Automated method for cutting biological materials

Abstract

The present invention relates to an automated method for cutting biological material from a region of interest (30) within a tissue sample disposed on a planar substrate, using a cutting tool comprising an internal cavity and a scraping blade (125) disposed at an inlet of the internal cavity. The method comprises: ·identifying the boundaries of the region of interest; ·calculating a scraping path for the scraping blade based on the identified boundaries, such that the scraping blade engages all the material within the ROI and scrapes all the material within the ROI; and ·controlling the position of the cutting tool relative to the planar substrate such that the scraping blade follows the calculated scraping path, whereby the calculated scraping path includes one or more individual scraping operations in which the blade (125) is pressed against the planar substrate at a starting location (61, 62) within the identified boundaries and is moved forward through the ROI until it reaches a stopping location (71, 72) and is lifted from the planar substrate. The method of the present invention includes calculating the scraping path such that the stopping location of each individual scraping operation is positioned within the already scraped area within the identified boundaries.

Description

Technical Field

[0001] The present invention relates to an automated method of cutting biological material from a sample disposed on a planar substrate such as a slide glass using a cutting tool having a scraping blade and a tool body into which the scraped material is collected during cutting.

Background Art

[0002] Such a method, and an apparatus for performing the method, are known from WO 2020 / 054250. The position of the cutting tool relative to the sample is controlled such that the scraping blade selectively engages the material within an identified region of interest (ROI). The blade is brought into contact with the slide glass and pushed forward through the biological material within the ROI to scrape the material and collect it within the tool. Further examples are disclosed in WO 2022 / 063695, where the blade is disposed in an orifice and suction is applied during cutting, and the sample material separated by scraping is drawn into the orifice and the internal cavity of the tool and collected on the back side of a filter element spanning the internal cavity. The sample material is then transferred to a collection tube by disposing a tube airtight around the tool orifice and generating a pressure pulse that discharges the material into the tube.

[0003] To optimize the quality of subsequent analysis, it is important that only the material from the ROI is collected and that the material of the ROI is not left on the slide glass. The ROI material must be torn in order to be collected within the cavity of the tool. At the end of the scraping operation, the blade is lifted from the slide. Any scraped material that remains connected to the material on the slide where the blade was lifted can be pulled out of the cavity. The inventors have found that when using suction, it can be ensured that tearing occurs by applying a sufficiently large suction force during cutting. However, this has the drawback of increasing the adhesion of the collected material on the back side of the filter and making reliable discharge into the collection tube more difficult.

[0004] Therefore, there is still room for improvement regarding defining a scraping method that does not require a high suction force and overcomes the problem that the sample material remains on the slide.

Prior Art Documents

Patent Documents

[0005]

Patent Document 1

Patent Document 2

Summary of the Invention

Means for Solving the Problems

[0006] The present invention belongs to an automated method of cutting biological material from a region of interest (ROI) within a sample disposed on a planar substrate using a cutting tool having a scraping blade, the scraping blade being disposed at an opening of an internal cavity within the tool into which the scraped material is collected during cutting. The method comprises: · identifying the boundaries of the region of interest; and · calculating a scraping path for the scraping blade based on the identified boundaries, the step being such that the blade engages all the material within the ROI and scrapes all the material within the ROI; · controlling the position of the cutting tool relative to the planar substrate such that the scraping blade follows the calculated scraping path and includes.

[0007] The scraping path includes one or more individual scraping operations in which the blade is pressed against the planar substrate at a starting location within or on the identified boundaries and is moved forward through the ROI until it reaches a stopping location and is lifted from the planar substrate. According to the present invention, the scraping path is calculated such that the stopping location of each individual scraping operation is positioned within an already scraped area within the identified ROI boundary.

[0008] In one embodiment, the calculated scraping path includes at least one scraping operation in which the identified ROI boundary is scraped. In one example, this is the first scraping operation. The blade has a leading edge that is moved forward during cutting so as to scrape material from a planar substrate, such as a slide glass. The blade further has an inner edge and an outer edge that cut a scraping lane through the sample material. When scraping the ROI boundary, the position of the blade is controlled such that the outer edge of the blade is at the ROI boundary and is used to cut off material from the adjacent sample material. The blade follows the identified ROI boundary and generates a first scraping lane. The blade returns to the starting location and is only lifted after advancing beyond the starting location, such that the stopping location is positioned on the first scraping lane, i.e., the already scraped portion of the ROI boundary.

[0009] The cutting tool is preferably attached to the tool carrier of the cutting device that executes the method so as to rotate about a vertical axis of rotation perpendicular to the slide glass on which the sample is disposed. The vertical axis of rotation preferably coincides with the outer edge of the scraping blade. As will be appreciated, the apparatus further comprises an actuator that enables control of the movement of the blade in the x and y directions with respect to the planar substrate during scraping, and an actuator that enables lowering and lifting of the blade.

[0010] In an automated cutting method, accuracy is of utmost importance, but optimization of speed and efficiency is also desirable. The most efficient solution is considered to be lifting the blade just before it returns to the starting location, with the remaining distance being smaller than the width of the blade and a small un-scraped fragment remaining. The blade can then be rotated so that the leading edge of the blade faces the inner region of the ROI, and this can be scraped by starting a subsequent scraping operation at the un-scraped portion of the identified ROI boundary.

[0011] As described above, this poses a risk that the material will be drawn out of the internal cavity of the tool when the blade is lifted. Furthermore, the inventors have found that during scraping, for example, the leading edge of the scraping blade, which is thin at 0.03 - 0.1 mm but thicker than the typical tissue sample thickness of 0.003 - 0.01 mm, pushes the material forward. This material pushed forward may also be left behind if the blade is lifted before reaching the already scraped area, which further reduces the accuracy of the scraping process.

[0012] After the boundary has been scraped, perhaps after being scraped in a first scraping operation, the internal area of the ROI can then be scraped in several subsequent scraping operations. The blade is brought into contact with the slide glass at the starting location on the first scraping lane generated, and is moved inward until it reaches the stopping locations in different areas of the first scraping lane.

[0013] In the above example, the first scraping operation of the determined scraping path generates a first scraping lane where the outer contour coincides with the ROI boundary.

[0014] In an alternative embodiment that can be advantageously applied when the tissue sample has a high tearing strength and shows flaking during scraping, the first scraping operation starts in the internal area within the identified ROI boundary. Flaking may occur in paraffin-embedded samples where the tumor tissue has a high density and the paraffin embedding is locally incomplete. As a result, when the scraping blade is pushed forward through the sample material, the tissue sample may tear at relatively weak points without being precisely cut off from the adjacent material by the outer edge of the blade. Therefore, a scraping operation following the outer contour of the ROI boundary poses a risk of collecting unnecessary samples.

[0015] In an alternative embodiment, the starting location of the first scraping operation is within the ROI boundary. In one example, the blade is programmed to follow a circuit that completely surrounds the internal region within the ROI boundary. As before, the blade is lifted only after returning to the starting location and passing through it. In a further example, the first scraping operation is programmed such that after the blade has scraped the entire internal region of the ROI, it is lifted from the slide at a previously scraped portion of the internal region as described above.

[0016] In subsequent scraping operations, the starting location of the blade is on the ROI boundary, and the blade is oriented substantially parallel to the local contour of the boundary with the leading edge facing the interior of the ROI. The stopping location of each subsequent scraping operation is preferably on the first scraping lane or on a previously generated scraping lane.

[0017] In an embodiment where the first scraping operation defines a first scraping lane that winds around the region of the ROI, the stopping location is positioned on the first scraping lane after the circuit is completed and the blade has returned to the starting location and passed through it by a predetermined distance. The predetermined distance is based on the known width of the blade. In an example where the blade width is 1.0 mm, the predetermined distance is between 20 - 150% of the blade width and is preferably greater than the positioning tolerance of the blade of the device being used. In an example of a typical cutting device, the positioning tolerance is 0.1 mm.

[0018] In a further development, the scraping path includes a first scraping operation that starts within the ROI, and subsequent scraping operations of the scraping path are calculated only after it has been determined whether the tissue sample being cut shows flaking.

[0019] The device used to perform automated cutting according to the present invention typically comprises an imaging system used to obtain an image of the slide. This system comprises an imaging sensor having a position relative to a pre-calibrated scraping blade.

[0020] The method can advantageously further include the step of capturing an image of at least a portion of a first scraping lane generated during a first scraping operation, and the step of processing the captured image to determine whether the outer edge / boundary of the generated scraping lane portion coincides with a programmed path of the blade outer edge. It is preferable that the portion of the first scraping lane that functions as a "test lane" follows a straight line.

[0021] If it is determined that the boundary of the scraped portion described above is cleanly cut off from the adjacent material, i.e., the scraped lane does not contain sample material, then the scraping path is calculated to remove the remaining ROI sample material from the slide, whereby one of the subsequent scraping operations is programmed to scrape the outer contour of the identified ROI boundary as described for the first embodiment.

[0022] As a result of image processing, when the boundary of the scraped portion deviates from the programmed path of the blade outer edge, the scraping path calculated to remove the remaining ROI material includes subsequent scraping operations where the blade is positioned on the identified ROI boundary in a direction parallel to the local contour and moved inward as described above.

[0023] The step of processing the captured image can preferably include detecting the boundary of the scraped portion and comparing the detected boundary with the programmed path, and calculating the deviation distance if there is a deviation distance between them. The threshold value may be set based on, for example, an average absolute deviation of 0.1 mm and / or a standard deviation, whereby if the threshold value is exceeded, it is determined that there is a deviation from the programmed path.

[0024] The scraping path is calculated based on the identified ROI boundary. Typically, the boundary includes local contour portions that intersect at an angle, and the method includes estimating the lengths of the intersecting portions and the angles at which they intersect. Advantageously, the scraping path is calculated according to one or more additional rules that improve the accuracy, speed, and efficiency of the cutting.

[0025] The leading edge of the scraping blade has a maximum effective width when oriented in a direction perpendicular to the forward translation direction. This creates a scraping lane of the corresponding width as the blade moves forward through the ROI material. The effective width can be reduced by changing the angle at which the leading edge of the blade faces with respect to the translation direction.

[0026] An additional rule that can be applied when calculating the scraping path is that if the blade width does not exceed the local width of the ROI, the leading edge of the blade is oriented in a direction perpendicular to the translation direction. Thus, the step of calculating the scraping path can include estimating the local width of the ROI that is scraped in a particular scraping operation. When the local width is wider than the maximum blade width, the leading edge of the blade is oriented in a direction perpendicular to the translation direction. This maximizes the efficiency of a particular operation. When the estimated local width is narrower than the maximum blade width, the blade is rotated around the vertical axis to adjust the orientation of the leading edge of the blade with respect to the translation direction and reduce the effective width as needed.

[0027] An additional rule that can be applied is that the operation of the blade does not stop at the local contour of the identified boundary with the leading edge parallel to and facing the local contour. This prevents the ROI material from being pushed outside the boundary.

[0028] When the identified boundary includes a corner, i.e., when it includes first and second local contour portions that intersect at an angle of 95 - 100 degrees or less, the intersection angle is estimated. In an embodiment where the boundary of the ROI is scraped using the outer edge of the blade, a further rule that can be applied when calculating the associated scraping operation of the scraping path is that the blade is rotated backward around a vertical axis that coincides with the outer edge before reaching the corner. As described above, the default orientation angle at which the blade faces the first local contour portion of the ROI boundary is 90 degrees, i.e., the leading edge is perpendicular to the first local contour portion. It is preferable that the forward movement of the blade is stopped before the inner edge of the blade reaches the second local contour portion, and after adjusting the angle at which the blade faces the first local contour portion to be smaller than the estimated angle of the corner, the forward movement is continued.

[0029] Subsequent scraping operations of the calculated scraping path advantageously generate scraping lanes that are straight and parallel to each other. This helps to optimize the speed and efficiency of the cutting process. For greater accuracy, the scraping path may be determined such that adjacent scraping lines overlap each other by, for example, 5 - 20% of the lane width. As a further development, the method can include identifying the direction in which the region of interest is the maximum length, or the direction in which the local contour portion of the ROI boundary is the maximum length. The scraping path is then calculated such that scraping lanes are performed parallel to the identified maximum length direction.

[0030] The scraping blade of the cutting tool used in the method of the present invention has a straight leading edge having a width of, for example, 1.0 mm. Any of the cutting tools disclosed in International Publication No. 2022063695 can be used in the method of the present invention, and the content of this document is incorporated by reference. The disclosed tool includes a filter element across an internal cavity such that scraped ROI material sucked into the tool is captured on the back surface of the filter element. The scraped material is hermetically disposed around the distal end of the cutting tool in a tube and transferred to a collection tube by generating a pressure pulse that discharges the material.

[0031] It is preferred that the transfer be performed after all of the ROI material has been scraped from the slide. Depending on the size of the ROI, the filter may become somewhat clogged before all of the material is collected, which may reduce the effectiveness of the suction during cutting. For certain tools with filters, a threshold value can be defined corresponding to the surface area of the material to be scraped that poses a risk of clogging. The surface area to be scraped is determined by the known width of the blade and the length advanced during scraping. For the length advanced by the blade, a corresponding threshold value may be determined. When the threshold value is reached, it is beneficial to interrupt the scraping and perform an intermediate transfer action.

[0032] Accordingly, the method can further include calculating the length advanced by the blade during scraping and, after completion of the scraping operation, interrupting the scraping path when the calculated length reaches a predetermined threshold value. The tool is lifted only after the blade has entered the already scraped area and can then, for example, move to a collection stage where the scraped material is transferred. The tool is then returned to the slide and the remainder of the calculated scraping path is executed.

[0033] Those skilled in the art will understand that any two or more of the above-described embodiments, implementations, and / or aspects of the present invention may be combined in any useful way.

[0034] Next, the present invention will be further described with reference to the embodiments described below.

Brief Description of the Drawings

[0035]

Figure 1

Figure 2a

Figure 2b

Figure 2c

Figure 3a

Figure 3b

Figure 4a

Figure 4b

Figure 4c

Figure 5

Figure 6

Best Mode for Carrying Out the Invention

[0036] It should be noted that items having the same reference numerals in different figures have the same structural features and the same functions, or are the same signals. If the functions and / or structures of such items are described, there is no need to repeat those descriptions in the detailed description.

[0037] Pathological diagnosis research of biological materials such as tissues and cells forms the basis for many treatment decisions, especially in oncology. For example, genomic-based tests are performed to inform treatment options for individual patients diagnosed with cancer. Biological materials / tissues can be obtained from biopsies and then, for example, embedded in paraffin, cut into thin slices, and fixed onto glass slides. These thin slices are called tissue samples. Other methods of obtaining and preparing biological materials are known. An example of a tissue sample disposed on a glass slide 10 is shown in FIG. 1.

[0038] The tissue sample 20 has a region of interest (ROI) 30 that includes the material within the boundary 35 of the ROI to be diagnostically examined. This material must be physically separated from the slide and the unwanted sample material 25 outside the boundary. The ROI and associated boundary 35 can be identified by staining or, alternatively, a pathologist may mark it on a reference slide after microscopic analysis. The ROI can also be identified via processing of the digital image of the sample. Once the ROI is identified, the material is removed / cut from the slide and then transferred to an analysis construct. Typically, the material is transferred to a collection tube and then steps of a sample preparation process such as cell lysis, purification, and amplification, as well as further necessary processing steps, are performed. As understood, the reliability and accuracy of the analysis are optimized by ensuring that only the material cut from the ROI is present, but also by maximizing the amount of ROI material collected.

[0039] The present invention defines an automated method for cutting biological material from a region of interest (ROI) within a sample disposed on a planar substrate such as a slide glass, using a tool having a scraping blade that is pressed against the slide and moved forward through the sample to scrape the ROI material and collect it within a cavity in the tool. In a first step, the boundaries of the ROI are identified. Then, based on the identified boundaries, a scraping path for the blade is calculated. The position of the tool is controlled relative to the slide such that the scraping blade follows the calculated scraping path. In the method of the present invention, the scraping path is calculated according to one or more rules designed to optimize the accuracy of the cutting by ensuring that material from the region of interest is not left behind on the slide glass.

[0040] Figure 2a shows a schematic arrangement of a configuration for performing the method of the present invention.

[0041] The apparatus 100 comprises a platform 110 for supporting a slide glass 10 on which a tissue sample such as the sample 20 shown in FIG. 1 is disposed. The apparatus comprises a cutting tool 120 having a scraping head / scraping blade 125. The cutting tool 120 is preferably fixedly connected to a robotic stage 130 comprising a series of actuators for performing the movements necessary during cutting. The robotic stage 130 preferably comprises: - a rotational actuator for rotating the tool about a vertical axis R perpendicular to the platform 110, - an X-Y stage for translational movement, and - a Z stage for vertical movement. The Z stage can be provided with hinge bearings for position control to ensure that a constant and accurate downward force is applied during cutting.

[0042] As will be appreciated, it is also possible for one or more of the necessary actuators to be connected to the platform 110.

[0043] The device further comprises an imaging system 150 that can be used to identify the boundaries of the ROI. This imaging system includes an imaging sensor, and the position of the scraping blade relative to the sensor is pre-calibrated. The device further comprises a processor for calculating an appropriate scraping path based on the identified boundaries, and a controller 140 that receives the calculated scraping path and controls the robotic stage accordingly. As a result, the scraping blade of the cutting tool is moved relative to the platform to scrape all the material within the identified ROI. Preferably, the device further comprises a vacuum generator 160 that generates an uplifting air flow at the scraping blade 125 so that the scraped material is sucked into the internal cavity of the tool.

[0044] A cross-sectional side view of the tool is shown in FIG. 2b. The tool 120 in the illustrated example comprises a thin-walled tube having an internal cavity 128 and a scraping head at the entrance to the internal cavity. The front face or leading edge of the tube functions as the scraping blade 125 of the scraping head. The scraping blade has a bottom 125a that is brought into contact with the upper surface of the slide 10. The leading edge of the scraping blade is also in contact with the tissue sample 20 in a scraping zone formed by the bottom 125a and the opposing side portions 125b, 125c of the tube in the contact area. Thus, when relative movement occurs in the X direction, the material from the tissue sample is scraped from the slide 10 into the cavity 128 of the cutting tool. In the scraping zone, portion 125b defines the outer edge of the scraping blade; portion 125c defines the inner edge of the scraping blade, whereby the leading edge acts as if it were only as a whole. The scraping lane cut into the tissue sample 20 has a width w corresponding to the effective width of the scraping blade between the outer edge 125b and the inner edge 125c in the scraping area.

[0045] A further example of a cutting tool that can be attached to a cutting device for practicing the method of the present invention is shown in FIG. 2c. The tool 220 includes a body portion 226 formed from a single piece in which a tool internal cavity extends. A portion of the internal cavity 228 tapers in diameter towards the tool orifice 223. A further portion of the internal cavity is formed by a conical recess 450 that is adapted to be precisely aligned with a correspondingly shaped conical protrusion of the tool carrier of the cutting device to connect the tool.

[0046] The tool body portion 226 further includes a seating portion 230 for axially installing a filter element 229. The scraping blade 225 of the tool is provided on a second portion 227 joined to the body portion 226 and extends obliquely with respect to the longitudinal central axis of the tool internal cavity. In the illustrated example, the second portion 427 may be overmolded onto the scraping blade 225 and irreversibly joined to the body portion 226 by a form fit, adhesive bonding, or other suitable joining method. In other examples, the entire tool body is overmolded onto the scraping blade.

[0047] The scraping path calculated for a particular scraping blade and a particular ROI includes several individual scraping operations in which the scraping blade is moved relative to the slide until it contacts the slide at the starting location, reaches the stopping location, and is lifted from the slide. As shown in FIG. 3a, when the scraping blade 125 is moved forward through the ROI material, the material can be separated in the form of a ribbon 30a that is guided or sucked into the tool cavity during cutting. When the leading edge of the ribbon 30a remains attached to the adjacent sample material and the blade is lifted, as shown in FIG. 3b, the ribbon of material can be pulled out of the cavity. The front surface / leading edge of the scraping blade also pushes the sample material forward during cutting, which creates an accumulation 30b or "mount" of ROI material in front of the blade. This material may be left behind if the accumulation in front of the blade is not considered in the scraping path.

[0048] According to the present invention, the scraping paths are calculated such that each individual scraping operation ends at a previously scraped position within the identified ROI boundary.

[0049] In a preferred embodiment, the scraping path starts from a first scraping operation in which the identified ROI boundary 35 is scraped. Referring to FIG. 4a showing the same tissue sample as depicted in FIG. 1, the scraping blade 125 is brought into contact with the slide at the starting location 61 of the identified boundary. The boundary typically includes intersecting local contour portions, and one possible way to select the starting location is to identify the longest portion among the local contour portions and start at those locations. The blade is oriented such that its leading edge is substantially perpendicular to the boundary at that location, and further positioned such that its outer edge coincides with the outer contour of the boundary. The blade is then moved forward in a first translational movement indicated by arrow a. The blade then follows the boundary in several subsequent translational movements, whereby, as much as possible, the leading edge is kept perpendicular to the local contour of the boundary and the outer edge is used to cut the ROI material from the unwanted sample material 25. When passing through a corner with an angle of 95 degrees to 100 degrees or less, typically a change in orientation is required, which is explained with reference to FIGS. 4b and 4c.

[0050] In FIG. 4a, the blade 125 is shown at various locations during the subsequent translations when a first scraping lane 51 following the boundary is generated. In the final translational movement, as indicated by arrow q, the blade returns to the starting location 61 and moves beyond it, for example, by 50% of the blade width, and is lifted from the slide at the stopping location 71, which is the already scraped area.

[0051] FIGS. 4b and 4c show an example of an ROI boundary having local contour portions 35a, 35b that intersect at an angle less than 90 degrees and thus generate a corner 38. Further, the blade 125 is shown at several different positions 1 - 7 when passing through the corner 38 during the first scraping operation.

[0052] The step of identifying the boundary 35 of the ROI preferably includes estimating the angle of any corner within the boundary. In the illustrated example, corner 38 has an angle of 70 degrees. As it approaches the corner and moves forward between position 1 and position 2, blade 125 is oriented perpendicular to the local contour 35a, and the outer edge 125b of the blade coincides with the outer edge of the local contour. This forward movement is preferably stopped at position 2 before the inner edge 125c of the blade meets the local contour 35b. This prevents the accumulated ROI material (such as that schematically shown in Fig. 3b) from being extruded outside the boundary. The blade is then rotated backward, for example, by an angle of 30 degrees to position 3 around a vertical axis of rotation R (see Fig. 2) that coincides with the outer edge 125b of the blade. At position 3, the blade has an orientation angle θ with respect to the local contour portion 35a, which is smaller than the estimated angle of the corner. The blade is then moved forward to position 4, where the outer edge is approximately located at the intersection of the contour portions 35a and 35b. As shown in Fig. 4b, the blade is then rotated backward again to position 5 and then adjusted in angle and moved forward to position 6, where the blade is perpendicular to the local contour 35b and the outer edge 125b of the blade coincides with the local contour 35b. The blade is then linearly moved forward to position 7.

[0053] During individual scraping operations, the blade remains in contact with the slide. Optionally, while moving continuously or stepwise and while stopping the linear movement, the blade can be adjusted in angle around the vertical axis of rotation.

[0054] After a first scraping lane 51 with an outer contour that coincides with the ROI boundary is generated, the interior of the ROI 30 can then be scraped in several subsequent scraping operations. Referring again to FIG. 4a, a second scraping lane 52 can be generated by bringing the blade into contact with the slide 10 at a second starting location 62 on the first scraping lane 51 and moving the blade forward to a second stopping location 72 on the opposite side of the first scraping lane. Subsequent scraping operations are preferably parallel to each other and can overlap each other by, for example, 10-20% of the blade width. The number of additional scraping operations depends on the width of the blade and the size of the area to be scraped.

[0055] The most efficient cutting method is to move the blade forward with the leading edge perpendicular to the translational direction so that the width of the blade is maximized. This may not always be possible, for example, when passing through a corner. There may also be a need to change the orientation of the blade to cope with the local width of the ROI.

[0056] FIG. 5 shows a further example of an ROI 530 with a boundary 535. A portion of the ROI 530a has a local width s that is narrower than the maximum effective width w of the scraping blade 125. The blade is also shown at several different positions 1-7 during the scraping operation. When the blade moves from position 1 to 2, the local width of the ROI is wider than w, and the blade is oriented such that the leading edge is perpendicular to the translational direction. As the blade moves forward to position 3, it is rotated backward about a vertical axis of rotation that coincides with the outer edge 125b of the blade, and as the blade enters the narrow portion 535a, the orientation angle is further adjusted to reduce the effective width of the blade and it moves forward to position 4. As shown, as the blade moves further to positions 5, 6, and 7, the angle of orientation is adjusted based on the local width of the ROI.

[0057] Generally, the most efficient way to scrape a particular ROI is, as described above with respect to FIG. 4a, to start from the boundary of the ROI and then scrape the internal region in a number of subsequent scraping operations. Depending on the type of tissue sample, for example, in the case of a tissue sample with high tearing strength as described above, if the outer edge of the blade is used to cut off the ROI from the adjacent sample material at the ROI boundary, there is a risk of flaking. In this case, it is advisable to use a different type of scraping path that includes placing the blade at the ROI boundary, aligning the leading edge substantially parallel to the local contour, and making a number of scraping movements that move forward towards the internal region of the ROI. If the length of the local contour portion is shorter than the width of the blade, the blade is not moved forward but is only pressed against that portion so as to be cut off from the adjacent unwanted material. The cut-off portion can then be scraped in a subsequent scraping operation where the outer edge of the blade follows the local contour.

[0058] According to the present invention, each scraping operation ends after the blade has reached the area that has already been scraped, i.e., the blade is lifted from the slide glass. The first scraping operation of the scraping path is then programmed to start from the internal region of the ROI.

[0059] Examples of possible first and second scraping lanes are shown in FIG. 6. The blade is placed on slide 10 at starting location 661 inside the identified ROI boundary 635 and is moved forward to create a continuous first scraping lane 651 that completely surrounds the area 630a of ROI material within the boundary. For simplicity, a substantially rectangular scraping lane 651 is depicted, but other enclosed shapes are possible. The blade returns to the first starting location 661, is moved in a "counterclockwise" direction until it passes through there, and is then lifted at stopping location 671. In the second scraping operation, the blade is placed on the ROI boundary 635 at the second starting location 662 and is moved forward to create a second scraping lane 652. The second scraping operation ends at a stopping location 672 that is part of the first scraping lane 681. In subsequent scraping operations, the blade is returned to a new starting location on the ROI boundary and is moved inward until it reaches the previously created scraping lane. The process stops when all of the ROI material has been collected.

[0060] Examples, embodiments, or optional features, whether shown as non-limiting or not, should not be understood as limiting the claimed invention. The above-described embodiments are illustrative rather than limiting, and it should be noted that those skilled in the art can design many alternative embodiments without departing from the scope of the appended claims.

[0061] In the claims, any reference signs placed between parentheses shall not be construed as limiting the claim. The use of the verb "comprise" and its conjugations does not exclude the presence of elements or steps other than those recited in the claim. The article "a" or "an" preceding an element does not exclude the presence of a plurality of such elements. The present invention can be implemented by hardware comprising several distinct elements and by a suitably programmed computer. In device claims enumerating several means, several of these means may be embodied by one and the same item of hardware. The mere fact that certain measures are recited in mutually different dependent claims does not indicate that a combination of these measures cannot be used to advantage.

Explanation of Signs

[0062] 1 - 7 Positions of the scraping blade at different instants within the scraping operation 10 Slide glass 20 Tissue sample 25 Unwanted tissue material 30 Region of interest (ROI) 30a Ribbon - shaped tissue material cut by the scraping operation 30b Accumulation of tissue material pushed forward by the scraping operation 35, 535, 635 Boundaries of the ROI 35a, 35b Intersecting parts of the ROI boundaries 38 Corners within the ROI boundary (parts intersecting at an angle of 90 degrees or less) 51, 651 First scraping lane 52, 652 Second scraping lane 61, 661 Starting points of the first scraping lane 62, 662 Starting points of the second scraping lane 71, 671 Ending points of the first scraping lane 72, 672 Ending points of the second scraping lane 100 Cutting device 110 Platform for supporting a slide glass 120, 220 Cutting tool 125, 225 Scraping blade of the cutting tool 125a Bottom of the tubular scraping blade 125b Outer edge of the scraping blade 125c Inner edge of the scraping blade 128, 228 Internal cavity of the cutting tool 130 Robot stage for adjusting the position of the blade relative to the slide glass 140 Controller 150 Imaging system 160 Vacuum generator 223 Orifice of the cutting tool 226 Body part of the cutting tool 427 Second tool body part 229 Filter element 230 Seating part for holding the filter element 530, 630a ROI part a First translational motion of the scraping operation q Final translational motion of the scraping operation R Vertical axis of rotation of the cutting tool s Local width of a part of the ROI w Width of the scraping blade / scraping lane θ Orientation angle of the blade with respect to the local contour part

Claims

1. An automated method for cutting biological material from a region of interest (30, 530, 630) within a tissue sample (20) disposed on a planar substrate (10) using a cutting tool (120, 220) comprising internal cavities (128, 228) and scraping blades (125) disposed at the entrances of the internal cavities, the method comprising: - identifying the boundaries (35, 535, 635) of the region of interest; - calculating a scraping path for the scraping blade based on the identified boundaries, the scraping path being such that the scraping blade engages all the material within the region of interest and scrapes all the material within the region of interest; - controlling the position of the cutting tool relative to the planar substrate such that the scraping blade follows the calculated scraping path; whereby the calculated scraping path includes one or more individual scraping operations in which the blade (125) is pressed against the planar substrate (10) at a starting location (61, 62, 661, 662) within the identified boundaries and is moved forward through the ROI until it reaches a stopping location (71, 72, 671, 672) and is lifted from the planar substrate; the method being characterized in that the scraping path is calculated such that the stopping location of each individual scraping operation is positioned within the already scraped area within the identified boundaries.

2. The method according to claim 1, wherein the cutting tool is mounted to rotate about a vertical axis of rotation (R) that coincides with the outer edge (125b) of the scraping blade.

3. The method according to claim 2, wherein the calculated scraping path includes a scraping operation in which a scraping lane (51, 651) is generated that completely surrounds at least a portion of the region of interest, whereby the position of the blade (125) is controlled to return the blade to the starting location (61, 661) and follow a circuit that continues beyond the starting location such that the stopping location (71, 671) is positioned within the already scraped portion of the generated scraping lane.

4. The method according to claim 3, wherein the stopping location is separated from the starting location by 20 - 150% of the width of the scraping blade, this distance being greater than the positioning tolerance of the cutting device to which the cutting tool is mounted.

5. - the outer contour of the generated scraping lane (51) coincides with the identified boundary (35); The method according to claim 3 or 4, wherein the position of the scraping blade (125, 225) is controlled during the scraping operation such that the outer edge (125b) of the scraping blade follows the identified boundary (35) to cut the material at the boundary from the adjacent material (25) of the sample (20).

6. The step of identifying the boundary includes identifying local contour portions (35a, 35b) of the boundary that intersect at a corner (38) having an angle of 100 degrees or less, and estimating the intersection angle. Before reaching the corner (38), the position of the scraping blade (125, 225) is controlled during the scraping operation such that the leading edge of the scraping blade (125, 225) is rotated backward about the vertical axis of rotation (R) so as to be oriented at an angle (θ) smaller than the estimated intersection angle. The method according to claim 5.

7. The scraping operation in which the outer edge of the blade follows the identified boundary (35, 535) is the first scraping operation, and the interior of the region of interest (30) is scraped in several subsequent scraping operations. The method according to claim 5 or 6.

8. The method according to any one of claims 1 to 3, wherein the first scraping operation and the associated first scraping lane (651) are inside the identified boundary (635).

9. The calculated scraping path includes several subsequent scraping operations having a starting point (662) on the identified boundary. The leading edge of the blade (125, 225) is oriented parallel to the local contour of the boundary (635) at the corresponding starting point, whereby the blade moves forward into the region of interest until it reaches a stopping point (672) on the first scraping lane (651) or on a scraping lane generated previously. The method according to claim 8.

10. The method according to claim 7 or 9, wherein the subsequent scraping operations generate scraping lanes (52, 652) parallel to each other.

11. The method according to claim 7, 9, or 10, wherein adjacent scraping lanes overlap each other by an amount corresponding to 5 - 20% of the width (w) of the scraping blade (125).

12. Identifying the direction in which the region of interest has its maximum length, or the direction in which the local contour portion of the ROI boundary (35) has its maximum length, and calculating the scraping path such that scraping lanes parallel to the identified direction of maximum length are executed The method according to claim 10 or 11, further comprising **Claim 13** The method according to any one of claims 1 to 12, wherein the step of calculating the scraping path further comprises estimating the local width (s) of the region of interest to be scraped in a specific scraping operation, and if the width (w) of the blade is not smaller than the estimated local width, further comprising orienting the leading edge of the scraping blade (125, 225) perpendicular to the translational direction. **Claim 14** The cutting tool is attached to a device comprising an imaging system (150) and a processor, and the method further comprises the step of capturing an image of at least a part of the first scraping lane (651), and the step of processing the captured image, and the processing step comprises - Detecting the boundary of the first scraping lane portion; - Comparing the detected boundary with the programmed path of the corresponding edge (125a, 125b) of the scraping blade (125, 225); - Determining whether the detected boundary deviates from the programmed path, and if it is determined that a deviation has occurred, - the calculated scraping path includes several subsequent scraping operations as defined in claim 9, and if it is not determined that a deviation has occurred, - the calculated scraping path includes subsequent scraping operations as defined in claim 5, The method according to claim 8, further comprising **Claim 15** - The cutting tool (220) comprises a filter element (229) across the internal cavity (228) of the scraping tool; - A vacuum generator (160) for generating an upward airflow that sucks the scraped material into the tool and holds it on the back surface of the filter element (229) is connected to the inlet (223) of the tool cavity; - The method includes the step of calculating the distance the scraping blade (225) has advanced during scraping; - After completion of the scraping operation, if the calculated distance exceeds a predetermined threshold, interrupting the scraping path, the threshold corresponding to the maximum surface area of the sample material that can be held on the back surface of the filter element (229) without adversely affecting the suction performance, and restarting the scraping path after the scraped material has been transferred to the collection tube. The method according to any one of claims 1 to 14, further comprising **Claim 16** A cutting tool having internal cavities (128, 228) and scraping blades (125) disposed at the entrances of the internal cavities, A platform (110) for supporting a planar substrate (10) on which a tissue sample (20) is disposed, An imaging system (150) for identifying boundaries (35, 535, 636) of an area of interest (30) within the tissue sample, A processor for calculating a scraping path that will engage the scraping blade (125, 225) with all sample material within the identified boundaries, A series of actuators (130) for moving the cutting tool in the X, Y, and Z directions relative to the platform (110) and for rotating the tool about a vertical axis of rotation (R), A controller (140) for receiving the calculated scraping path and controlling the series of actuators (130) so that the scraping blade follows the calculated scraping path Comprising, A cutting device (100), characterized in that a processor is configured to calculate a scraping path such that automated cutting is performed according to the method of any one of claims 1 to 14.

17. Further comprising a vacuum generator (160) for generating an upward airflow at an entrance (223) to an internal cavity (228) of the tool, the tool further comprising a filter element (129) extending across the internal cavity, the apparatus being configured to perform the method of claim 15, the cutting device (100) according to claim 16.

Images