embodiment of the fourth aspect of the invention, at least one embedded artificial intelligence model is an anomaly detection model configured to detect abnormal movement or operation of the robotic structure, using as input data from at least one of: accelerometer, gyroscope, magnetometer, encoder, and current sensors.

[0421] According to another preferred embodiment of the fourth aspect of the invention, at least one embedded artificial intelligence model is configured to detect inclination or loss of horizontal stability of the device, using as input data from at least one of: accelerometer, gyroscope, magnetometer, and encoder sensors, and to output a warning if the inclination exceeds a predetermined threshold. According to another preferred embodiment of the fourth aspect of the invention, at least one embedded artificial intelligence model is a regression model configured to estimate the quality index of the support structure of the device, based on inertial sensor data.

[0422] According to another preferred embodiment of the fourth aspect of the invention, at least one embedded artificial intelligence model is a regression model configured to estimate the wear of the compressor or hydraulic pumps, based on a comparison of estimated and actual usage time, and / or flow sensor data.

[0423] According to another preferred embodiment of the fourth aspect of the invention, at least one embedded artificial intelligence model is a regression model configured to estimate the wear of the reaction chamber heaters, based on a comparison of applied electrical power and measured thermal output, using as input data from temperature sensors and power measurement data.

[0424] According to another preferred embodiment of the fourth aspect of the invention, at least one embedded artificial intelligence model is a classification model configured to detect the activation state of hydraulic pumps, using as input data from accelerometer, gyroscope, and magnetometer sensors.

[0425] According to another preferred embodiment of the fourth aspect of the invention, at least one embedded artificial intelligence model is a classification model configured to detect the liquid level in a reagent vial, using as input data from a capacitive sensor and a Z-axis encoder, and optionally further comprising as input the Z-axis speed, Z-axis position, liquid identification, and / or a basic liquid detection algorithm.

[0426] According to a fifth aspect of the invention, an automatic staining device for dispensing staining reagents over one or more biological samples is disclosed, the automatic staining device comprising: a pressure-driven fluid displacement mechanism; a memory and a processor in communication with said memory; one or more sample holder supports (42) suitable to support one or more sample holders (41); one or more reagent vials (51); a robotic arm (10) comprising at least one dispensing probe (80), wherein the dispensing probe (80) is fluidly connected to the pressure-driven fluid displacement mechanism; at least one inertial sensor selected from: accelerometer, gyroscope, and magnetometer. Wherein the device is characterized in that the memory comprises instructions that, when executed by the processor, cause the processor to: collect data from the at least one inertial sensor during execution of a staining protocol; provide said data to an embedded artificial intelligence model trained to detect loss of horizontal stability or excessive inclination of the device or its support table; compare the output of the artificial intelligence model to a predetermined threshold; and if the output indicates a loss of stability or inclination exceeding the threshold, generate a warning signal and / or interrupt the staining protocol.

[0427] According to another preferred embodiment of the fourth aspect of the invention, the warning signal is displayed on a user interface and / or transmitted to a remote monitoring system.

[0428] CLAUSES

[0429] 1. A computer-implemented method for performing treatments of histological staining, immunoenzymatic staining and hybridisation on tissue sections or cell smears, comprising the steps of: i. disposing one or more previously prepared samples on at least one sample holder, optionally labelled with identification codes, in a staining or hybridisation area, segmented into a plurality of compartments that act as reaction chambers for a plurality of staining or hybridisation treatments on said samples disposed therein; ii. capturing at least one image by means of a high-precision photographic image capture device, of the position of each sample holder in the respective reaction chamber and, optionally, the identification code disposed on the sample holder; iii. capturing at least one high-precision image of one or more samples disposed on said sample holder by a high-precision photographic image capture device; iv. capturing at least one high-precision image of one or more reagent vials; v. performing said treatments on the samples disposed in the compartments acting as reaction chambers, by means of the provision of a robotic device, configured for its three- dimensional displacement along the three axes XYZ, and adapted to be positioned at any point of said staining or hybridisation area of the compartments located in the horizontal plane XY, and which carries out, by means of a general purpose microcontroller with a computer program, the control of the processes that comprise:

[0430] - processing at least one image of the sample holder by Artificial Intelligence (Al) and, optionally, of the label with codes disposed thereon, wherein the processing of the at least one image of the sample holder comprises determining the position of each sample holder;

[0431] - dispensing specific reagents or common reagents based on a pre-programmed protocol of processes to be applied, depending on:

[0432] - the reading of identification codes disposed on specific reagent reservoirs; - the quantity, location and time of dispensing;

[0433] - image capture of at least one sample, processed by Artificial Intelligence (Al), wherein the processing comprises tissue detection directly performed by a CNN neural network model;

[0434] - image capture of at least one reagent vial, processed by Al, wherein the processing of said image of the reagent vials includes the detection of whether the vial is open or closed;

[0435] - performing washes after each process protocol and / or each process independently; and

[0436] - obtaining data from the images captured of each sample and, optionally, related to each identification code of the sample holders. The method according to claim 1 , wherein the labels of the sample holders have codes that allow identifying the sample, the case to which it corresponds, the protocol to be operated therein and the order of priority to be applied to each sample and / or each protocol. The method according to claim 1 , characterised in that the microcontroller carries out the control of the following functions: a) start the method; b) check that the lid is closed and activate a lock to prevent it from opening during operation; c) check the correct placement of all the elements, particularly the sample holders in the reaction chambers; d) check that all equipments involved in the method during the execution of the process protocols are ready, including, at least, among said checks, the washing and purging process protocols of all reagent, air and any other solution dispensing probes used during the rest of said process protocols; e) optionally, read, by means of at least one high-precision image capture camera, all the labels of each sample holder, to identify the sample to be treated, the case from which it comes, the process protocol(s) to be executed, in the treatment of said sample; the order of priority of said process protocols; as well as carry out the detection of the tissue or cell smear existing on each sample holder and also carry out the delimitation of the position and amount of tissue or cells in each area of each sample holder; f) if all the initial checks have been passed, begin to execute one or more process protocols, by dispensing reagents in the samples for which an image capture of the identification codes of the respective specific reagent reservoirs and mixing tubes is carried out by Al if there is mixture in the series; g) calculate the time stamps and prioritise the process protocols to be executed, based on the previous readings and, based on the specific process protocol, determining the reagents and the amount of said reagents to be dispensed in each area of the sample holder; h) periodically update the timestamps and prioritisation of the process protocols of step g), until all processes of all protocols have been completed; and i) finish, when all the final maintenance tasks are concluded, then activating a low energy consumption routine. The method according to any of the preceding claims, wherein prior to its start, a list of tissue samples or cell smears to be processed, each of which identified with a sample code and the process protocol to be developed in those tissues or cell smears, as well as the corresponding code of the temporal priority of each sample and the personal data of the subject from whom it has been extracted, has been loaded into the database of the computer program comprised in the microprocessor. The method, according to any of the preceding claims, wherein prior to its start, a list of protocols has been loaded into the database of the computer program comprised in the microprocessor, each of which details the reagents and processes to be followed to perform a certain treatment on a type of tissue sample or cell smear. The method according to anyone of the preceding claims, wherein, during the performance of a treatment, the process protocols may be in any of the following states:

[0437] - READY: when a protocol is ready to begin its execution;

[0438] - EXEC: when a protocol is in the execution state; and

[0439] - EXEC_OK: when a protocol has completed its execution. The method according to any of the preceding claims, wherein, depending on the specific action they are carrying out, the processes of each protocol may in turn be in one of the following states: - Not ready: when a process is not ready for execution;

[0440] - READY: when a process is ready for execution;

[0441] - EXEC: when a process is in an execution state;

[0442] - EXEC_OK: when a process has finished its execution, but is waiting for the next process to start; and

[0443] - FINISHED: when a process has completed its execution and the next process has started its own.

[0444] 8. The method according to any of the preceding claims, wherein the robotic device (8) also comprises at least one reagent spreader roller (33).

[0445] 9. The method according to any of the preceding claims, comprising carrying out maintenance processes, based on changes in environmental conditions of temperature and humidity.

[0446] 10. The method according to any of the preceding claims, wherein the dispensing of reagents by a robotic device (8) is replaced by a manual dispensing.

[0447] 11 . The method according to any one of the preceding claims, wherein the probes and reagent spreader roller (33) are washed each time they are used during the performance of any process.

[0448] 12. The method according to any of the preceding claim, wherein the method further comprises a step of monitoring the temperature in the plurality of compartments acting as reaction chambers by means of an infrared (IR) sensor.

[0449] 13. The method according to any of the preceding claims, wherein the method further comprises a dynamic planning step comprising steps for adaptively programming tasks of the robotic device (robotic tasks) in a time interval, wherein the robotic device is coupled to a staining or hybridisation apparatus, wherein the robotic device treats slides coupled to the staining or hybridisation apparatus according to a treatment protocol with reagents in reagent or fluid reservoirs coupled to the staining or hybridisation apparatus, wherein the steps comprise:

[0450] - creating a task list of the robotic device comprising all robotic tasks that are ready for execution within the time interval;

[0451] - calculating a robotic task priority for each robotic task on the robotic task list;

[0452] - sorting the list of robotic tasks in descending order of priority of robotic tasks;

[0453] - adding robotic tasks by continuously prioritising;

[0454] - changing the order of the processes of the robotic device as it acts, and

[0455] - reporting the state of each of the processes of the robotic device. A system for performing treatments of histological staining, immunoenzymatic staining and hybridisation on tissue sections or cell smears comprising:

[0456] - a general-purpose microcontroller;

[0457] - means for storing data in one or more databases;

[0458] - a USB bus;

[0459] - a CAN bus;

[0460] - a main electronic card, connected to the USB bus configured to allow communication with the microcontroller, and on the other hand to the CAN bus configured to propagate and receive the information from the other cards, and control:

[0461] • motors of a robotic device (8), together with its encoders and end-of-stroke sensors, for the X and Y axes;

[0462] • reagent presence and level detection sensors;

[0463] • open / close detection sensors;

[0464] • indicator light to indicate the system state;

[0465] • cold table for reagent maintenance;

[0466] • interface (34) to control at least one precision probe (32);

[0467] - an electronic motion control board, connected to the can bus, configured to control the motors of the robotic device (8) based on the different axes XYZ (30) configured to control: motors of the robotic device (8), together with its encoders and end-of-stroke sensors; interface for at least one liquid level detection (LLD) sensor;

[0468] - a hydraulic control board that is connected to the CAN bus, configured to control hydraulic circuits of the instrument:

[0469] • peristaltic pumps with stepper motors;

[0470] • direct current pumps;

[0471] • direct current valves;

[0472] • air compressors;

[0473] • blowing system air flow rate sensor (36);

[0474] - a compartment control card connected to the CAN bus, configured to control the thermal actuators of reaction chambers disposed in said compartments, and which also controls:

[0475] • actuators for the control of resistive heaters;

[0476] • interfaces of the resistive temperature sensors;

[0477] • actuators to control light indicators (12) indicative of the state of each reaction chamber; and further comprising: i. a staining or hybridisation area (2), segmented into various compartments (9) adapted for the function of reaction chambers of the different treatments to be executed in them, in each of which one or more previously prepared samples are disposed on sample holders (14), optionally labelled with identification codes; ii. a robotic device (8), which has means to move freely along the three axes XYZ (30), adapted to be positioned at any point in the area (2) of the compartments (9), wherein the area (2) is located in the horizontal plane XY, and comprising the following elements:

[0478] - at least one high-precision image capture camera (31) adapted to capture images of the position of the sample holders and / or of the physical characteristics of the samples and, optionally, of the identification codes, disposed on the sample holders (14), as well as of the identification codes disposed on specific reagent reservoirs (22);

[0479] - at least one probe adapted for the dispensing of specific reagents and at least one probe adapted for the dispensing of common reagents; - at least one probe adapted for washing; and

[0480] - at least one general-purpose microcontroller with a computer program for the control of all the protocols, processes, members and equipment that form or operate the system.

[0481] 15. The system according to claim 14, wherein the probes comprise:

[0482] - at least one comb (35) with at least one common reagent dispensing probe (35a);

[0483] - at least two washing probes of a diameter greater than the dispensing probes, adapted for performing washes and connected directly to the common reagent reservoirs (5a); and

[0484] - at least one reagent spreader roller (33), adapted for spreading reagents dispensed onto the sample holders (14).

[0485] 16. The system according to claim 14 or 15, wherein each of said compartments (9) comprises a lower face that rests on a thermal blanket (15), on a vibration system (16) and on a waste tray (3) and by its upper face, bevelled, grooved or milled, the lower face of the sample holder (14) of size and shape adapted to that bevelling, grooving or milling is supported, keeping the perimeter of the upper face of the sample holder (14) in flight and isolated, preventing the reagent that is dispensed on it from spilling out of the area by surface tension; and wherein, between each bevel, groove or milling, grooves (13) are placed transversely for the expulsion of excess waste towards a waste tray (3).

[0486] 17. The system according to any one of claims 14 to 16, further comprising a dual drainage device which, through two pumps, connects the waste tray (3) of all the compartments with at least two waste tanks, one for hazardous waste and one for non-hazardous waste.

[0487] 18. The system according to any one of claims 14 to 17, wherein the dispensing probes are connected with at least one area (5) of common reagent reservoirs and at least one area of specific reagents (6).

[0488] 19. The system according to any one of claims 14 to 18, further comprising at least one air blow drying system (36).

[0489] 20. The system according to any one of claims 14 to 19, further comprising automatic washing devices (25) using washing solutions. The system according to any one of claims 15 to 20, wherein the common reagent dispensing comb (35) is formed by at least six dispensing probes (35a) and at least six common reagent tubes. The system according to any one of claims 14 to 21 , further comprising at least one washing station (25) of the probes of the system. The system according to any one of claims 14 to 22, comprising at least one washing (25) and drying station for the spreader roller (33) and the comb (35) of probes (35a). The system according to any one of claims 14 to 23, comprising at least one automatic reagent mixing area (27). The system according to any one of claims 14 to 24, comprising at least one gas removal filter (41). The system according to any one of claims 16 to 25, wherein the thermal blanket (15) is provided with a thermal probe for the control and maintenance of the temperatures of the different protocols carried out by the system. The system according to any one of claims 14 to 26, wherein each compartment has a light indicator (12) to identify the fault state or the state of the protocol in execution in said compartment. The system according to any one of claims 17 to 27, wherein the waste tray (3) has a level sensor. The system according to any one of claims 19 to 28, wherein the air blow drying system (36) integrates a compressor and a flow sensor. The system according to any one of claims 14 to 29, further comprising an automatic washing device (25) of the compartments (9), consisting of a tube connected to a pump disposed in a container containing a specific washing solution determined by the control device, based on the specific process protocol being developed in said compartment. The system according to any one of claims 18 to 30, wherein the specific reagent reservoir area comprises a thermostatted rack (23), the temperature of which is controlled by the programmable device that controls the system. The system according to any one of claims 18 to 31 , wherein the common reagent reservoir area (5) has a weight volume sensor for each reservoir (5a). The system according to any one of claims 18 to 32, wherein the specific reagent reservoir area has a capacitance volume sensor in the probe to detect the reagent level in each reservoir. The system according to any one of claims 14 to 33, wherein the system (1) comprises at least one infrared sensor (42) for monitoring the temperature in the plurality of compartments (9) adapted for the function of reaction chambers. The system according to any one of claims 14 to 34, wherein the system is configured to execute a dynamic planning method comprising steps for adaptively programming tasks of the robotic device (robotic tasks) in a time interval, wherein the robotic device is coupled to the staining or hybridisation system, wherein the robotic device treats slides coupled to the staining or hybridisation system according to a treatment protocol with reagents in reagent or fluid reservoirs coupled to the staining or hybridisation system, wherein the steps comprise:

[0490] - creating a task list of the robotic device comprising all robotic tasks that are ready for execution within the time interval;

[0491] - calculating a robotic task priority for each robotic task on the robotic task list;

[0492] - sorting the list of robotic tasks in descending order of priority of robotic tasks;

[0493] - adding robotic tasks by continuously prioritising;

[0494] - changing the order of the processes of the robotic device as it acts, and

[0495] - reporting the state of each of the processes of the robotic device. The system of any one of claims 14 to 35, adapted to perform the method of any one of claims 1 to 13. The system according to claim 14 wherein the system can be operated manually, without the intervention of a robotic device (8) adapted for capturing high-precision images and dispensing reagents. An apparatus (1) for the performance of histological staining, immunoenzymatic staining and hybridisation treatments on tissue sections or cell smears comprising: i. a staining or hybridisation area (2), covered by a lid (100), segmented into various compartments (9) adapted for the function of reaction chambers of the different treatments to be executed therein, in each of which one or more previously prepared sample...

Claims

CLAIMS1. An automatic staining device (1) for dispensing staining reagents over one or more biological samples, wherein one or more reagents are delivered through a controlled pressure-driven flow, and wherein the automatic staining device (1) comprises: a. a pressure-driven fluid displacement mechanism; b. a memory and a processor in communication with said memory, wherein the memory comprises at least one of: i. information of at least one protocol, said protocol comprising information related to at least one reagent dispensing step and at least one blowing step; ii. information related to one or more biological samples; c. one or more sample holder supports (42) suitable to support one or more sample holders (41); d. one or more reagent vials (51); e. an air blowing unit comprising an air blower (60) and, preferably, an air flow sensor; and f. a robotic arm (10) comprising at least one dispensing probe (80), wherein the dispensing probe (80) is fluidly connected to the pressure-driven fluid displacement mechanism, preferably wherein the air blowing unit is also comprised in the robotic arm (10); characterized in that the memory comprises instructions that, when executed by the processor, cause the processor, during a blowing step, to regulate the flow or speed of air in the air blowing unit, the time duration of air blowing, the robotic arm displacement or speed during the air blowing step, and / or the distance of the air blower (60) to the sample or sample holder (41), based on information stored in the memory regarding a dispensing step of the protocol, preferably a previous, current or following reagent dispensing step and / or information regarding the biological sample.

2. The automatic staining device (1) according to any one of the previous claims, wherein, during an air blowing step, the flow or speed of air in the air blowing unit, the time duration of air blowing, the robotic arm displacement or speed, and / or the distance of the air blower (60) to the sample or sample holder (41) is regulated based on a previous, current or following reagent dispensing step, preferably based on a following reagent dispensing step of the protocol.

3. The automatic staining device (1) according to any one of the previous claims, wherein the air blowing unit is configured to remove a reagent from the sample and / or sample holders by blowing air over said reagent; and / or blow air over the reagent only until an adequate moisture level is left on the sample and / or sample holder (41) to facilitate further reagent dispensing or reagent spreading over said sample and / or sample holder (41).

4. The automatic staining device (1) according to any one of the previous claims, wherein the air blower (60) has a shape and orientation adequate to evenly spread and / or remove the reagents over the sample and / or sample holder (41), preferably such that the air is blowed at an angle between 10 to 80 degrees with respect to the normal to the surface of the sample holder, more preferably between 25 and 65 degrees, even more preferably between 35 and 55.

5. The automatic staining device (1) according to any one of the previous claims, wherein the air blower (60) comprises one or more orifices (64) to blow the air, and: a. the longitudinal axis (62) of the air blower has a length (61) substantially the same as the width (45) of the sample holders (41) and / or sample holder supports (42), or has a difference in length with said width of ± 25%; and / or b. if the air blower comprises only one orifice, said orifice is elongated in the direction of the longitudinal axis (62) of the air blower (60), wherein the maximum length of said orifice is substantially the same as the width (45) of the sample holders (41) and / or sample holder supports (42), or has a difference in length with said width of ± 25%; and / or c. if the air blower comprises more than one orifice, said orifices are placed along the longitudinal axis (62) of the air blower (60), wherein the two more distanced orifices with respect to each other are separated such that at least part of one of the orifices is separated from at least part of the other orifice a distance substantially the same as the width (45) of the sample holders (41) and / or sample holder supports (42), or has a difference in length with said width of ± 25%.

6. The automatic staining device (1) according to any one of the preceding claims, wherein each blowing step is related to a blowing profile, each profile defining at least one parameter of the air flow selected from: air speed or flow rate, position or distance to the sample, robotic arm speed, robotic arm position, power applied by the pressure source, and travel path or segment of the sample holder, preferably wherein theprotocol comprises a plurality of dispensing steps related each to at least one reagent or fluid and optionally to a blowing profile, more preferably said blowing profile determined based on a reagent associated to a dispensing step, more preferably the previous, current or following dispensing step.

7. The automatic staining device (1) according to any one of the preceding claims, further comprising an adjustable orientation mechanism configured to vary the angle at which air is blown with respect to the sample holder (41).

8. The automatic staining device (1) according to any one of the preceding claims, further comprising a protective element for protecting at least one orifice (64) of the air blower (60), preferably said element blocking a path between a dispensing probe and the air blower orifice (64).

9. The automatic staining device (1) according to any one of the preceding claims, wherein the air blowing unit is configured to perform multiple passes of air blowing in a given blowing step, preferably each pass having at least one parameter that may differ from a previous pass.

10. The automatic staining device (1) according to any one of the preceding claims, further comprising a sensor arrangement for verifying air flow rate or pressure, wherein the processor is configured to adjust the air blowing flow or speed and / or the position or distance of the air blower (60) with respect to the sample holder (41) during a dispensing or blowing step.

11. The automatic staining device (1) according to any one of the preceding claims, wherein the memory further comprises instructions that, when executed by the processor, cause the processor to select a specific blowing step, a blowing profile or to regulate one or more blowing parameters, based on the viscosity of the reagent just dispensed, the type of sample holder (41), information related to a reagent dispensing step and / or on the characteristics of the biological sample.

12. The automatic staining device (1) according to any one of the preceding claims, wherein the air blower (60) is coupled to the robotic arm (10) such that the robotic arm (10) can move the air blower (60) from a parking position to a working position, preferably at different distances from the sample holder (41).

13. An automatic staining device (1) for dispensing staining reagents over one or more biological samples, wherein one or more reagents are delivered through a controlled pressure-driven flow, and wherein the automatic staining device (1) comprises: a. a pressure-driven fluid displacement mechanism;b. a memory and a processor in communication with said memory, the memory comprising information of at least one protocol, said protocol comprising information related to at least one reagent dispensing step and at least one spreading step; c. one or more sample holder supports (42) suitable to support one or more sample holders (41); d. one or more reagent vials (51); e. a robotic arm (10) comprising at least one dispensing probe (80), wherein the dispensing probe (80) is fluidly connected to the pressure-driven fluid displacement mechanism; characterized in that the automatic staining device (10) further comprises: f. a spreading unit (30), preferably comprised in the robotic arm (10), comprising a non-rotatable spreading element (31), wherein a longitudinal axis of said spreading element (31) is or can be placed parallel to the surface of the one or more sample holders (41) and / or the surface of the sample holder supports (42), and wherein the memory comprises instructions that, when executed by the processor, cause the device (1) to perform a spreading step based on the previous reagent dispensing step, more preferably based on the viscosity of the reagent previously dispensed.

14. The automatic staining device (1) according to claim 13, wherein the spreading element (31) is coupled to the robotic arm via a mechanism that allows guided displacement along the vertical axis (Z axis) over a limited distance.

15. The automatic staining device (1) according to claim 13 or 14, wherein the spreading element (31) alone or in combination with the supporting element (32), weights between 1 and 50 grams, preferably between 1 and 20 grams, even more preferably between 2 and 10 grams, even more preferably 5 ± 1 grams.

16. The automatic staining device (1) according to any of claims 13 to 15, wherein the spreading unit (30) further comprises a supporting element (32) attached to the spreading element (31), and a supporting piece (33) preferably attached to the robotic arm, wherein the spreading element (31) is supported by the supporting element (32), and the spreading element (31) can be displaced parallel to a longitudinal axis of the supporting element (32) and / or in the direction perpendicular to the longitudinal axis of the spreading element (31), between a first configuration wherein the supporting piece(33) supports the weight of at least the spreading element (31) , preferably supports the weight of the spreading element (31) and of the supporting element (32), and a second configuration wherein the spreading element (31) is not supported by the supporting piece (33), such as when is supported by the sample holder (41) and / or the sample holder support (42) during a spreading step.

17. The automatic staining device (1) according to any of claims 13 to 16, wherein the spreading unit (30) further comprises a supporting element (32) attached to the spreading element (31), and a supporting piece (33) preferably attached to the robotic arm, wherein the spreader supporting element (32) can slide or traverse the supporting piece (33) until certain point allowing to move the spreading unit (30) the length of the supporting piece (33) or less.

18. The automatic staining device (1) according to claim 16 or 17, wherein the supporting element (32) and / or the spreading unit (31) can tilt an angle <|), preferably between -10 and 10 degrees with respect to the supporting piece (33) or the normal direction to the surface of the sample holder, more preferably can tilt between -6 and 6 degrees..

19. The automatic staining device (1) according any one of claims 13 to 18, wherein the longitudinal length of the spreading element is substantially the same as the width (45) of the sample holders (41) and / or sample holder supports (42), or has a difference in length size with respect to said width of ± 50%, preferably ± 25%.

20. The automatic staining device (1) according to any one of claims 13 to 19, wherein the spreading element (31) is cylindrical or a prism, preferably cylindrical.21 . The automatic staining device (1) according to any one of claims 13 to 20, wherein the spreading element (31) comprises a distal part (311), a proximal part (312), and a central part (313), wherein a diameter (D1) of the distal (311) and proximal part (312) is the same for both distal (311) and proximal (312) parts, and is bigger than a diameter (D2) of the central part (313), wherein the diameter (D1) of the distal (311) and proximal(312) parts is between 0.02 and 2 mm longer than the diameter (D2) of the central part(313), preferably is between 0.06 and 0.6 mm longer, more preferably is between 0.1 and 0.3 mm longer.

22. The automatic staining device (1) according to any one of claims 13 to 21 , wherein the spreading element (31) is made of a material selected from: polymer, metal, ceramic, and / or hybrid composite.

23. The automatic staining device (1) according to any one of claims 13 to 22, wherein the spreading element (31) is configured to adapt its height (Z-axis) dynamically during spreading, thereby following the topography of the sample holder (41) or sample, and / or is configured to vary its inclination angle with respect to the Z-axis to optimize reagent spreading.

24. The automatic staining device (1) according to any one of claims 13 to 23, wherein the spreading element (31) is coupled to a force control mechanism configured to limit the pressure exerted on the sample holder (41) during spreading, thereby preserving the integrity of fragile substrates or sensitive samples.

25. The automatic staining device (1) according to any one of claims 13 to 24, wherein the spreading element (31) is configured to be raised from the sample holder (41) after a spreading step, and displaced to a washing station.

26. The automatic staining device (1) according to any one of claims 13 to 25, further comprising a washing receptacle (79) suitable to fit the length of the spreading element.

27. The automatic staining device (1) according to claim 26, wherein the washing receptacle (79) comprises a cover or protective element configured to prevent splashing of cleaning solution outside the washing area or over the spreading element (31) or a dispensing probe.

28. The automatic staining device (1) according to claim 26 or 27, wherein the washing receptacle (79) is configured to clean the spreading element (31) by at least one of: direct fluid dispensing, preferably from one or more dispensing orifices within the washing receptable (79), immersion of the spreading element (31) in a cleaning solution, or a combination thereof, preferably for a predetermined period of time based on the previous reagent dispensing steps.

29. The automatic staining device (1) according to any one of claims 13 to 28, further comprising a drying station (78) for the spreading element (31), said drying station comprising an absorbent surface suitable to remove residual cleaning solution from the spreading element (31) by contact, preferably wherein the spreading element (31) is displaced over the absorbent surface in one or more passes.

30. The automatic staining device (1) according to any one of the previous claims, further comprising at least one camera (20), and wherein the memory comprises further instructions that, when executed by the processor, cause the processor to:a. receive one or more images captured by the at least one camera (20), the images comprising at least one sample or sample holder (41); b. determine the presence, size, and / or position of the sample or sample holder (41) in the images using a trained machine learning model; and c. optionally, adapt at least one parameter of the blowing step and / or the spreading step based on the determined presence, size, and / or position of the sample or sample holder (41).

31. An automatic staining device (1) for dispensing staining reagents over one or more biological samples, comprising: a. a pressure-driven fluid displacement mechanism; b. a memory and a processor in communication with said memory; c. one or more sample holder supports (42) suitable to support one or more sample holders (41); d. one or more reagent vials (51); e. a robotic arm (10) comprising at least one dispensing probe (80), wherein the dispensing probe (80) is fluidly connected to the pressure-driven fluid displacement mechanism; f. one or more sensors selected from: accelerometers, gyroscopes, magnetometers, encoders, current sensors, temperature sensors, flow sensors, and capacitive sensors; characterized in that the memory comprises instructions that, when executed by the processor, cause the processor to: i. collect data from at least one or more of said sensors over a temporal window, thereby generating a data matrix of sensor values; ii. provide said data matrix as input to at least one embedded artificial intelligence model configured to perform at least one of:- classification of the activity of the robotic structure,- detection of anomalies in the robotic functionality,- detection of anomalies in the robotic structure,- detection of inclination or loss of horizontal stability,- determination of the quality of the support structure,- estimation of compressor or pump wear,detection of activation of hydraulic pumps, estimation of heater wear, detection of liquid level, wherein the output of the embedded artificial intelligence model is used to generate a diagnostic, warning, or control signal for the operation of the automatic staining device (1).

32. The automatic staining device (1) according to claim 31 , wherein the memory comprises a plurality of embedded artificial intelligence models, each configured to process a respective subset of sensor data and to output a respective diagnostic or classification result, wherein the processor is configured to execute said models in parallel or sequentially.

33. The automatic staining device (1) according to claim 31 or 32, wherein at least one embedded artificial intelligence model is a neural network trained for N-class classification of the activity of the robotic structure, using as input data from at least one of: accelerometer, gyroscope, magnetometer, encoder, and current sensors.

34. The automatic staining device (1) according to any one of claims 31 to 33, wherein at least one embedded artificial intelligence model is an anomaly detection model configured to detect abnormal movement or operation of the robotic structure, using as input data from at least one of: accelerometer, gyroscope, magnetometer, encoder, and current sensors.

35. The automatic staining device (1) according to any one of claims 31 to 34, wherein at least one embedded artificial intelligence model is configured to detect inclination or loss of horizontal stability of the device, using as input data from at least one of: accelerometer, gyroscope, magnetometer, and encoder sensors, and to output a warning if the inclination exceeds a predetermined threshold.

36. The automatic staining device (1) according to any one of claims 31 to 35, wherein at least one embedded artificial intelligence model is a regression model configured to estimate the quality index of the support structure of the device, based on inertial sensor data.

37. The automatic staining device (1) according to any one of claims 31 to 36, wherein at least one embedded artificial intelligence model is a regression model configured to estimate the wear of the compressor or hydraulic pumps, based on a comparison ofestimated and actual usage time, and / or sensor data, preferably sensor data comprises data from an accelerometer, a gyroscope, a magnetometer, and / or air flow sensor data.

38. The automatic staining device (1) according to any one of claims 31 to 37, wherein at least one embedded artificial intelligence model is a regression model configured to estimate the wear of the reaction chamber heaters, based on a comparison of applied electrical power and measured thermal output, using as input data from temperature sensors and power measurement data.

39. The automatic staining device (1) according to any one of claims 31 to 38, wherein at least one embedded artificial intelligence model is a classification model configured to detect the activation state of hydraulic pumps, using as input data from accelerometer, gyroscope, and magnetometer sensors.

40. The automatic staining device (1) according to any one of claims 31 to 39, wherein at least one embedded artificial intelligence model is a classification model configured to detect the liquid level in a reagent vial, using as input data from a capacitive sensor and a Z-axis encoder, and optionally further comprising as input the Z-axis speed, Z- axis position, liquid identification, and / or a basic liquid detection algorithm.

41. An automatic staining device (1) for dispensing staining reagents over one or more biological samples, comprising: a. a pressure-driven fluid displacement mechanism; b. a memory and a processor in communication with said memory; c. one or more sample holder supports (42) suitable to support one or more sample holders (41); d. one or more reagent vials (51); e. a robotic arm (10) comprising at least one dispensing probe (80), wherein the dispensing probe (80) is fluidly connected to the pressure-driven fluid displacement mechanism; and f. at least one inertial sensor selected from: accelerometer, gyroscope, and magnetometer; characterized in that the memory comprises instructions that, when executed by the processor, cause the processor to: i. collect data from the at least one inertial sensor during execution of a staining protocol;ii. provide said data to an embedded artificial intelligence model trained to detect loss of horizontal stability or excessive inclination of the device or its support table; iii. compare the output of the artificial intelligence model to a predetermined threshold; and iv. if the output indicates a loss of stability or inclination exceeding the threshold, generate a warning signal and / or interrupt the staining protocol.

42. The automatic staining device (1) according to claim 31 , 35 or 41 , wherein the warning signal is displayed on a user interface and / or transmitted to a remote monitoring system.

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