Sectioning and Quality Control in the Microtome Method

Abstract

[0003] The present disclosure relates to systems and methods for sectioning and quality control determination in microtomy. In some embodiments, a histology system is provided that includes a microtome configured to progressively remove one or more sections from a tissue block, the tissue block comprising a tissue sample embedded in an embedding material, a vision system associated with the microtome, the illumination system configured to illuminate the tissue block comprising the tissue sample embedded in the embedding material, an imaging system configured to image the tissue block and generate imaging data associated with the tissue block, and a processor in communication with the vision system, the processor programmed to receive the imaging data and perform sectioning and quality control analysis.

Description

Technical Field

[0001] (Cross - Reference to Related Applications) This application claims the benefit and priority of U.S. Provisional Application No. 62 / 980,201, filed Feb. 22, 2020; U.S. Provisional Application No. 62 / 980,203, filed Feb. 22, 2020; U.S. Provisional Application No. 62 / 980,202, filed Feb. 22, 2020; U.S. Provisional Application No. 62 / 980,194, filed Feb. 22, 2020; and U.S. Provisional Application No. 63 / 134,399, filed Jan. 6, 2021; U.S. Utility Application No. 17 / 182,139, filed Feb. 22, 2021; U.S. Utility Application No. 17 / 182,133, filed Feb. 22, 2021; and U.S. Utility Application No. 17 / 182,166, filed Feb. 22, 2021, all of which are hereby incorporated by reference in their entirety herein.

[0002] The present invention relates to quality control of cut tissue sections transferred from a biological tissue sample block to a slide, including sectioning, tracking, and mechanical quality control. In particular, this can be achieved using an automated system.

Background Art

[0003] The generation of postage stamp-sized micron-thin tissue sections for microscopic visualization, which is a conventional microtome method, is a delicate and time-consuming manual task. In the process, the microtome cuts a tissue block consisting of tissue samples, which is encapsulated within a support block of an embedding material such as paraffin wax. The microtome holds a blade that is aligned to cut slices from one face of the tissue block, i.e., the block cutting face. A common type of rotary microtome linearly oscillates a chuck that holds the block with a cutting face along the blade cutting face. In combination with an incremental advancement of the block cutting face to the cutting face, the microtome continuously shaves thin tissue sections from the block cutting face. For sections with a paraffin wax embedding medium, the operator carefully picks up these tissue sections and floats them on warm water. The water gently removes wrinkles and reduces deformation from cutting. Finally, the operator moves the sections from the water onto a microscope slide for further processing.

[0004] Recent advancements in digital imaging of tissue sample sections have made it desirable to slice the sample block very rapidly. As an example, when tissue is sectioned as part of a clinical procedure, time is an important variable in improving patient care. For instance, when examining the margins of a lung cancer to determine if sufficient tissue has been removed, all the time that can be saved during the sectioning of tissue for intraoperative use in anatomic pathology is clinically valuable. To create a large number of sample sections rapidly, it is desirable to automate the process of cutting tissue sections from a sample block with a microtome blade and facilitate the transfer of the cut tissue sections to an adhesive tape or other transfer medium without reducing the section quality. Additionally, a large number of tissue sample sections cut from the block need to be transferred to a microscope slide for evaluation.

[0005] Quality control regarding tissue samples is important. Inadequate quality control can have an adverse effect on pathology and lead to an inaccurate assessment of the tissue. However, currently, quality control of tissue sections deposited on a glass slide is a resource-consuming task.

[0006] This comparison is done manually for tissue transfer to slides, however, in many cases, it is difficult to distinguish tissue from paraffin, leading to inaccurate comparisons and thus improper quality control. Further, the inability to properly assess the tissue results in a lack of knowledge that insufficient tissue sections are placed on the slide. This also hinders the assessment of whether the tissue sections on the slide are damaged.

[0007] The present disclosure overcomes current workflow problems and deficiencies by implementing methods and systems that eliminate or at least significantly reduce quality control issues and the risk of discrepancies between slide labeling and tissue sections. SUMMARY OF THE INVENTION MEANS FOR SOLVING THE PROBLEM

[0008] The present disclosure relates to systems and methods for sectioning tissue blocks. In some embodiments, a method for sectioning a tissue block includes imaging the tissue block to generate imaging data of the tissue block, the tissue block comprising a tissue sample embedded within an embedding material; estimating a depth profile of the tissue block based on the imaging data, the depth profile comprising the thickness of the embedding material to be removed to expose the tissue sample to a predetermined reference; and removing the thickness of the embedding material to expose the tissue to the predetermined reference.

[0009] In some embodiments, the method may further include progressively removing one or more sections from a tissue block comprising a tissue sample embedded within an embedding material, imaging the one or more sections, generating imaging data associated with the one or more sections, and based on the imaging data, verifying that the tissue sample is exposed to a predetermined criterion. In some embodiments, the tissue block is imaged using structured light to determine a depth profile.

[0010] In some embodiments, a method for sectioning a tissue block is provided, the method including progressively removing one or more sections from a tissue block comprising a tissue sample embedded within an embedding material, imaging the one or more sections removed from the tissue block, generating imaging data associated with the one or more sections, and based on the imaging data, determining when a sufficient number of one or more sections have been removed from the tissue block to expose the tissue sample to a predetermined criterion.

[0011] In some embodiments, the method may further include imaging the tissue block and generating baseline imaging data of the tissue sample prior to removing the one or more sections. In some embodiments, the method may further include determining an expected contour, size, or shape of the tissue sample from the baseline imaging data. In some embodiments, the method may further include determining a depth profile of the embedding material from the baseline imaging data and illuminating the tissue block with structured light within the UV range to remove a sufficient amount of the embedding material to expose the tissue sample to a predetermined criterion.

[0012] In some embodiments, the method can further include comparing imaging data of a section including the tissue sample to baseline imaging data to determine when the tissue sample is sufficiently exposed. In some embodiments, the imaging data of the section comprises imaging data of one or more sections on a tissue block, on a transfer medium, or on a slide. In some embodiments, the contour, size, or shape of the tissue sample in one or more sections is compared to the contour, size, or shape of the tissue sample expected from the baseline imaging data. In some embodiments, the method can further include determining a depth profile by one or more of parallax, focus, or light field imaging, and increasing the contrast between the tissue sample and the embedding material.

[0013] In some embodiments, there is provided a method for sectioning a tissue block, including removing a thickness of an embedding material configured to expose a tissue sample within the embedding material to a predetermined criterion, following the step of removing the thickness, progressively removing one or more sections from the tissue block, imaging the one or more sections removed from the tissue block and generating imaging data associated with the one or more sections, and confirming from the imaging data that the tissue sample is exposed to the predetermined criterion.

[0014] In some embodiments, a microtome configured to progressively remove one or more sections from a tissue block, where the tissue block comprises a tissue sample embedded within an embedding material, and a vision system associated with the microtome, can be provided. The vision system can include an illumination system configured to illuminate a tissue block comprising a tissue sample embedded within an embedding material, an imaging system configured to image the tissue block and generate imaging data associated with the tissue block, and a processor in communication with the vision system, where the processor is programmed to receive the imaging data and determine when the tissue block has been sufficiently sectioned by the microtome based on the imaging data.

[0015] In some embodiments, the processor is further programmed to determine when the tissue block has been sufficiently sectioned by recognizing the amount of tissue sample that is exposed. In some embodiments, the processor is further programmed to determine the expected contour, size, or shape of the tissue sample from baseline imaging data generated by imaging the tissue block using structured light prior to the step of removing one or more sections from the tissue block. In some embodiments, the illumination system is configured to illuminate the tissue block using structured light.

[0016] In some embodiments, the histology system can further include a transfer medium for transferring one or more sections comprising the tissue sample from the tissue block to one or more slides, and the processor is further programmed to compare one or more sections on the tissue block, on the transfer medium, or on one or more slides to baseline imaging data generated by imaging the tissue block using UV light prior to the step of removing one or more sections from the tissue block.

[0017] In some embodiments, a vision system is provided that includes an illumination system configured to illuminate a tissue block comprising a tissue sample embedded within an embedding material, an imaging system configured to image the tissue block and generate imaging data of the tissue block, and a processor in communication with the imaging system, the processor being programmed to receive the imaging data and determine, based on the imaging data, an exposure of the tissue sample up to a predetermined criterion.

[0018] The present disclosure also relates to systems and methods for quality control in a histology system. In some embodiments, a method is provided that includes receiving a tissue block comprising a tissue sample embedded within an embedding material, imaging the tissue block and creating first imaging data of the tissue sample in a tissue section on the tissue block, removing the tissue section from the tissue block, the tissue section comprising a portion of the tissue sample, imaging the tissue section and creating second imaging data of the tissue sample in the tissue section, and comparing the first imaging data to the second imaging data and verifying a correspondence of the tissue sample in the first imaging data and the second imaging data based on one or more quality control parameters.

[0019] In some embodiments, if there is no correspondence of any one or more quality control parameters in the tissue sample in the first imaging data and the second imaging data, the tissue section is unqualified. In some embodiments, one or more quality control parameters include one or more of the shape of the tissue sample, the size of the tissue sample, or one or more mechanical damages. In some embodiments, the method can further include the step of transferring the tissue section to a slide using a transfer medium, and the second imaging data comprises imaging data of the tissue section on the transfer medium or imaging data of the tissue section on the slide. In some embodiments, the method can further include the step of comparing at least two of the first imaging data, the imaging data of the tissue section on the transfer medium, or the imaging data of the tissue section on the slide.

[0020] In some embodiments, if there is no correspondence of the shape or size of the tissue sample in the first imaging data and the second imaging data, the tissue section is unqualified. In some embodiments, one or more mechanical damages are selected from the group consisting of tearing, shredding, blade marks, wrinkles, cracks, air bubbles, insufficient tissue samples, and incomplete tissue samples. In some embodiments, the method can further include the step of identifying the tissue section as unqualified if one or more mechanical damages are present in the tissue sample in the second imaging data but not in the tissue sample in the first imaging data. In some embodiments, the method can further include the step of adjusting one or more operating parameters associated with the removal of the tissue section to correct one or more mechanical damages. In some embodiments, the method can further include the step of approving the tissue section if no mechanical damages are present in the tissue sample in the first imaging data and the second imaging data. In some embodiments, the method can further include the step of rejecting the tissue block if one or more mechanical damages are present in both the first imaging data and the second imaging data.

[0021] In some embodiments, one or both of the steps of imaging comprise illuminating the tissue sample with UV light, imaging the tissue sample with a visible-range camera, and creating first imaging data or second imaging data. In some embodiments, the method can further comprise illuminating the tissue section to enhance the contrast between the tissue sample and the embedded material within the tissue sample. In some embodiments, one or both of the steps of imaging comprise imaging the tissue section in one or more wavelength ranges, creating imaging data of the tissue section, segmenting the tissue sample from the embedded material based on color and intensity information in the color imaging data, and identifying the size, shape, or edge of the tissue sample in the tissue section.

[0022] In some embodiments, the method can further comprise imaging the tissue block and generating baseline imaging data of the tissue sample prior to removing one or more sections. In some embodiments, the method can further comprise illuminating the tissue block with UV light. In some embodiments, the method can further comprise comparing the first imaging data, the second imaging data, or both with the baseline imaging data. In some embodiments, the method can further comprise comparing the outline, size, or shape of the tissue sample in the first imaging data, the second imaging data, or both with the expected outline, size, or shape of the tissue sample from the baseline imaging data.

[0023] In some embodiments, provided is a vision system that includes an illumination system configured to illuminate a tissue sample, an imaging system configured to create imaging data of a tissue section illuminated by the illumination system, and a processor that receives the imaging data and communicates with the imaging system to perform one or more quality control analyses based on the imaging data. In some embodiments, the one or more quality control analyses are one or more of a comparative analysis of tissue blocks and tissue sections on slides, an analysis of mechanical properties of the tissue sections, an analysis of sufficiency of the tissue sample, or an analysis of sample representation on the slide.

[0024] In some embodiments, provided is a histology system that may include a microtome configured to generate one or more tissue sections from a tissue block, a transfer system configured to transfer one or more tissue sections from the microtome to one or more slides, and a vision system. The vision system can include an illumination system configured to illuminate a tissue sample and an imaging system configured to create imaging data of a tissue section illuminated by the illumination system. A processor receives the imaging data and communicates with the imaging system to perform one or more quality control analyses based on the imaging data. In some embodiments, the one or more quality control analyses are one or more of a comparative analysis of at least two of the tissue sections on the tissue block, the tissue sections on the transfer system, and the tissue sections on the slide, an analysis of mechanical properties of the tissue sections, an analysis of sufficiency of the tissue sample, or an analysis of sample representation on the slide.

[0025] The present disclosure also relates to systems and methods for tracking and printing within a histological system. In some embodiments, an information reader configured to read identification data associated with a tissue block, a microtome configured to cut one or more tissue sections from the tissue block, one or more slides for receiving the one or more tissue sections, and a printer configured to receive the identification data and print one or more labels for the one or more slides after the one or more tissue sections have been cut from the tissue block, wherein the one or more labels comprise information associating the one or more tissue sections on the one or more slides with the tissue block, are provided.

[0026] In some embodiments, the system can further include a transfer medium configured to transfer one or more tissue sections from the microtome to the one or more slides. In some embodiments, the transfer medium includes markings indicating identification data for the one or more tissue sections, and the markings are configured to associate the one or more tissue sections with the tissue block. In some embodiments, the system can further include a transfer medium marking device for marking the transfer medium with markings indicating identification data for the one or more tissue sections, and the markings are configured to associate the one or more tissue sections with the tissue block.

[0027] In some embodiments, the system can further include a visualization system configured to track one or more tissue sections from the microtome to one or more slides. In some embodiments, the visualization system is configured to perform a comparison between one or more tissue sections on one or more slides and one or more images of the tissue blocks or images of the sections on the transfer medium. In some embodiments, the visualization system is configured to perform a comparison between one or more tissue sections on one or more slides, on the tissue blocks, or on the transfer medium and a baseline image of the tissue sample in the tissue block generated by imaging the tissue block using UV light prior to the step of removing one or more sections from the tissue block. For example, the comparison is based on the size, shape, and contour of the tissue sample in one or more tissue sections. In some embodiments, the visualization system is configured to read one or more labels on the slide and confirm their association with identification data regarding the sample block. In some embodiments, the printer prints labels individually for one or more samples.

[0028] In some embodiments, a system can be provided that includes an information reader configured to read identification data from a tissue block, a microtome configured to cut one or more tissue sections from the tissue block, a transfer medium configured to transfer one or more tissue sections to one or more slides, and a printer. A processor is configured to receive the identification data, cause the microtome to cut one or more tissue sections, and subsequently cause the printer to print one or more labels regarding one or more slides, where the one or more labels can comprise information associating one or more tissue sections on one or more slides with the tissue block.

[0029] In some embodiments, the transfer medium includes markings indicating identification data for one or more tissue sections, and the markings are configured to associate one or more tissue sections with a tissue block. In some embodiments, the system further includes a transfer medium marking device for marking the transfer medium with markings indicating identification data for one or more tissue sections, and the markings can be configured to associate one or more tissue sections with a tissue block. In some embodiments, the system further can include a visualization system configured to track one or more tissue sections from the microtome to one or more slides. In some embodiments, the visualization system is configured to perform a comparison between one or more tissue sections on one or more slides and one or more sections on the tissue block. For example, the comparison is based on the size and edges of the tissue in one or more tissue sections.

[0030] In some embodiments, there is provided a method for tracking a sample in a microtome method, including the steps of reading identification data from a tissue block, cutting a first set of one or more tissue sections from the tissue block, and subsequent to the cutting, printing one or more labels for one or more slides, wherein the one or more labels comprise information associating one or more tissue sections on one or more slides with the tissue block, and transferring one or more tissue sections to one or more slides and labeling the one or more slides with the one or more labels.

[0031] In some embodiments, the method can further include comparing one or more tissue sections on a slide with one or more tissue sections on a block and verifying an association between the one or more tissue sections on one or more slides and the tissue block. In some embodiments, the method can further include imaging the tissue block with UV light prior to removing one or more sections from the tissue block and comparing the one or more tissue sections on one or more slides with a baseline image of the tissue sample in the tissue block generated thereby. One or more tissue sections on one or more slides can be placed on one or more slides and labeled with one or more labels, and only then can a second set of one or more tissue sections be cut. In some embodiments, the method can further include cutting a second set of one or more tissue sections only after one or more tissue sections on one or more slides have been placed on one or more slides and labeled with one or more labels. In some embodiments, the method can further include comparing one or more tissue sections on one or more slides with a baseline image of the tissue sample in the tissue block generated by imaging the tissue block with UV light prior to removing one or more sections from the tissue block. This specification also provides, for example, the following items. (Item 1) A method for sectioning a tissue block, comprising: imaging the tissue block to generate imaging data of the tissue block, wherein the tissue block comprises a tissue sample embedded in an embedding material; estimating a depth profile of the tissue block based on the imaging data, wherein the depth profile comprises the thickness of the embedding material to be removed to expose the tissue sample to a predetermined reference; removing the thickness of the embedding material to expose the tissue sample to the predetermined reference and including a method. (Item 2) progressively removing one or more sections from a tissue block comprising a tissue sample embedded in an embedding material; imaging the one or more sections and generating imaging data associated with the one or more sections; confirming that the tissue sample is exposed to the predetermined reference based on the imaging data and further including the method according to Item 1. (Item 3) The method according to Item 1, wherein the tissue block is imaged using structured light to determine the depth profile. (Item 4) A method for sectioning a tissue block, comprising: progressively removing one or more sections from a tissue block comprising a tissue sample embedded in an embedding material; imaging the one or more sections removed from the tissue block and generating imaging data associated with the one or more sections; determining when a sufficient number of the one or more sections have been removed from the tissue block to expose the tissue sample to a predetermined reference based on the imaging data and including a method. (Item 5) The method according to Item 4, further comprising imaging the tissue block and generating baseline imaging data of the tissue sample prior to removing the one or more sections. (Item 6) The method according to Item 5, further comprising determining an expected contour, size, or shape of the tissue sample from the baseline imaging data. (Item 7) The method according to item 5, further comprising the step of determining a depth profile of the embedding material from the baseline imaging data in order to remove a sufficient amount of the embedding material to expose the tissue sample to the predetermined standard. (Item 8) The method according to item 5, further comprising the step of illuminating the tissue block with structured light within the UV range. (Item 9) The method according to item 5, further comprising the step of comparing imaging data of the section including the tissue sample with the baseline imaging data to determine when the tissue sample is sufficiently exposed. (Item 10) The method according to item 9, wherein the imaging data of the section comprises imaging data of the one or more sections on the tissue block, on the transfer medium, or on the slide. (Item 11) The method according to item 9, wherein the contour, size, or shape of the tissue sample in the one or more sections is compared with the contour, size, or shape of the tissue sample expected from the baseline imaging data. (Item 12) The method according to item 4, further comprising the step of determining a depth profile by one or more of parallax, focus, or light irradiation field imaging. (Item 13) The method according to item 4, further comprising the step of increasing the contrast between the tissue sample and the embedding material. (Item 14) A method for sectioning a tissue block, comprising: removing a thickness of the embedding material configured to expose the tissue sample to a predetermined standard from a tissue block comprising a tissue sample embedded in the embedding material; subsequent to the step of removing the thickness, progressively removing one or more sections from the tissue block; imaging the one or more sections removed from the tissue block and generating imaging data associated with the one or more sections; confirming from the imaging data that the tissue sample is exposed to the predetermined standard and including. (Item 15) A histological system, comprising: a microtome configured to progressively remove one or more sections from a tissue block, the tissue block comprising a tissue sample embedded in an embedding material; a vision system associated with the microtome, An illumination system configured to illuminate the tissue block comprising the tissue sample embedded within the embedding material, an imaging system configured to image the tissue block and generate imaging data associated with the tissue block, a processor in communication with the vision system, the processor being programmed to receive the imaging data and determine, based on the imaging data, when the tissue block has been sufficiently sectioned by the microtome, a vision system comprising a histology system comprising (Item 16) The histology system of item 15, wherein the processor is further programmed to determine when the tissue block has been sufficiently sectioned by recognizing the amount of tissue sample being exposed. (Item 17) The histology system of item 15, wherein the processor is further programmed to determine an expected contour, size, or shape of the tissue sample from baseline imaging data generated by imaging the tissue block using structured light prior to the step of removing one or more sections from the tissue block. (Item 18) The histology system of item 15, wherein the illumination system is configured to illuminate the tissue block using structured light. (Item 19) The histology system of item 17, further comprising a transfer medium for transferring one or more sections comprising the tissue sample from the tissue block to one or more slides, the processor being further programmed to compare one or more sections on the tissue block, on the transfer medium, or on the one or more slides to baseline imaging data generated by imaging the tissue block using UV light prior to the step of removing the one or more sections from the tissue block. (Item 20) A vision system comprising an illumination system configured to illuminate a tissue block comprising a tissue sample embedded within an embedding material, an imaging system configured to image the tissue block and generate imaging data of the tissue block, A processor that communicates with the imaging system, wherein the processor is programmed to receive the imaging data and determine the exposure of the tissue sample up to a predetermined standard. A vision system comprising the same. (Item 21) A method for quality control in a histological system, comprising: Receiving a tissue block comprising a tissue sample embedded in an embedding material; Imaging the tissue block and creating first imaging data of the tissue sample in a tissue section on the tissue block; Removing the tissue section from the tissue block, wherein the tissue section comprises a part of the tissue sample; Imaging the tissue section and creating second imaging data of the tissue sample in the tissue section; Comparing the first imaging data with the second imaging data and verifying the correspondence of the tissue sample in the first imaging data and the second imaging data based on one or more quality control parameters; A method comprising the above steps. (Item 22) If there is no correspondence of one or more quality control parameters in the tissue sample in the first imaging data and the second imaging data, the tissue section is unqualified. The method according to Item 21. (Item 23) The method according to Item 22, wherein the one or more quality control parameters include one or more of the shape of the tissue sample, the size of the tissue sample, or one or more mechanical damages. (Item 24) The method according to Item 21, further comprising transferring the tissue section to a slide using a transfer medium, wherein the second imaging data comprises imaging data of the tissue section on the transfer medium or imaging data of the tissue section on the slide. (Item 25) The method according to Item 24, further comprising comparing at least two of the first imaging data, the imaging data of the tissue section on the transfer medium, or the imaging data of the tissue section on the slide. (Item 26) If there is no correspondence of the shape or size of the tissue sample in the first imaging data and the second imaging data, the tissue section is unqualified. The method according to Item 23. (Item 27) The one or more mechanical damages described in item 23 are selected from the group consisting of tearing, shredding, blade marks, wrinkles, cracks, air bubbles, insufficient tissue samples, and incomplete tissue samples, according to the method described in item 23. 1. If one or more mechanical damages exist in the tissue sample in the second imaging data but do not exist in the tissue sample in the first imaging data, the method according to item 27 further includes the step of identifying the tissue section as ineligible. (Item 28) The method according to item 28 further includes the step of adjusting one or more operating parameters associated with the removal of the tissue section to correct one or more mechanical damages. (Item 29) If no mechanical damage exists in the tissue sample in the first imaging data and the second imaging data, the method according to item 28 further includes the step of approving the tissue section. (Item 30) If one or more mechanical damages exist in both the first imaging data and the second imaging data, the method according to item 28 further includes the step of rejecting the tissue block. (Item 31) One or both of the imaging steps include illuminating the tissue sample with UV light, imaging the tissue sample using a visible range camera, and creating the first imaging data or the second imaging data, according to the method described in item 21. (Item 32) The method according to item 21 further includes the step of illuminating the tissue section to enhance the contrast between the tissue sample and the embedding material within the tissue sample. (Item 33) One or both of the imaging steps include imaging the tissue section in one or more wavelength ranges, creating imaging data of the tissue section, segmenting the tissue sample from the embedding material based on color and intensity information in the color imaging data, and identifying the size, shape, or edge of the tissue sample in the tissue section and are included in the method described in item 21. (Item 34) Before the step of removing the one or more sections, the method according to item 21 further includes the step of imaging the tissue block to generate baseline imaging data of the tissue sample. (Item 35) The method according to item 34, further comprising the step of illuminating the tissue block with UV light. (Item 36) The method according to item 34, further comprising the step of comparing the first imaging data, the second imaging data, or both with the baseline imaging data. (Item 37) The method according to item 34, further comprising the step of comparing the contour, size, or shape of the tissue sample in the first imaging data, the second imaging data, or both with the contour, size, or shape of the tissue sample expected from the baseline imaging data. (Item 38) A vision system, An illumination system configured to illuminate a tissue section, An imaging system configured to create imaging data of the tissue section illuminated by the illumination system, A processor that receives the imaging data and communicates with the imaging system to perform one or more quality control analyses based on the imaging data A vision system comprising. (Item 39) The vision system according to item 38, wherein the one or more quality control analyses are one or more of a comparative analysis of the tissue block and the tissue section on the slide, an analysis of the mechanical properties of the tissue section, an analysis of the sufficiency of the tissue sample, or an analysis of the sample representation on the slide. (Item 40) A histological system, A microtome configured to generate one or more tissue sections from a tissue block, A transfer system configured to transfer the one or more tissue sections from the microtome to one or more slides, A vision system, An illumination system configured to illuminate a tissue sample, An imaging system configured to create imaging data of the tissue section illuminated by the illumination system A vision system comprising, A processor that receives the imaging data and communicates with the imaging system to perform one or more quality control analyses based on the imaging data A histological system comprising. (Item 41) The one or more quality control analyses as described above include at least two of comparative analyses of the tissue section on the tissue block, the tissue section on the transfer system, and the tissue section on the slide, analysis of mechanical properties of the tissue section, analysis of sufficiency of the tissue sample, or analysis of sample representation on the slide, and the histological system according to item 40 includes one or more of these analyses. (Item 42) A system comprising: An information reader configured to read identification data associated with a tissue block; A microtome configured to cut one or more tissue sections from the tissue block; One or more slides for receiving the one or more tissue sections; A printer configured to receive the identification data and, after the one or more tissue sections are cut from the tissue block, print one or more labels for the one or more slides, wherein the one or more labels comprise information associating the one or more tissue sections on the one or more slides with the tissue block; A system comprising the above components. (Item 43) The system according to item 42, further comprising a transfer medium configured to transfer the one or more tissue sections from the microtome to the one or more slides. (Item 44) The system according to item 43, wherein the transfer medium includes markings indicating the identification data for the one or more tissue sections, and the markings are configured to associate the one or more tissue sections with the tissue block. (Item 45) The system according to item 43, further comprising a transfer medium marking device for marking the transfer medium with markings indicating the identification data for the one or more tissue sections, and the markings are configured to associate the one or more tissue sections with the tissue block. (Item 46) The system according to item 42, further comprising a visualization system configured to track the one or more tissue sections from the microtome to the one or more slides. (Item 47) The visualization system of claim 46, configured to perform a comparison between the one or more tissue sections on the one or more slides and one or more images of the tissue block or an image of the section on a transfer medium. (Item 48) The visualization system of claim 47, configured to perform a comparison between the one or more tissue sections on the one or more slides, on the tissue block, or on the transfer medium and a baseline image of the tissue sample in the tissue block generated by imaging the tissue block using UV light prior to the step of removing the one or more sections from the tissue block. (Item 49) The system of claim 48, wherein the comparison is based on the size, shape, and contour of the tissue sample in the one or more tissue sections. (Item 50) The visualization system of claim 46, configured to read the one or more labels on the slide and confirm their association with the identification data regarding the sample block. (Item 51) The system of claim 42, wherein the printer prints the label individually for the one or more samples. (Item 52) A system comprising: an information reader configured to read identification data from a tissue block; a microtome configured to cut one or more tissue sections from the tissue block; a transfer medium configured to transfer the one or more tissue sections to one or more slides; a printer; and a processor configured to receive the identification data, cause the microtome to cut the one or more tissue sections, and subsequently cause the printer to print one or more labels regarding the one or more slides, the one or more labels comprising information associating the one or more tissue sections on the one or more slides with the tissue block. The system comprising the above components. (Item 53) The transfer medium includes markings indicating the identification data for the one or more tissue sections, and the markings are configured to associate the one or more tissue sections with the tissue block, the system according to item 53. (Item 54) The system according to item 53 further comprises a transfer medium marking device for marking the transfer medium with markings indicating the identification data for the one or more tissue sections, and the markings are configured to associate the one or more tissue sections with the tissue block. (Item 55) The system according to item 53 further comprises a visualization system configured to track the one or more tissue sections from the microtome to the one or more slides. (Item 56) The system according to item 53 further comprises a visualization system configured to perform a comparison between the one or more tissue sections on the one or more slides and the one or more sections on the tissue block. (Item 57) The comparison is based on the size and edges of the tissue in the one or more tissue sections, the system according to item 56. (Item 58) A method for tracking samples in the microtome method, the method comprising: reading identification data from a tissue block; cutting a first set of one or more tissue sections from the tissue block; subsequent to cutting, printing one or more labels for one or more slides, the one or more labels comprising information associating the one or more tissue sections on the one or more slides with the tissue block; transferring the one or more tissue sections to the one or more slides and labeling the one or more slides using the one or more labels and including. (Item 59) The method according to item 58 further comprises comparing the one or more tissue sections on the slide with the one or more tissue sections on the block and verifying the association between the one or more tissue sections on the one or more slides and the tissue block. (Item 60) The method according to item 58, further comprising the step of cutting a second set of one or more tissue sections only after the first set of one or more tissue sections is placed on the one or more slides and labeled with the one or more labels. (Item 61) The method according to item 58, further comprising the step of comparing the one or more tissue sections on the one or more slides with a baseline image of a tissue sample in the tissue block generated by imaging the tissue block using UV light prior to the step of removing the one or more sections from the tissue block.

Brief Description of the Drawings

[0032] The preferred embodiments will be described in detail below with reference to the drawings so that those skilled in the art to which the subject invention pertains can more readily understand the method of fabricating and using the surgical devices and systems disclosed herein.

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[0069] Detailed Description The present disclosure provides systems and methods for various quality control analyses on tissue samples during the microtome process. Exemplary embodiments of systems and methods for quality control of tissue samples are shown in FIG. 1. Such a system can be configured to provide various types / aspects of quality control 100, including printing and tracking 102, tissue sectioning analysis 104, and mechanical tissue integrity analysis 106.

[0070] As shown, tissue sections can be cut from tissue samples such as sample blocks. Various quality control analyses can be performed in relation to the tissue sections to determine their integrity. As will be explained in more detail below, one or more analyses can be performed while the tissue sections are being transported from the sample block to a slide or other medium, or once the tissue sections are located on a slide or other medium. The process can be performed manually or using an automated system.

[0071] To perform various quality control analyses, various images of tissue blocks and tissue sections for comparison can be taken. For example, without limitation, images before tissue sectioning to create a baseline image of the block, the face of the tissue block, the tissue section after being cut from the tissue block, the tissue section while it is being transported via a transport system or tape, and the tissue section positioned on a slide can be taken. Comparison of any of these images is used to determine whether the tissue section is sufficient and can be used once positioned on a slide. If the comparison reveals any problems or errors, depending on the type of problem or error identified, the tissue slice can be discarded, the tissue block can be discarded, and / or adjustments can be made to any of the physical components of the system. Various operating parameters can be adjusted based on the type of mechanical damage or defect detected using the vision system of the present disclosure. Such operating parameters include, without limitation, sharpening or replacing the microtome blade, adjusting the hydration time or temperature, replacing the transfer medium, or adjusting the operating parameters of the transfer medium (e.g., speed) or the pressure applied to the tissue section by the transfer medium.

[0072] In some embodiments, an analysis comparing the tissue section to a sample block is performed to match the tissue section to the sample block. In some embodiments, an analysis determining the condition of the tissue section is performed either during transport or after slide placement. Additional analyses of the tissue section can be performed, as will be discussed in more detail below. Further, various methods can be used to perform quality control, including the use of a visualization / imaging system configured to image one or more of the tissue sample block, the tissue section during transport, and the tissue section on the slide so that the images can be used to perform one or more quality control analyses.

[0073] In some embodiments, a histology system can provide tracking and comparative analysis of cut tissue sections in a manual or automated tissue transfer device / system. A determination system analyzes / compares one or more parameters or characteristics / features of a cut tissue section on a microscope slide and a sample block. Alternatively, or in addition, the system analyzes / compares one or more parameters or characteristics / features of a cut tissue section on a microscope slide and a cut tissue section (slice) immediately after it was cut from a sample block. These systems ensure that the section on the glass slide properly matches the tissue sample block. This is the comparative aspect of a quality control system that improves sample tracking in a laboratory.

[0074] In some embodiments, a system that can provide feedback is provided to enable self-correction or adaptation. This feedback system provides another aspect of quality control.

[0075] In some embodiments, the system also provides a quality control feedback system. In some embodiments, for example, once the quality control system sends a flag, the feedback process can be triggered, and as a result, various actions can be automatically performed without human intervention. For example, when a flag, e.g., a discrepancy, exists, examples of downstream actions that can be taken include recalibrating the instructions or algorithms, thus enabling the system / process to self-correct or adapt. In some embodiments, the feedback process can interact with the block sectioning determination. In the quality control process when a defect is detected, the root cause is explored to solve the problem. Thus, when a defect is detected by the quality control process, a predetermined set of possible root causes will be checked. For example, if there are air bubbles under a tissue section on the glass, this could mean that the transfer medium (tape) to the glass applicator roller has a defect, and the user can be warned about this. Or, if the tissue is shredded in the automated QC image, the system will force a new sectioning blade change in the microtome. Once the root cause is identified, one or more operating parameters can be changed as described above.

[0076] For example, in an exemplary embodiment of a feedback system, the QC system suddenly begins to generate an increased number of flags regarding section quality on a certain day. This can indicate that the conditions in the system have changed and corrective action is required. The increased mismatch rate can trigger a set of self-tests, one of which can then trigger a correction mechanism (e.g., replace the microtome blade). As another example, assume that it is noted that the tissue begins to disappear on the slide at a high rate. This can mean that the sectioning algorithm has failed or the tissue has detached from the tape, and again, the self-tests can trigger corrective action. Other actions in response to various events are also envisioned in the quality control feedback system, so it should be understood that these are provided merely as examples.

[0077] It should be understood that the feedback loop can be used in combination with other quality systems disclosed herein.

[0078] It should be understood that any transfer medium other than the tape (also referred to as a transport medium) can be utilized. Thus, since the systems and methods disclosed herein are fully applicable to transfer media other than the tape, the reference to tape in this specification is used for convenience.

[0079] Sample Preparation

[0080] The sample can be a tissue, an organ, an organism, a frozen liquid, or other biological sample. In some embodiments, the sample can be pre-stained or pre-treated in some other manner to facilitate sectioning, as will be discussed in more detail below. The process of initial preparation of the sample can, in some embodiments, include a) transfer of the biological tissue, when removed from the organism, into a fixative, such as a formalin container, b) after fixation, transfer of the tissue into a labeled tissue cassette, c) after transfer to the cassette, the tissue being processed by i) dehydration via immersion in alcohol to remove water and formalin, ii) clearing via a solvent to remove the alcohol, and iii) application of an embedding material, such as paraffin wax, to surround the tissue within a large block of the molten material to create a "sample block". The paraffin is poured over the dried and chemically treated tissue within the mold. In some cases, the histotechnologist compresses the tissue during the molding process, but paraffin is always present between the bottom of the mold and the tissue sample. When the block solidifies, this provides a support matrix during the tissue sectioning process. That is, when the paraffin cools, it is removed from the mold and thus takes the form of a paraffin block in which the tissue is embedded. The molded tissue-paraffin combination is supported on a plastic cassette. The plastic cassette provides features for holding the tissue within a microtome clamp. On the opposite side of the cassette where this is clamped, the paraffin / tissue combination is cantilevered. This side corresponds to the bottom of the mold in the previous step. Excess paraffin resulting from the molding needs to be removed so that the tissue sections can be identified and analyzed / evaluated. The thickness of the excess paraffin varies significantly from tens of microns to hundreds of microns.

[0081] Plastic cassettes that hold the tissue within the paraffin mold can have different colors. In some embodiments, the quality control system detects the color of the cassette that holds the sample block to cross-check the sample type. In some laboratories, the laboratory assigns these colors to a certain tissue type. For example, the laboratory may choose to use a pink cassette for breast tissue. When the system detects a certain color that is significant to the laboratory, this can cross-check the sample type using laboratory data to ensure that the colors match. This provides an additional backup for the quality control system. Also, this data can be used during image processing such that variations in the image background are effectively filtered out and removed.

[0082] When the tissue block is placed in the chuck for cutting (sectioning), the paraffin side faces the cutting blade of the microtome, but the molding process is not complete and the tissue is not on the surface of the paraffin but under the paraffin layer. The new block first undergoes sectioning with relatively thick sections, removing a 0.1 mm to 1 mm layer of paraffin wax on top of the tissue sample. After removal of the paraffin layer on this surface, when the complete contour of the tissue sample is exposed, the block is in a state where it can be sectioned. Once this paraffin layer is removed, in clinical and research settings, the tissue is typically sectioned to a thickness of 3 μm to 5 μm. This process of removing the paraffin layer and exposing a large cross-section of the tissue is referred to as block sectioning. After removal of the paraffin layer on this surface, the tissue sample is exposed and in a state where it can be sectioned and is placed on a tape for transfer to a glass slide for analysis, for example, pathology or histology. That is, when sufficient paraffin is removed (the block is said to be "sectioned"), subsequent block sectioning provides tissue sections for placement on a glass slide for analysis (which is further processed for evaluation).

[0083] It should be noted that although paraffin is described herein as an embedding material, other embedding materials, including frozen sections, may also be utilized.

[0084] Visualization System

[0085] In some embodiments, the histological system includes a visualization system that may include an illumination system and an imaging system. The illumination system aids in imaging / discriminating / distinguishing the tissue and paraffin, which can then be imaged by the imaging system for evaluation. Thus, the discrimination of tissue from paraffin is enhanced with respect to the images taken on the sample block, tape, and / or slide. Various illumination systems and imaging systems are discussed below. In some embodiments, the visualization system utilizes an appropriate optical system (illumination system / method, imaging system / method, detection system / method), followed by calculation processing. This calculation processing provides a comparative assessment of the images for quality control, as will be described in more detail below.

[0086] It should be understood that the term "image" includes data in any form or format generated by an imaging system that represents an image or can be analyzed to determine information about the object being imaged otherwise. The images taken by the visualization system can be used such that the images themselves can be processed, analyzed, and / or compared for quality control analysis, or any data representing the image or the object being imaged, including data for creating an image or the image or data representing the object being imaged, can be used.

[0087] In some embodiments, the visualization system can be configured to a) image i) a tissue sample block containing tissue embedded in an embedding material, ii) a block face of the tissue sample block, iii) a tissue section after being cut from the sample block, iv) a transfer medium transporting the cut tissue section, or v) a slide containing the cut tissue section, and b) image a cut section from the sample block on either the slide or the transfer medium (e.g., tape). In some embodiments, this can be done to enhance the distinction of the tissue from the embedding material in which the tissue is embedded in order to enhance the images captured by the imaging device.

[0088] FIG. 2 illustrates an embodiment of a visualization system 110 that includes an illumination system 112 and an imaging system 114 that can be used to image one or more of a tissue block, a tissue section associated with a transport system, and a tissue section associated with a slide. As shown, the illumination system 112 illuminates a sample block 116 supported on a sample support chuck 118. Note that reference numeral 122 illustrates the sample block under high-contrast illumination where the contrast between paraffin and tissue is prominent, while reference numeral 120 illustrates the sample block under low-contrast illumination where it is more difficult to distinguish between the tissue and the paraffin. The illumination can alternatively or additionally illuminate the tissue section after being cut from the block. Various types of illumination systems are described below. An imaging device, e.g., a camera, images the sample block 116 for evaluation. Classical image processing techniques as described below can be utilized for evaluation. More recent artificial intelligence-based image processing techniques can also be utilized, and the teaching and testing phases are all conducted at the development site.

[0089] As shown, the imaging system 114, e.g., a camera, images the block face of the sample block 116 when the illumination system 112 illuminates the block face such that the tissue-to-paraffin contrast is increased. The imaging system in some embodiments can be triggered by software or hardware to take successive images of the block 116 as it is sectioned, which is done without human intervention. Classical image processing techniques based on edge detection (finding the boundaries of objects in an image by detecting discontinuities in luminance), color, hue, or intensity tracking in successive images, or similar attributes of each pixel and their variations in the context of the image can be utilized. The imaging system can be part of an automated sectioning system, which has the ability to synchronize imaging with block sectioning, whether through software operations or through hardware triggers such as position-sensing switches. As described above, images of the cut tissue sections after cutting by the microtome can be taken alternatively or in addition.

[0090] In some embodiments, line scanning is utilized. Line scanning involves constructing an image one line at a time using a line sensor that passes in a linear motion across an object. Thus, in the present method, the area of the block-tissue ribbon interface in the blade during cutting is scanned after the tissue block has been cut by the blade. Software then stitches the lines together to create an image of the cut section. The line of the section directly above the blade can be imaged, and a 2D image can be constructed from a series of line images. The line imaging device in some embodiments can be installed inside the roller for line scanning as the tape moves past the roller or as the roller moves along the tape. In some embodiments, the imaging device can be installed outside the roller. In either case, imaging can be associated with tape transport with transmitted light on one side of the tape. The illumination device can be inside or outside the roller. Some advantages of line scanning compared to taking a 2D image of an already cut tissue section are that i) the reconstructed line scan image will be a “flat” representation of the tissue surface and will have no risk of wrinkles or curls, ii) it is easy to determine the boundaries at the start and end of the section, and iii) since the blade and cutting force provide a consistent location for the tissue section, all parts of the image will be in focus.

[0091] The system can utilize one-dimensional scanning across samples, profiles, or images at all various degrees of sampling resolution.

[0092] Imaging tissue sections on a slide can also provide downstream uses related to digital pathology. When scanning and computationally processing the tissue in a slide scanning device, it is beneficial to first (geometrically) determine the location within the slide where the tissue is seated. This is beneficial for the autofocus algorithm in the scanning device or to reduce the computational steps in computer-aided diagnosis where the tissue is segmented. The systems described herein, for example, the UV illumination fluorescence mode, provide sharp visibility to enable easy segmentation. Such segmentation / location information can be passed back to the laboratory information system so that it is integrated downstream with the scanning device or computer-aided diagnosis platform when the slide with the tissue section is being used.

[0093] In some embodiments, illumination and signal capture using Raman spectroscopy can be used to identify the paraffin matrix across the tissue. Raman spectroscopy can be used to determine the vibrational, rotational, or other low-frequency modes of molecules and provide a structural guide for identifying molecules. When light excites a molecule, the molecule reflects light at different wavelengths, which thereby enables the detection of the composition. That is, Raman spectroscopy quantitatively detects a substance based on scattered light. The tissue itself is infiltrated with paraffin, but the density is lower than that of the paraffin in other areas of the block. Progressive image capture using Raman spectroscopy of the block after each section provides a quantified metric for use in comparison. This method can be used in parallel with visible light imaging to empirically determine the tissue area and will enhance the efficiency of the method implemented.

[0094] In some embodiments, the imaging system comprises a sensor and a radiation system. The method / system captures signals from the paraffin portion of the tissue block or sectioned slice such that as the paraffin across the tissue becomes thinner, the signal level decreases. This is due to the fact that different materials absorb different radiation wavelengths and the tissue and paraffin radiation absorption wavelengths are different. As the paraffin layer across the embedded tissue is removed, the absorption spectrum (e.g., IR absorption spectrum) will change. By tracking this change across successive images, the absorption level correlates with the amount of paraffin across the embedded tissue. Thus, the amount (or removal) of paraffin can be determined by a comparison or calculation process.

[0095] For tissue on a taper or slide, either a reflection mode or a transmission mode may be used. From the perspective of the location of the light source and camera, it may be beneficial to utilize the transmission mode. In particular, imaging of sections on a tape can be performed by embedding either the light source or the camera inside a transparent roller (or a roller with a transparent window) using either a light source or a line camera.

[0096] The illumination system can emit light, for example, within the UV, near IR, IR, or visible / broadband range. The emitted light can have different colors. In some embodiments, the illumination system can include LEDs, OLEDs, lasers, light bulbs, or similar light-emitting devices or materials. Also, as shown, the imaging system can include various areas for taking images including on a tissue block or on a tape or glass slide. A calculation system processes the images, for example, quantitatively, for identification and tracking.

[0097] Other lighting systems can be utilized to enhance the distinction between tissue and paraffin (or other embedding materials), other imaging systems can be utilized, and other calculation systems or comparison processing systems can be utilized for identification tracking and sample integrity checks. It should be understood that these various lighting and imaging systems and various associated methodologies described herein are provided as examples. Also, any combination of lighting systems and / or any combination of imaging systems can be utilized.

[0098] Light (coherent or incoherent) can be used via absorption, refraction, scattering, Raman scattering, fluorescence, phosphorescence, interference, and the wavelength can be a continuous or discontinuous distribution anywhere within the spectrum from X-rays to radio waves, or any combination of these modalities. Transmission in reflectance mode can be utilized.

[0099] In some embodiments, the imaging system can include a plurality of imaging devices for imaging a sample block for quality control as described herein. The imaging devices can function together to create a single 3D image of the sample block. Various types of imaging devices can be used, including but not limited to visible light cameras, spectrometers, multispectral cameras, hyperspectral cameras, mid-wavelength infrared (MWIR) cameras, and Raman spectroscopy cameras.

[0100] Hyperspectral imaging of tissue within a paraffin block enables the discrimination of the unique absorption of paraffin to be identified separately from various tissue reflection peaks in a single image. In a hyperspectral cube based on 3 - 5 nm steps in frequency, specific reflectance peaks of paraffin and tissue can be separated and the depth of the tissue can be identified.

[0101] Multi-spectral imaging that utilizes an array of illumination sources or a series of illumination sources from UV to infrared can enable the measurement of the responses of tissues and paraffins in unique spectral lines with multiple exposures at different color frequencies that elicit the specific absorption or reflection characteristics of the tissue.

[0102] In some embodiments, the tissue block may be illuminated using structured light, and the returned light can be used to determine various characteristics of the tissue block, tissue sample, or both. For example, the contour or cross-sectional area of the tissue sample or the depth profile of the tissue block can be determined using the structure. In some embodiments, the depth profile is the thickness of the embedded material to be removed. In some embodiments, such removal of the thickness can expose the tissue sample to a predetermined reference. In some embodiments, structured light refers to the illumination of the tissue block in a specific pattern. In some embodiments, the structured light may be spatially structured, i.e., the tissue is illuminated in a geometrically structured pattern such as a grid, stripes, concentric circles, etc. In some embodiments, the structured light may be spectrally structured, i.e., the tissue is illuminated simultaneously with light having different wavelengths. In some embodiments, the wavelengths may be selected from different intensities, bands, or colors. In some embodiments, the spectrally structured light is within the same, or predominantly the same, intensity range (e.g., UV), but can have different specific wavelengths within that intensity range. In some embodiments, the spectrally structured light may predominantly consist of light from one or more frequency bands, such bands being tailored to the optical properties of the tissue molecules, such optical properties including, for example, fluorescence absorption and emission spectra. As an example, a wavelength range predominantly with UV radiation can produce strong autofluorescence from a certain tissue compartment and facilitate subsequent processing steps in the present invention.

[0103] Laser dot diffusion can also be used. A green laser source at 515 nm and a diffractive optical element (DOE) are used to create an array of laser dots on a paraffin tissue block, thereby allowing the sampling of dispersion across the entire block in a single image. Different DOEs can be used to project laser arrays of different sizes onto the surface of the block. Alternatively, the block can be moved and one or more images taken to increase the laser dot density on the surface of the block. An image of the tissue block is taken while it is illuminated using the laser DOE. One or more channels of this image can be analyzed to determine light scattering. For example, when a green laser source is used to illuminate the block, the green channel will have the highest response. Also, the gray values of the image can be used for analysis.

[0104] In some embodiments, multiple cameras are provided adjacent to the block surface to simultaneously capture images of the block surface. The cameras and their software can use the unique geometric features of the block or the mechanism surrounding the block to orient each image to the same reference geometry. Images from multiple cameras can be used to construct a 3D image of the tissue within the block. Such a 3D image would increase the speed at which the block is sectioned because the microtome could pre-determine the amount by which the block is sectioned.

[0105] Images from multiple cameras can also be used to enhance 2D image quality by merging multiple images taken from different angles. The sharpness of the image will increase when more images are processed to create a single image. This can be used for image enhancement of the sample block surface, the cut tissue section, and / or the slide on which the cut tissue section is deposited.

[0106] Another way to enhance imaging is to mount the imaging device, e.g., a camera, on a computer-controlled motion stage, which is movable to various positions. In this way, multiple images of the sample block can be taken from different positions along the motion path of the stage. Alternatively, the block or tissue on the glass slide may be moved relative to the camera to have a similar effect. Thus, the relative motion of the imaging device with respect to the block or the sample can be utilized, and the relative movement represents the movement of the imaging device, the movement of the block or the glass slide, or the movement of both the imaging device and the block or the glass slide. The images can be processed to create a single image or to select the images for comparative analysis.

[0107] In some embodiments, the imaging device, e.g., a digital camera, is stored within a closed chamber. Within the closed chamber, a radiation source with a controlled polarity and wavelength can be provided to illuminate the biological tissue sample inside the closed chamber for enhancing imaging. The radiation source can have a dynamically adjustable polarity and wavelength while the biological sample is being imaged. Such illumination can be provided for the tissue on the sample block and / or the sectioned tissue sections on the slide. The system can include various software algorithms for performing different analyses. For example, an algorithm can be utilized to determine the depth of the biological tissue sample under the paraffin, an algorithm can be utilized to determine the overall or partial 3D shape of the biological tissue embedded in the paraffin, and / or an algorithm can be utilized to determine the largest surface area cross-section of the biological tissue and the depth of the cross-section. Another algorithm that can be provided is to extract the physical properties of the biological tissue from the image.

[0108] For example, the algorithm can determine physical properties including, but not limited to, the largest tissue contour, the number of tissue fragments within the sample (paraffin) block, and the depth of each fragment within the block. In some uses, the paraffin block may have multiple fragments of tissue therein. These may be the same tissue cut into multiple fragments or multiple tissue samples collected at multiple points from a patient. Placing multiple tissue fragments within the same block helps the laboratory reduce tissue staining costs. The feature (algorithm) ensures that all tissue fragments are cut in the same cross-section, i.e., that they are in the same plane.

[0109] In some embodiments, for comparative analysis, a projection system projects an orientation pattern onto the block surface and the same pattern is projected onto the slide. A software algorithm can be used to determine the relative orientation of the biological tissue sample within the paraffin block and the tissue on the slide.

[0110] The identity matching quality control system can be used in combination with any of the devices / systems and methods described herein.

[0111] Tissue Block Analysis

[0112] In some embodiments, one or more images can be taken prior to sectioning of the complete tissue block. In some embodiments, cross-sectional images can be taken at a known depth into the tissue block to estimate the location, size, and shape of the tissue within the block. One or more of these images can be used to create a depth profile of the tissue embedded within the tissue block. In some embodiments, an image of the composition of the entire tissue block can be created. The tissue block can be illuminated using a visualization system as described above such that the illumination light causes fluorescence in the tissue inside the tissue block. Various illumination types can be used, but in some embodiments, the tissue block can be illuminated with UV light such that the UV penetrates through the embedding material surrounding the tissue and causes fluorescence in the tissue sample embedded within the embedding material. Thus, the tissue embedded within the embedding material can be fully contoured within the block. This can be provided as a baseline image that can be used for various comparisons to the exposed tissue on the surface of the tissue block on the transport system and on the slide after transport. An exemplary baseline image of the entire tissue block 132 is shown in FIG. 3. As shown, the baseline image 130 includes the tissue sample 134 embedded within the embedding material 136 to form the tissue block 132.

[0113] This baseline image of the overall composition of the tissue block can be used in the various methods disclosed below. For example, when determining whether a tissue block is to be sectioned, the imaged surface of the block can be compared to the baseline image. Thus, a comparison can be made between the "sectioned" area of the tissue and the "embedded" area of the tissue from the baseline image since this is related to the block sectioning.

[0114] The baseline image can also be used for other quality control analyses. For example, an image of a tissue section cut from a tissue block, a tissue section on a tape, or a tissue section on a slide can be compared to the baseline image. The tissue section can be compared to the "embedded" area within the baseline image to determine the integrity, completeness, and / or physical properties of the tissue section or to confirm that the tissue section is from a specific block.

[0115] For example, as described in more detail below in connection with FIGS. 4A-4F, the initial section has only the embedding material (FIG. 4A), while subsequent sections (FIGS. 4B-4F) contain a tissue sample, and the contour of the tissue sample in the section can be compared to the expected contour from the baseline image for sectioning, tracking, and determination of mechanical integrity. In some embodiments, the system can determine the contour of the tissue on the sample block and the contour of the cut tissue section on the transfer medium or on the slide to determine whether a match exists.

[0116] Block Sectioning Methodology

[0117] In some embodiments, provided are systems and methods for sectioning biological sample tissue, including promoting and improving block sectioning determination, i.e., detection of complete sectioning of a tissue block, for determining when a tissue layer embedded within paraffin or other embedding material has been reached. In some embodiments, this can include determination systems and methods for tissue block sectioning assessment and detection in an automated device. The system / method increases the speed of sectioning and the quality of the final sections. In some embodiments, sectioning the tissue block is related to the amount of embedding material that needs to be removed. For example, a depth profile of the tissue block can be used, and thus the depth profile is related to the thickness of the embedding material that can be removed to expose the tissue sample to a predetermined reference that is related to the distance between the surface of the embedding material and the tissue sample. For example, the predetermined reference can be an amount or depth of material such that when removed from the face of the block, it can reach the surface of the tissue sample, or the predetermined reference can be an amount of material such that when removed, a sufficient cross-section of the tissue sample on the block face is exposed. The cross-sectional area of the tissue that is exposed can vary. In some embodiments, the reference can be 20% - 60% of the cross-sectional area of the tissue sample. In some embodiments, the reference can be 20% - 80% of the cross-sectional area of the tissue sample. In some embodiments, the reference can be 40% - 60% of the cross-sectional area of the tissue sample. It should be understood that the amount of the cross-sectional area of the tissue sample that is exposed varies and can be any amount.

[0118] In some embodiments, an image-based determination system and method for tissue block sectioning assessment and detection can be used. A machine vision system can be used to facilitate and improve block fragmentation determination, i.e., detection of complete sectioning of a tissue block, for determining when a tissue layer embedded in paraffin has been reached. This machine vision system for automating block sectioning determination has an application in the automated transfer of cut tissue sections to a support (carrier) medium, such as a tape, for subsequent transfer to a microscope slide. In some embodiments, a system and method of calculation processing is provided after a visualization system utilizing an optical system as disclosed herein has been operated. Thus, the visualization system utilizes an appropriate optical system (lighting system / method, imaging system / method, detection system / method) and subsequently the calculation processing continues. This calculation processing provides a comparative assessment of images for determining the status of block sectioning.

[0119] In some embodiments, the illumination and imaging system can be used to increase the contrast between an histological section and a paraffin section of a tissue block, as shown in the progression of the images of FIGS. 4A - 4F. For example, the illumination system can emit an illumination wavelength to enhance the contrast between the tissue and the paraffin. In some embodiments, this enables the system to determine the status of the sectioning of a block containing a sample of tissue embedded within an embedding material. The method can include the steps of a) increasing the contrast of the tissue and the embedding material to enhance the discrimination therebetween, including illuminating one or more of i) the block face of the tissue sample block, or ii) the section transport medium, or iii) the slide (e.g., using UV radiation); b) imaging the tissue and the embedding material; c) processing the image to assess when the block is sectioned; and d) transferring the tissue to a tissue transport medium after the block has been sectioned. The progression shows five images as an example to illustrate the process from the initial unprocessed block in Image 1 to the fully sectioned block in Image 5. FIGS. 4A - 4F illustrate a series of six images, however, it should be understood that any number of images can be used to achieve the purpose. Six images are shown for ease of explanation. More specifically, in the images shown in FIGS. 4A and 4B, the target histological section is completely covered by paraffin, thus the first n sections will not have a tissue sample and will be followed by sections with a faint outline of the distinguishable tissue. In the image shown in FIG. 4C, the paraffin sections have been removed, but the tissue is still covered by paraffin, and a larger tissue outline is shown. In the image shown in FIG. 4D, more paraffin has been removed, but a thin layer of paraffin remains over the tissue. In the image shown in FIG. 4E, the histological section is shown within a white outline and still has paraffin around the edges (outside the boundary / periphery defined by the outline). In the image shown in FIG. 4F, the fully sectioned block is illustrated and the entire histological section is exposed.The paraffin block is shown schematically as having a top (exposed) planar surface, however, it should be noted that it is understood that the block does not necessarily have a flat surface as shown.

[0120] The sectioning determination can be achieved using a manual or automated system. In some embodiments, an automated system for determining the status of sectioning of a block containing a sample of tissue embedded within an embedding material is provided in an automated apparatus, including an imaging device for distinguishing tissue from an embedding material such as paraffin, and in response to the distinction of the tissue and the embedding material, the apparatus determines when the block is completely sectioned, and in response to determining when the block is completely sectioned, the apparatus automatically cuts and transfers a tissue section from the block for subsequent analysis. In some embodiments, after determining that the sectioning of the block is complete, the system automatically stops the sectioning. In some embodiments, after determining that the sectioning of the block is complete and the system has automatically stopped the sectioning, the cutting and transfer of the tissue section to tape is automatically initiated, and the tissue section can be mounted on a slide.

[0121] Various parameters / characteristics can be used to assess the status of sectioning of the block and to make a determination as to whether the block is completely sectioned. In some embodiments, the characteristics of the sample block or tissue section can be determined after being cut from the block. In some embodiments, an evaluation, e.g., measurement, of the sample block itself or the cut tissue section itself can be performed. In some embodiments, a comparative analysis of the sample block or cut tissue section can be performed against a pre-evaluation, e.g., measurement. In some embodiments, the sample block can be cut to a predetermined depth. It should be understood that any combination of these various parameters / characteristics can be used to make the sectioning determination.

[0122] In some embodiments, an automated system is provided to enhance / facilitate block sectioning determination in an automated tissue transfer device where a sample block is sectioned, the tissue is transferred to a tape or other medium, and then to a glass substrate suitable for microscopic analysis after further processing.

[0123] The sample block can be examined at various angles such as an angle perpendicular to the front face, an angle at the angle of incidence, an angle perpendicular to the side face, or an angle at the angle of incidence, or combinations thereof.

[0124] Block measurements can be used, or only cut sections can be used, or both can be combined.

[0125] In some embodiments, the automated determination / decision regarding block sectioning is achieved by enhancing the distinction between paraffin and tissue and determining when block sectioning is complete via an imaging system. In some embodiments, this can be achieved using the illumination system and imaging system described herein. The illumination system can assist in the discrimination / distinction of tissue and paraffin, which can then be imaged by the imaging system for evaluation. Thus, the distinction of tissue from paraffin is enhanced and images are taken on the block, holder, tape, and / or other locations. For example, a machine vision system can enhance the assessment when it reaches the tissue layer embedded in the paraffin, i.e., when the thick section of paraffin wax over the tissue sample on the sample block is removed to expose the tissue sample so that the tissue sample can be sectioned, transferred to a glass slide, and further processed for evaluation.

[0126] As shown in FIG. 5, in some embodiments, the baseline image of the tissue sample can be used to assist in the sectioning determination. In particular, at step 140, the cross-sectional area of the embedded tissue can be determined using UV fluorescence to create the baseline image. At step 142, when the tissue section is removed from the tissue block, the cross-sectional area of the tissue can be determined using one of many alternative methods. At step 144, the baseline image and the image of the block face can be compared to make a determination regarding sectioning. For example, as shown in FIGS. 4A-4F, as progressive sections are removed from the tissue block, the contour of the tissue sample within these sections will change progressively. In some embodiments, the system can determine when sectioning is complete based on the expected contour of the tissue sample based on the knowledge of the baseline image. For example, sectioning can be completed when only the upper portion of the tissue has been reached. Alternatively, sectioning may be completed when the middle section of the tissue sample has been reached. Such a determination can be made by comparing the contour of the tissue sample in the section to the expected contour of the tissue sample from the baseline image. At step 146, to confirm the sectioning determination made at step 144, the area of the embedded tissue relative to the area of the section on the tape is confirmed (e.g., using a UV illumination image on the tape).

[0127] In some embodiments, as shown in the flowchart of FIG. 6, the UV method / system can be utilized for cross-section determination. The steps are as follows, namely, i) illumination with a preselected range of wavelengths (e.g., the UV range) (step 150), ii) using an appropriate optical system in step 152 to create an image on a color camera, i.e., a camera that simultaneously captures images within a plurality of wavelength ranges such as, for example, an RGB camera, iii) in step 154, using the color and intensity information from the resulting image to segment and extract the portion of the block where the tissue is present, iv) when the cross-section cut is made, monitoring the size of the tissue region and the edge of the tissue region in step 156, and v) in step 158, detecting an appropriate change in the amount monitored in step (iv) that leads to a "cross-section determination". As discussed above, these steps are automated and do not require user input during the process. Note that the intensity in grayscale can be used as an alternative to the color image. FIGS. 7 and 8 are exemplary images of tissue ribbons under UV illumination.

[0128] In some embodiments, as shown in the flowchart of FIG. 9, UV radiation or other wavelengths can be used to enhance tissue / paraffin contrast, but observations of both the block face and the cut ribbon are made. After applying light of the illumination wavelength at step 160, the imaging system creates color images of the block face (step 162) and the cut ribbon (step 164). In this method, the first appearance of tissue fluorescence in the cut section indicates that "sectioning" has occurred. This can be done based on the total fluorescence from the ribbon at step 166 (such that there is no need to image the ribbon completely face up), or alternatively, the ribbon can be imaged face up at step 168. Such imaging can be facilitated in some embodiments by having the ribbon on a tape and comparing an image of the ribbon (using UV illumination respectively) to an image of the block face. The images are processed and evaluated at step 170 for tissue segmentation (or other characteristics / parameters described herein) so that the size and edges of the tissue regions can be assessed and quantified (step 172) and changes detected (step 174) to determine the status and completion of sectioning. In another alternative method, imaging is performed only on the cut ribbon, for example, when it is on and / or away from a tape, as an alternative to block imaging rather than as a supplement.

[0129] The flowchart of FIG. 10 illustrates various systems that can be utilized as examples for determining block sectioning of an unprocessed or partially sectioned block (step 180) or a fully sectioned block (step 182).

[0130] There are various methods for cutting into a tissue block to expose tissue embedded within paraffin or other embedding materials. Evaluation of the tissue can be performed on the sample block itself, after cutting from the block, or both. In some embodiments, a predetermined amount of paraffin can be removed from the tissue block to expose the tissue. For example, in an untreated or partially sectioned block (the latter can be the result, for example, of a failed sectioning procedure in a previous step), determination of the sectioned block can be made by taking N sections at a thickness of M μm for removal of paraffin at a fixed / pre-programmed depth (step 184). After such cutting, the block sectioning is considered complete (step 186).

[0131] In some embodiments, evaluation of the tissue block is performed after a series of one or more cuts have been made to the tissue block, determining whether the block has been sectioned after each cut. Sections are cut from the block and evaluated through imaging or other techniques of the block, the cut tissue sections, and / or the cut tissue sections on tape (or other transfer media) until a determination is made that the block has been fully sectioned. For example, X sections at a thickness of Y μm are taken (step 190), then the sections are cut from the block (step 192), an evaluation of the cut sections is performed (step 194), and it is confirmed that the block has been sectioned (step 196). This check can be by taking an image of the cut tissue sections or the cut tissue sections on tape (or other transfer media).

[0132] Evaluation of the sample block and / or the cut tissue (on or off the tape), e.g., measurement (step 202), can be performed using each cut section (step 200), and thus this leads to a decision to continue sectioning (not yet fully sectioned) or to stop sectioning (fully sectioned as in step 204). In some embodiments, the decision to continue sectioning can lead to a decision to cut several sections prior to evaluation, e.g., measurement. In some embodiments, several consecutive sections can be cut prior to leading to an evaluation or decision.

[0133] In some embodiments, the decision can be made based on an evaluation of the sample block and / or a specific cut section (on or off the tape). In some embodiments, the decision can be made by comparing a second measured value (or other criterion or parameter) to a first measured value (or other criterion or parameter) according to an algorithm. The decision can also be made based on a fixed depth rather than an evaluation of a specific block face or cut section.

[0134] In some embodiments, depth measurement can be used to (empirically) determine the tissue depth under an embedding material such as paraffin (step 210). In some embodiments, the tissue depth can be determined from a baseline image, for example, as shown in FIGS. 4A-4F. 3D imaging techniques can be used to determine the tissue depth, for example, without any further measurements. The sample block is cut to the measured depth to expose the entire surface of the tissue at step 212. At this point, the block can be considered to be fully sectioned (step 214). In some embodiments, after the sample block is cut to a predetermined depth, additional images, for example, images of the cut tissue sections on a tape (or other transfer medium) to confirm that the entire surface has been reached, can be taken at steps 216 and 218. In some embodiments, depth measurement can be used to determine the tissue depth under the embedding material at step 220. After the sample block is cut to a predetermined depth at step 222, sections are cut from the block (step 224) and evaluated, for example, by images of the cut tissue sections or the cut tissue sections on a tape (or other transfer medium), to confirm that the block is sectioned at step 226. If not fully sectioned, another section is cut from the block and checked. This continues at step 228 until a determination is made that the block is sectioned. It should be understood that other methods, including but not limited to ultrasound, X-ray imaging, and other non-light-based detection methods, as well as comparison with baseline images, can also be used to determine depth. One way to achieve this is by an imaging device that creates multiple images.

[0135] As described above, in some embodiments, determining when block sectioning has occurred is accomplished by monitoring the paraffin on the tissue section by using a signature of the paraffin that differentiates it from the tissue. Using this signature, this can be detected when all of the paraffin has been removed and sufficient tissue is exposed, which will then trigger a determination that "the block has been sectioned." Various methods can be utilized for such detection, and two methods for determining the depth of paraffin on the tissue are discussed herein as examples.

[0136] In some embodiments, the block is illuminated using infrared radiation and an image is taken. For example, infrared illumination in the wavenumber range of 2,800 - 3,000 cm^ -1 (3.3 - 3.6 μm optical wavelength range) can be used because paraffin has strong absorption within this range due to the stretching mode of CH bonds. If the block is narrowly illuminated within this spectral range (such as by using a quantum cascade type laser such as a narrowband QC laser at 3.28 μm) and an image is taken using an infrared camera, the block will initially appear black (when all block surfaces are paraffin), and then brighter regions will appear around the areas where tissue is exposed as the block is cut.

[0137] Based on the fact that paraffin scatters light, in some embodiments, one sharp light spot (or an array thereof) is illuminated on the block surface. Looking at the diffuse reflected light, the "spot size" should correlate with the depth of the paraffin, meaning that thicker paraffin means more diffusion of the light. For example, if UV illumination is used (e.g., using a laser with a sharp optical spot), this will generate a point light source of fluorescence within the block. The scattering of the incident UV light and the scattering of the fluorescence upon exit will also broaden / diffuse the spot. Thus, the spot size of the fluorescence emission will correspond to the thickness of the paraffin and thus provide a method for monitoring the depth of the paraffin.

[0138] As described above, hyperspectral imaging can be used by the illumination system. FIGS. 11A and 11B illustrate exemplary images and their associated hyperspectral cubes for adipose tissue 230 embedded within paraffin 232. The low counts of the adipose tissue layer compared to the paraffin indicate that this is still below the paraffin within the block. The significantly higher tissue reflectance in FIGS. 12A and 12B indicates that this is at the surface. Thus, the layer depth of the tissue can be identified.

[0139] As described above, laser dot diffusion using a diffractive optical element can be used for imaging, which can determine depth. For example, FIG. 13 shows the green channel of a tissue block image. Using an image processing tool, the gray values of the image can be extracted as shown in FIG. 14, which illustrates a column of 2 dots sampled at 1 pixel across a block of laser dots on the tissue block. Here, a single pixel column across the dots is sampled. When the gray value is below a certain count due to tissue absorption, for example 20 in this case, it can be declared that the area of the tissue is sectioned. The area between peaks 2-4 indicates the sectioned area of FIG. 14. The gray values from each row can be combined to obtain a 3D map of the tissue sectioning metric across the surface of the block. In other words, the method can be used as an absolute measure of sectioning without a progressive image. However, using the method for a progressive image improves its efficiency.

[0140] The method can include other illumination modalities such as UV or white light to determine the boundaries of the tissue within the paraffin block. This boundary can be used as a mask that can be overlaid on the image obtained using the laser dot array. This narrowed pixel range helps improve the accuracy of the sectioning decision-making algorithm.

[0141] In some embodiments, the system can be used in conjunction with already sectioned blocks. N sections at a thickness of M μm are made to complete the block sectioning. After the block is sectioned, it is removed from the microtome and can be hydrated. After hydration, it is returned onto the microtome. The tissue within the block can absorb more moisture than paraffin and can protrude irregularly from the paraffin matrix. Also, there is a very high likelihood that the blade will be exchanged on the microtome between sectioning and slicing. The paraffin block will need to be aligned to the blade after grinding and cutting when the pathologist will take the sections that will be stained and analyzed.

[0142] In some embodiments of the automated system, once the block is sectioned, the automated device will automatically stop the sectioning and begin taking sections from the tissue block for transfer to the tape. Thus, the automated slicing device is programmed to take tissue sections from the block face once it determines that the block is sectioned, and this transition occurs without any user intervention or input. In some embodiments, once the automated determination that the block is sectioned is made, the automated device will automatically stop the sectioning, but user input is required to initiate the tissue slicing / tape transfer process. The system can include feedback to indicate when the block is completely sectioned.

[0143] In some embodiments, the micro-scratches 240 (referred to herein as "blade trajectories") left by the blade on the block face are evaluated to make a sectioning determination, as illustrated in FIG. 15. On the initial cut section from the sample block, the blade scratches will be present only on a portion of the block. However, when the block is sectioned, the scratches will span the entire block face. This is because the block is cut with a sharp blade, but the surface of the block is not flat. On the tissue itself, the scratches are interrupted because the reflection coefficient of the tissue is different from that of the paraffin block. More specifically, even the best blades have an imperfection that leaves a trace (blade trajectory) on the tissue block during sectioning. These marks will initially be present on a portion of the block because the block face is not flat and it cannot align perfectly with the blade. In a progressive image, the blade trajectory (scratch) is left on the block face over an increasingly larger area of the block as deeper cuts are made. In other words, when sectioning the block, the entire surface is not polished at once. As this is polished, the imperfections of the block appear like fine lines on the block. When the entire surface of the block is reached, micro-scratch lines are present over the entire area of the paraffin block. The reflection of the blade trajectory lines is different on the paraffin and the tissue due to different reflection constants. The system compares the normal image of the tissue and the image captured by the blade trajectory method. This comparison provides a quantitative metric of the amount by which the block is sectioned and thus can identify whether / when the block is fully sectioned. Note that these micro-scratches are not visible under white light but are visible when illuminated at UV wavelengths, and thus the illumination system disclosed herein can be utilized with the present imaging system for blade micro-scratches.

[0144] In some embodiments, successive images are taken and compared to a prior image by a device that processes the images. In some embodiments, comparing successive images to one or more previous images assesses the change in color over time as the block is cut. In some embodiments, the size and edges of the tissue and / or paraffin are evaluated to assess the status. In some embodiments, qualitative changes in the color of the image are evaluated and quantified. In some embodiments, physical properties of the tissue, such as shape, size, edges, contours, etc., are determined / evaluated to assess the status. In some embodiments, physical properties of the tissue are extracted from the image, and the physical properties include the number of tissue fragments within the sample block and the depth of each tissue fragment.

[0145] For a completely sectioned block, this can be beneficial when not only the tissue but also the entire rectangular paraffin area of the block profile is captured. For example, a rhomboid profile can be considered "sectioned" with respect to conventional sectioning if sufficient tissue sections are captured but only the paraffin-only corners are not captured. The same is not suitable for tape transfer because the tape is applied across the entire block and would catch on the blade when sectioning at the recessed block corners. Therefore, it would be beneficial to determine where the paraffin is missing. That is, it can be beneficial to look at the removed paraffin material. This is because in the middle of the cut, if a fragment is missing, the blade does not make contact and thus the cut is deeper. This visual inspection / evaluation can be achieved by using a video or a series of photographs when the block is cut to make the sectioning decision, i.e., by visually inspecting the cut material or trimming as it is formed away from the blade edge during the cut. The location where the trimming is formed is where the blade makes contact. Looking at the location where the blade and the trimming are formed, i.e., looking at how the blade interacts with the block surface at the edge, is similar to performing a line scan of the location where the blade contacts the block. An image of the location where the trimming is formed vertically on the block can be useful. For example, if a previously sectioned block has an area sunken from drying in the middle of the tissue, it can be difficult to distinguish from a photograph. The tissue can have a few microns of depression around the peripheral paraffin and retain the sheen of the sectioned block. It is easier to perform a test cut and check across the entire block to see if trimming is formed.

[0146] There can be cases where the tissue is only slightly warped in an otherwise completely sectioned block. The sections are not taken immediately after sectioning; rather, after sectioning, there is an immersion in ice water for hydration, which typically lasts about 5 - 15 minutes and which tends to slightly modify, e.g., warp, the previously sectioned block. Usually, several disposable polished sections are taken to reset the sectioned block, but the imaging system can result in fewer discarded sections being taken.

[0147] In some embodiments, the algorithm can process the images collected at each section. This would compare it to one or more historical (pre - processed) images and determine whether the images have changed such that they indicate the blocks they were sectioned from. The system can take progressive images of the blocks as they are sectioned. In parallel, the image - processing system will evaluate each image, compare it to the historical image from the same tissue block, and determine whether to continue or stop sectioning the block. For example, a hue value can be detected in the initial unprocessed block, and as the section is cut, the hue value is compared to the initial value. The algorithm can subtract the values of successive images from the initial value, and if a predetermined value (error function) is exceeded, the system recognizes that the block is being sectioned. Other characteristics such as those described above can, alternatively or in addition, form an initial baseline for comparison calculations / assessment of successive images in order to determine the status and completion of block sectioning. It should be noted that hue is only one example of a parameter on which the algorithm's detection is based. In other algorithms, intensity changes between images at approximately the same location can be used to determine sectioning. On an unprocessed block (an unsectioned tissue block), due to the dispersive nature of the paraffin layer over the tissue, the image of the tissue will not be sharp and the boundaries of the tissue will be ambiguous. In technical terms, the intensity change between the paraffin and the tissue boundary will be gradual. As the tissue is sectioned and progressive images are taken, the paraffin layer across the tissue will be made thinner until it is completely removed. When calculating the intensity at the paraffin - tissue boundary in these images, the boundary will become increasingly sharper. With a suitable threshold, it can be determined whether the tissue is sectioned.

[0148] In some embodiments, people may teach the algorithm when the block is sectioned. Thus, in this alternative embodiment, unlike the foregoing embodiments, subjective input by people, e.g., by the user, provides the initial input. In a machine learning type algorithm, the progressive images taken as more cuts are made need to be annotated by a person to indicate whether sectioning has been achieved. This paragraph refers to these annotations.

[0149] In some embodiments, a 2D image or a 3D image can be utilized.

[0150] The block can be moved up and down in front of the imaging device in some embodiments. Alternatively, the imaging device can be moved relative to the sample block, or both the sample block and the imaging device can be moved relative to each other.

[0151] As shown in FIG. 2 and as described above, the imaging system takes an image of the tissue block to assess when sectioning has occurred. In some embodiments, an image of the tissue on the blade holder is taken as an alternative or in addition. That is, the imaging system takes an image of the cut tissue section located on the blade holder. The initial cut section will not have tissue as the untreated tissue block has a layer of paraffin over the tissue. As explained above, multiple cuts are required to remove the paraffin layer by layer to expose the tissue (see FIGS. 4A-4F). However, as the cut section progresses deeper into the sample block and a portion of the tissue begins to be cut, contrast-increasing illumination utilizing one or more than one of the illumination systems described herein will enable capture of an image of the tissue of the thin cut section (ribbon) on the blade holder. That is, under contrast-increasing illumination, the initial section (all or mostly containing paraffin) does not fluoresce, but as portions of the tissue are cut, the system will begin to detect these tissue portions. When the tissue on the ribbon resembles the contour of the tissue on the block, the system recognizes that the block is being sectioned. The imaging system in this embodiment will have an image of the entire surface of the tissue of the tissue block. Comparing the full surface image from the tissue block and the images collected from the layers removed during the sectioning operation on the blade holder, the system will determine the degree of proximity to which sectioning of the tissue is achieved such that an automated determination of complete sectioning can be affected. The system / image can also check that the paraffin portion of the section matches the profile of the block to prevent tape snagging. In an automated system, when this is recognized as the block being sectioned, in some embodiments, the apparatus can automatically stop the sectioning, i.e., the microtome can stop cutting the section.

[0152] Regarding light field imaging, in general terms, a light field camera captures both the intensity of light in a scene and the direction in which light rays are traveling through space. This is in contrast to conventional cameras that record only the intensity of light. One type of light field camera uses an array of microlenses placed in front of a conventional image sensor, otherwise, to sense intensity, color, and direction information. From this super-resolution variant spatial information, an accurate identification of the "depth" of tissue located within paraffin can be obtained. Various illuminations, multispectral, hyperspectral, and UV can be utilized to bring out the tissue within the paraffin.

[0153] Regarding depth from parallax, depth information can be obtained from 2D images by combining at least two 2D images taken from different positions with respect to an object. Keeping the camera in a fixed location, the paraffin block can be moved within a vertical plane and / or a horizontal plane to take multiple images. Then, based on the 2D images, the 3D features of the object can be calculated. Image processing tools can be used to determine depth information from multiple images of the same object. An algorithm can detect key points between stereo images to calculate the parallax. This information can then be used to calculate depth information.

[0154] Another method for identifying the depth of a set of tissues embedded within a paraffin block with respect to depth from a focus is to use the optical design of the imaging system to identify the depth of a focused tissue fragment inside the paraffin block. UV illumination fluorescence imaging enables the capture of tissues located "inside" an untreated paraffin block. A series of images of the tissue can be acquired using a stepwise method of moving the block forward towards the imaging system. The UV illumination will penetrate the untreated block and cause the tissues to fluoresce. The imaging system is set with a short depth of field (DoF). Then, a series of sequential images are captured only when the paraffin block is moved forward towards the camera system along the Z-axis in steps equal to the DoF at which the fluorescing tissue will be in focus and imaged at a given depth. An edge detection algorithm run on the green channel (grayscale) image attempts to identify the image with the greatest number of "edges" or focused details, and thus identify when most of the tissue is in focus from a series of displaced images. This is then correlated to the amount by which the block holder has been moved forward, taking into account that the depth is corrected by the refractive index (RI) of the paraffin wax in its solid form. This empirically calculated depth value can then be used to identify the number of slice cuts on the block facing the microtome required until the "sectioning" condition is reached. Since this does not require progressive images, this is a faster method for sectioning the block.

[0155] As discussed herein, the illumination system can include, for example, UV, IR, or visible / broadband. Also, in some embodiments, the imaging system can capture an image of one or more of an illuminated tissue block face, a cutting ribbon on a blade holder, a cut tissue section attached to a support medium such as a tape, or a cut tissue section on a slide. The image can, in some embodiments, be a video of the block face or ribbon during sectioning. A calculation system processes the image, for example, quantitatively, to determine when the block is being sectioned. At that point, the microtome cut sections from the sample block can be automatically transferred to the tape by device components in an automated tape transfer system for subsequent analysis of the tissue.

[0156] The system herein can also have a linked database for storing data from block sectioning to enable improvements in machine learning.

[0157] Comparison and Machine Analysis

[0158] The present disclosure provides systems and methods for quality control in a histology system. In some embodiments, the method includes receiving a tissue block comprising a tissue sample embedded in an embedding material, imaging the tissue block to create first imaging data of the tissue sample in a tissue section on the tissue block, removing a tissue section from the tissue block, wherein the tissue section comprises a portion of the tissue sample, imaging the tissue section to create second imaging data of the tissue sample in the tissue section, and comparing the first imaging data to the second imaging data to confirm the correspondence of the tissue sample in the first imaging data and the second imaging data based on one or more quality control parameters.

[0159] In some embodiments, if there is no correspondence of any one or more quality control parameters in the tissue sample in the first imaging data and the second imaging data, the tissue section is unqualified. In some embodiments, one or more quality control parameters include one or more of the shape of the tissue sample, the size of the tissue sample, or one or more mechanical damages. In some embodiments, the method can further include the step of transferring the tissue section onto a slide using a transfer medium, and the second imaging data comprises imaging data of the tissue section on the transfer medium or imaging data of the tissue section on the slide. In some embodiments, the method can further include the step of comparing at least two of the first imaging data, the imaging data of the tissue section on the transfer medium, or the imaging data of the tissue section on the slide.

[0160] In some embodiments, if there is no correspondence of the shape or size of the tissue sample in the first imaging data and the second imaging data, the tissue section is unqualified. In some embodiments, one or more mechanical damages are selected from the group consisting of tearing, shredding, blade marks, wrinkles, cracks, air bubbles, insufficient tissue samples, and incomplete tissue samples. In some embodiments, the method can further include the step of identifying the tissue section as unqualified if one or more mechanical damages are present in the tissue sample in the second imaging data but not in the tissue sample in the first imaging data. In some embodiments, the method can further include the step of adjusting one or more operating parameters associated with the removal of the tissue section to correct one or more mechanical damages. In some embodiments, the method can further include the step of approving the tissue section if no mechanical damages are present in the tissue sample in the first imaging data and the second imaging data. In some embodiments, the method can further include the step of rejecting the tissue block if one or more mechanical damages are present in both the first imaging data and the second imaging data.

[0161] After the block cross-sectioning determination is made, the cut tissue sections from the sample block are transferred to tape or (other transport medium), and subsequently transferred from the tape or other medium to a glass slide. The system ensures the following, namely: i) the sections are not mechanically lost and remain properly associated with the sample block (sample tracking), ii) the sections are not subject to mechanical damage such as wrinkles, tears, cracks, etc., or are not partially taken, which is suitable for working together, for example, suitable for pathology / histology, iii) the sections placed on the slide contain a sufficient amount of tissue and not too much paraffin to ensure that it is suitable for working together, and / or iv) it is possible to ensure one or more than one of the multi-fragmented tissues in the sample block are fully represented on the slide.

[0162] Exemplary embodiments of a system and method for mechanical quality control of tissue samples are shown in FIG. 16. Such a system includes: i) a comparative analysis 250 of the sections on the transfer medium to the slide or sample block or the sections immediately after being cut from the sample block to ensure the existence of appropriate correspondence (e.g., a check to ensure that the sections are not mechanically lost and remain properly associated with the sample block from which they were cut), and / or ii) a check 252 to ensure that the cut sections on the slide or transfer medium are not subject to mechanical damage such as wrinkles, tears, cracks, etc. that could have an adverse effect on pathology, iii) a check 254 to ensure that the cut sections on the slide or transfer medium contain sufficient tissue, and / or iv) a check 256 to ensure that the multi-fragmented tissue of the sample block is fully represented on the slide, and can be configured to provide various types / aspects of quality control. These multiple aspects can be used alone, or in combination with one of the other aspects, or in combination with two or more than two of the other aspects.

[0163] Although not limited to, various quality control analyses, including the following, can be performed on tissue samples during the microtome method. i) Comparative analysis of sections on a transfer medium to a slide or sample block or sections immediately after being cut from a sample block to ensure that there is an appropriate match (e.g., a check to ensure that the section is not mechanically lost and remains properly associated with the sample block from which it was cut), and / or ii) A check to ensure that the cut sections on the slide or transfer medium have not suffered mechanical damage such as wrinkles, tears, cracks, etc. that could adversely affect pathology. iii) A check to ensure that the cut sections on the slide or transfer medium contain sufficient tissue, and / or iv) A check to ensure that the multi-fragment tissue of the sample block is fully represented on the slide. These various quality control analyses can be used alone or in any combination thereof.

[0164] In some embodiments, a system is provided to check the condition of the tissue on a microscope slide. This provides another aspect of quality control to ensure that the tissue on the slide is in appropriate condition for further analysis / evaluation.

[0165] In some embodiments, a system is provided to check whether the sections on the slide contain a sufficient amount of tissue to render it suitable for analysis / evaluation. That is, the system ensures that there is not too much embedding material, e.g., paraffin, in the sections on the slide. This provides another aspect of quality control to ensure that the tissue on the slide is in appropriate condition for further analysis / evaluation.

[0166] In some embodiments, the system is provided to check tissue on a tape (or other transport medium) as an intermediate quality control check, for example, for tracking purposes or for tissue integrity purposes (under appropriate conditions for analysis).

[0167] In some embodiments, the system is provided to check that the multi-fragment tissue within a sample block is fully represented on a slide and to ensure that no fragments are missing. Such missing fragments may require the pathologist to request re-sectioning. This integrity check provides another aspect of quality control.

[0168] In some embodiments, tissue comparison can be facilitated by an illumination subsystem and an imaging subsystem. In some embodiments, this can be provided in an automated transfer system, which is described in more detail below. Thus, within an automated sectioning and cut tissue transfer device, an image-based automated tissue comparison system, i.e., a machine vision system for automating the tracking and quality control of tissue sections, is provided. This system can be used to compare the tissue contour on a glass slide with the tissue shape on a block face to ensure proper identification of the slide. In some embodiments, an image of a cut tissue section is taken on a transport medium, which transports the cut tissue section away from the sample block via a controller. In some embodiments, the comparing step includes determining the contour of the tissue on the sample block and the contour of a first cut tissue section. In these systems, an image of the block face just prior to taking the tissue section can be taken (alternatively, or in addition, an image of the slide after being cut from the sample block can be taken), and the image is used as a comparison image with respect to subsequent images of the same section on the slide. The calculating step compares the two images to ensure that there are no significant changes in one or more parameters or features. If a significant change is detected, e.g., if the change exceeds a predetermined parameter in some embodiments, this can be provided as a feedback signal for corrective action. The illumination system aids in the discrimination / distinction of tissue and paraffin that can be imaged by the imaging system for evaluation. Thus, in this system and method, the distinction of tissue from paraffin is enhanced, and images are taken on blocks, transfer media, slides, and / or other locations. For example, the step of comparing images includes evaluating and quantifying changes in the color of the images. In some embodiments, the step of comparing images includes determining whether there are changes in one or more of location, spatial shape, and integrity. In some embodiments, qualitative changes such as color and granularity of the images are evaluated and quantified.In some embodiments, grayscale imaging can be used. Thus, if the variation exceeds a predetermined parameter, a feedback signal is provided for corrective measures. Various lighting systems and imaging systems are discussed above.

[0169] Systems and methods of calculation processing can also be provided after a visualization system utilizing an optical system as disclosed herein is operated. Thus, the visualization system utilizes an appropriate optical system (lighting system / method, imaging system / method, detection system / method), and subsequently, the calculation processing continues. This calculation processing provides a comparison assessment of the images for determining other aspects of tracking or quality control.

[0170] The lighting system enhances the visual / imaging discrimination of the tissue and paraffin, and the imaging system captures an image of the illuminated tissue / paraffin for subsequent comparison. The lighting system enhances the visualization / imaging of the tissue on the block surface, and the imaging system captures an image of the illuminated tissue for subsequent comparison for quality control. Such imaging of individual tissue sections is performed prior to the separation of the cut tissue from the block or alternatively prior to transfer to a slide, providing a basis for comparison. Subsequently, such assessment of individual tissue sections is performed after the cut tissue section has been transferred to the slide. Various methods for discrimination are described below. Various imaging systems and various locations for imaging are also discussed below. Also discussed in detail below are various embodiments of the lighting system that create and / or enhance the contrast between the tissue and the paraffin, depending on the nature of the paraffin and the tissue. In some embodiments, the tissue of the sample block is embedded in an embedding material, and the method further includes i) illuminating the sample block using a wavelength of light to increase the contrast between the tissue of the sample block and the embedding medium of the sample block in which the tissue is embedded, and ii) illuminating the first slide and / or transport medium containing the first tissue section using a wavelength of light to increase the contrast between the tissue and the embedding medium in which the tissue is embedded, including one or both of these steps.

[0171] Comparative analysis (block vs. section)

[0172] There are various ways to achieve a comparison to determine whether a tissue section on a slide or during transport between a tissue block and a slide is within a pre-set criterion so as to correspond to the same fragment of tissue on a sample block. The system can, in addition or as an alternative, compare a tissue section on a microscope slide with a tissue section freshly cut from a sample block and determine whether they correspond to the same tissue sample comparison within a pre-set criterion. A tissue section on a transfer medium can, in addition or as an alternative, be compared with the tissue on a sample block or a microscope slide. A determination system can examine one or more parameters (characteristics / features) to make the comparison. The decision-making algorithm of the determination system then provides a queue or any action to be taken tentatively. These quality control systems speed up the process, improve the process with fewer errors, and result in fewer wasteful sections. In some embodiments, the comparison determines whether there is a matching identification between a tissue block and a tissue section, and can include, for example, matching tissue contours and edges, as well as barcodes, on sample block 260 and slide 262, as shown in FIGS. 17A and 17B. In some embodiments, the comparison determines a match between a tissue block and a tissue section during transport, as shown in FIGS. 18 and 19, which illustrate exemplary images taken from the tissue section during transport. It should be understood that the following are some examples, and other criteria, such as, for example, light, composition, mechanical properties, etc., can also be utilized.

[0173] Instead of comparing the tissue on the slide with the block face, it should be understood that a comparison can be made between the tissue section on the slide and the cut tissue section (slice) after cutting from the sample block. Thus, the discussion of systems for assessing block faces, including illumination and imaging systems, discussed herein, is fully applicable for assessing cut slices for comparison with tissue sections on slides.

[0174] The block sectioning determination can, in some embodiments, be determined according to the inventive concepts in a provisional application filed on the same day as this application and entitled "Systems and Methods for Assessment of Tissue Block Facing in Automated Tissue Transfer Systems" by the same applicant (the entire content of which is incorporated herein by reference). Other methods can also be utilized. Once the block is sectioned, the block is in a state where thin tissue sections can be cut for transfer to a transport medium (e.g., tape) and then to slide analysis. Thus, the quality control systems / methods disclosed herein can be utilized with such block sectioning determinations. In some embodiments, automated methods (processes) and systems can be used to automatically section the tissue within the paraffin block via a fully automated sectioning device, and once sectioned, the tissue is automatically cut from the block face and automatically transferred to the tape, and the tape is automatically moved via rollers to advance the cut tissue and position a subsequent portion of the tape across the block face for subsequent transfer of the cut tissue section to the tape. In some embodiments, the automated tissue sectioning apparatus also includes a slide station, and the tissue section held on the tape is automatically transported and transferred to a glass slide for analysis within the automated apparatus.

[0175] After this is cut from the block and adhered to the tape, an image of the cut tissue section on the tape (or other transfer medium) can be taken. After this is transferred from the tape to the slide, an image of the cut tissue section can be taken in addition or alternatively. An algorithm will process the images collected at each section using a calculation step, compare the images on the tape and / or slide with one or more historical (previously) images, and determine whether there are significant changes in characteristics such as location, spatial shape, and integrity. The location of the tissue section is on the (glass microscope) slide. The tissue section has a rectangular shape, and the larger ones are approximately 28 mm × 22 mm. The usable area of the glass slide is approximately 50 mm × 25 mm, and the tissue section can be transferred to any location on the slide, including rotational changes. However, such accidental transfers are not preferred. The mechatronics system can ensure that tissue sections of similar size are deposited in similar locations on the glass slide. In addition, the QC system will check whether the transferred tissue is in the expected location and orientation. The spatial shape can be considered part of the foregoing description. Tissue integrity refers to having no defects such as bubbles, tears, blade marks, shredding, cracks, and missing fragments under the tissue. Note that the historical (reference) image can include an image of the sample block before it is cut and / or an image of the section (slice) after it is cut and before it is placed on the tape or slide. Therefore, the system will take progressive images of the block as it is sectioned. The system will also take images of the cut tissue sections on the tape and / or slide. In parallel, the image processing system will evaluate each image, compare it with the historical images from the same tissue block, and ensure that it has not changed. For example, hue values can be detected in the same block, and when the section is cut, the hue value is compared with the initial value. The algorithm can subtract the values of consecutive images from the initial value and assess whether they match.As noted, hue is but one example of a detection-based parameter of the algorithm, as other characteristics as described herein may alternatively or additionally form an initial baseline for comparative calculations / assessments of successive images to determine compliance. To determine sectioning, intensity changes between images at approximately the same location can be used. On an untreated block (an unsectioned tissue block), due to the dispersive nature of the paraffin layer over the tissue, the image of the tissue will not be sharp and the boundaries of the tissue will be ambiguous. In technical terms, the intensity change between the paraffin and the tissue boundary will be gradual. As the tissue is sectioned and progressive images are taken, the paraffin layer over the tissue will be made thinner until it is completely removed. When calculating the intensity at the paraffin-tissue boundary in these images, the boundary will become increasingly sharper. With a suitable threshold, it is possible to determine whether the tissue has been sectioned.

[0176] In some embodiments, quality control is not a machine learning algorithm trained by people. Thus, in some embodiments, the methodology will not rely on expert humans to teach the machine learning algorithm how to assess quality or compliance. However, note that in some implementations, the results of classical image processing methods can be fed into the machine learning algorithm and it can be trained in the expectation that the machine learning algorithm can handle more general cases. The present method will cost-effectively increase the number of annotated images used for the AI algorithm teaching phase. In machine learning type algorithms, progressive images taken as more cuts are made need to be annotated by people to show whether sectioning has been achieved. However, alternatively, classical image processing techniques can be used to annotate images that are easier with respect to such algorithms (more structured), train the AI algorithm, and extend the applicability of the entire algorithm to more general (unstructured) data.

[0177] In some embodiments, the illumination system enhances the detection of tissue. In some cases, the processed and embedded tissue has a very faint color compared to the surrounding paraffin matrix, making it difficult to reliably capture the tissue outline using normal imaging. Thus, the application of a range of wavelengths can increase the contrast of the tissue against the paraffin. For example, UV light is used to increase the contrast over the range of 320 nm to 400 nm. Such enhanced detection facilitates the comparison of the tissue on the slide (or tape) with the block. This also facilitates checking the suitability of the tissue sections on the slide (or tape) according to other quality control aspects.

[0178] In some embodiments, the increase in contrast of the tissue from the paraffin is achieved using multi - spectral images, i.e., various wavelength emissions, to illuminate the tissue block in the same case of the block section. These images are combined to increase the contrast of the tissue sections within the tissue block. These higher - contrast images are easier to compare.

[0179] In some embodiments, the tissue sample emits fluorescence when illuminated by UV radiation, so the UV radiation can be used to illuminate the tissue. UV radiation as used herein can be construed broadly as having a wavelength shorter than optically visible blue light. However, it should be noted that in some embodiments, the actual wavelength range available may include a portion of the blue end of the optical spectrum. Since biological tissue has a number of fluorescent molecules relevant in this context, including NADH, FADH, the tissue sample glows (e.g., can glow green) under UV illumination using a diode. However, paraffin does not fluoresce under the same conditions, and the paraffin block scatters the visible portion of the UV light source and can appear in different shades or colors, e.g., can appear bluish. Three advantages of UV light can be understood. First, the UV light penetrates into the paraffin block and the fluorescence emission escapes from the paraffin block, so the method / process can clearly visualize the tissue sample embedded within the block. This is in contrast to viewing the block under visible light, which is strongly scattered by the paraffin wax, and the embedded tissue sample may not be clearly visible or may even be invisible. Second, the color (hue) of the emitted light (more precisely, the wavelength ranges of the fluorescence emission and the passive scattered emission) provides a sharp contrast between the tissue and the paraffin wax, thus enabling easy detection and segmentation of the tissue sample. Third, by imaging the cut sections on the tape (or other transport medium) and / or the slide, and by observing the clearly different fluorescence emissions emitted by the tissue, it is possible to precisely detect the tissue placement to check for integrity and to check that this matches the pre-cut tissue section.

[0180] In some embodiments, another range of illumination wavelengths utilized to illuminate block faces and tapes and / or cut tissue sections on slides is located within the infrared range. Paraffin wax has a characteristic infrared absorption spectrum. By selecting tissue portions from processing UV images, when the reflection spectrum is obtained from the face of a paraffin block in an imaging mode (or when reflection IR spectroscopy is performed in an imaging mode), the tissue can be detected when the IR spectral signature of the paraffin is decreased in a localized pattern across the tissue sample.

[0181] Infrared spectroscopy can be used in some embodiments to detect the hydration state of tissue within a block by utilizing the characteristic IR absorption spectrum of liquid water.

[0182] One method is shown in the flowchart of FIG. 20 that utilizes UV or infrared tissue illumination. The steps can include: i) illumination with a preselected range of wavelengths (e.g., the UV range) at step 270, ii) creating images of the block face (step 272) and of the tape and slide (step 274) using an appropriate optical system on a color camera, i.e., a camera that simultaneously captures images within a plurality of wavelength ranges such as, for example, an RGB camera, iii) using color and intensity information from the resulting images to segment and extract the portions where tissue is present (step 276), iv) monitoring the size and the edges of the tissue region (step 278), and v) comparison of the images for detecting conformance / change (step 280). Optionally, the comparison can also be made between the image and a baseline image (step 282). In particular, as explained above, the contour of the tissue sample can be compared to the expected contour of the tissue sample from the baseline image, and the comparison of the actual contour to the expected contour can confirm the origin of the tissue sample, i.e., that the tissue sample is derived from the tissue block having a barcode associated with the slide.

[0183] This information is used to compare the tissue on the tape with the image of the block and to compare the tissue image on the tape with the image of the block. These initial (tape and block) comparisons enable the identification of the tissue picked up from the block surface with the transfer medium (tape). The tissue can be picked up partially, or a part of the tissue can be torn and rotated. If the tissue on the tape is not of sufficiently high quality, this is not worth transferring to a glass slide. This saves time and resources. The second (slide and block) comparison. The tissue transferred to the glass slide can have air bubbles, tears, missing fragments, etc. These glass slides should not be given to the end user as a final product as they would be below standard or unhelpful. The algorithm may result in recutting of the sections or warn the user about bad blocks. Bad blocks can have tissue that was not properly embedded in the laboratory before it was introduced into this system.

[0184] As discussed above, in an automated device, these steps are automated and do not require user input during the process. Note that the intensity in grayscale can be used as an alternative to a color image. Thus, in this method, UV radiation or other wavelengths enhance the tissue / paraffin contrast as discussed herein, and observation of both the block surface and the cut tissue sections is performed. In this method, the appearance of tissue fluorescence in the cut tissue sections is detected. This can be done based on the total fluorescence from the ribbon (such that there is no need to image the ribbon completely face up), or alternatively, the ribbon can be imaged face up. Such imaging is performed by comparing the image of the tissue section on the tape or slide with the image of the block surface (each with UV illumination) on the cut tissue section (on the tape or slide). The images are processed and evaluated for tissue segmentation (or other characteristics / parameters described herein) so that the size and edges of the tissue regions can be appraised and quantified until they match.

[0185] In some embodiments, multispectral illumination can be used, and spectroscopy, i.e., obtaining information from multiple wavelengths or colors, can be employed, for example, Fourier transform infrared spectroscopy can be adopted. This can be done by simple spectroscopy from a spot selected in the imaging mode or by some other method (preliminary UV imaging) for focusing on top of the tissue. In either case, the goal is to (i) determine whether paraffin is present on top of the tissue or whether the paraffin has been removed and the tissue is exposed, or (ii) image the ribbon on the tape and detect the presence of the tissue and the reduction in the amount of paraffin, or (iii) image the ribbon on the slide away from the tape and detect the presence of the tissue and the reduction in the amount of paraffin by employing the characteristic infrared spectral signatures of paraffin and the tissue.

[0186] In some embodiments, visible / broadband illumination is used to image the block face and / or the tissue section on the tape and / or on the slide for comparison, to (i) detect qualitative changes in the image such as tissue parts with more visible / darker brown / more clearly defined edges on the block, or (ii) detect the tissue by using color / intensity information by looking at the cut tissue section.

[0187] It should be understood that the above methods can be combined such that more than one illumination system can be utilized.

[0188] In the methods herein, used individually or in combination, there are calculation steps as discussed herein where a multi - color image or spectrum is analyzed and appropriate changes are detected to ensure that the cut tissue section remains properly associated with the sample block. This calculation step can include image comparison with the original or baseline image or a pre - image, and comparisons such as hue, intensity, boundaries, etc. can be quantified for calculating the degree of difference between the images for assessment.

[0189] In the case of tissue imaging on a tape or a slide, the reference image can be an image of the tissue within the block.

[0190] The automation system in some embodiments can capture various resulting variables such as speed, temperature, humidity, timeout, etc. for input into machine learning to improve the process. If the QC system considers the section to be of good quality and this trend correlates with a range of operating temperatures, this can form the basis of a machine learning algorithm to ensure that all future blocks with similar tissue are cut under conditions that result in a good quality cut.

[0191] The decision-making process can determine that there is no appropriate correspondence (comment criteria) between the tissue on the slide and the tissue on the sample block. The decision-making process can also include finding an appropriate correspondence between the tissue on the slide and the sample block, but can also include the finding that the tissue is distorted or damaged and thus not usable.

[0192] Note that the decision-making algorithm provides a queue of recommended actions and it can be optional to follow the recommendations. However, in some embodiments, whether or not the queue is taken, the information is recorded in a database, which can potentially use machine learning to improve the decision-making algorithm.

[0193] In some embodiments, the system can provide a queue when another section to transfer to the slide should be selected. For example, the selection can be every nth section or when the images of consecutive sections vary by a certain amount of criteria.

[0194] Examples of incorrect matches will be described herein. As discussed herein, biological tissue is embedded within a paraffin matrix (or other embedding material) that forms a sample block. However, tissue embedded within a paraffin block may not necessarily have high contrast compared to the paraffin matrix. Distinguishing tissue from paraffin is more difficult and thus can make it more difficult to assess the shape of the tissue on the sample block, such as the contour, which can negatively impact image analysis. Accordingly, the illumination and imaging systems disclosed herein provide systems and methods for improving the distinction between tissue and paraffin, and thus the image of the tissue, and thus the determination (analysis) of the tissue shape, such as the contour, can be determined more accurately. This ensures a more accurate comparison of tissues, for example, by reducing potential distortion of the image due to inaccurately including paraffin as part of the tissue image. In other words, a “base” or “input” image, defined as the initial image intended to define the tissue contour on the sample block for subsequent comparison with the tissue contour on the slide, includes paraffin, and a “second” or “output” image, defined as a subsequent image of the tissue on the slide, is processed without paraffin, a false determination of non-match can occur. Similarly, a false determination of non-match can occur if the base image does not include paraffin but the second image of the tissue on the slide is defined with paraffin. Conversely, false matches can also occur if not accurately distinguished and processed.

[0195] The section thickness ranges from about 1 to about 15 μm, but most commonly is about 4 μm. At this thickness, the contrast ratio between the paraffin matrix and the tissue section is very low. In some embodiments, a quality control system ensures that the contrast ratio between the tissue and the paraffin is enhanced by the systems and methods disclosed herein in order to accurately compare the tissue itself or accurately recognize the tissue and paraffin on the image. Image post - processing efficiency increases when the image has a high contrast ratio, as explained above. When the tissue is illuminated with light in a certain wavelength range, it begins to fluoresce, while the paraffin does not fluoresce in the same wavelength range. This creates a high - contrast image. Thus, the sample block is illuminated with light in a range of wavelengths to increase the contrast ratio between the biological tissue and the paraffin matrix. Similarly, the glass slide containing the tissue section deposited thereon is illuminated with light in a range of wavelengths to increase the contrast between the biological tissue and the paraffin matrix. The slide with the tissue is illuminated from the rear, or alternatively, from the front, where the rear and front of the slide can be defined based on where the camera is placed relative to the glass slide. Thus, in these embodiments, the system can better distinguish the tissue from the paraffin for comparative analysis of the sample block and the slide.

[0196] In some embodiments, the wavelength of the light can be controlled using a filter or LED with a given range of wavelength emissions. In some embodiments, on the image capture side, a filter can be provided to enhance image capture. The set of light source, filter, and camera can be referred to as imaging hardware.

[0197] Mechanical property analysis

[0198] In the lighting and imaging systems disclosed herein, the system can be utilized to ensure that cut tissue sections do not suffer mechanical damage such as wrinkles, tears, cracks, etc. when being transferred to slides. Such a system can also be utilized to ensure that a sufficient amount of tissue is on the slide. When comparing the tape or tissue on the tissue to the block face, if there are significant differences in surface area, the algorithm can indicate that there is not sufficient tissue on the slide.

[0199] The composition can be measured by sampling the vapor of the sample information or by extracting substances from the sample by impact, and can be detected by mass spectrometry, vapor phase chromatography, or other methods. This is in the context of mass spectrometry in the sense of detecting substances in the vapor over the substance.

[0200] Mechanical properties can be measured by vibration, atomic force microscopy, or other methods. Vibration can be used to determine material properties, although in this case it is a stretch but is included here for completeness. In the case of atomic force microscopy, the attractive force on the probe and the material being probed can be used as an indication of the material properties. The reason for including this individually is for the completeness of the sectioning determination sensor. This is not an imaging modality.

[0201] In some embodiments, the quality control system can check for sample orientation and / or inversion. In some embodiments, the quality control system determines tissue orientation variations on one or both of the transfer medium and the slide and alerts the user whether components of the automation device require adjustment.

[0202] Sample sufficiency analysis

[0203] In some embodiments, the system can perform checks to ensure that the cut sections on the slide or transfer medium contain sufficient tissue. For example, the system will compare the surface area of the tissue within the block face and the surface of the section on the glass slide. If the two areas are similar within a predetermined range, tissue integrity is preserved. Another example is the presence of air bubbles under the section. Comparison of the intensities of two images will reveal whether air bubbles are present. This can also provide the location where air bubbles are present. If the air bubbles are present only within the paraffin matrix, this is not a critical failure. On the other hand, if the air bubbles are on the tissue, this indicates low-quality tissue on the slide.

[0204] Tissue sample integrity

[0205] In some embodiments, the system can perform checks to ensure that the multi-fragment tissue of the sample block is fully represented on the slide. In some tissue blocks, multiple fragments of tissue can exist. The histotechnician places the tissue fragments in a plastic cassette and while trying to push them to the bottom of the cassette, the histotechnician pours warm paraffin wax. In some cases, some of these tissues move and are not planar with the remaining parts. During sectioning, one way to confirm is to compare the block face image with the tissue on the tape or glass slide. If they have the same number of tissues, overall tissue integrity is preserved.

[0206] Tracking and Printing

[0207] In some embodiments, a just-in-time glass slide labeling printing protocol can be implemented, where labels for slides are printed after the tissue sample has been cut from the tissue block. In this manner, the glass slide is barcoded or labeled by a just-in-time printer using a barcode from a block that has just been sectioned in a microtome, and thus the freshly cut tissue section is then placed on the newly printed barcoded slide. In some embodiments, the next tissue section is cut and the label is printed only after a preceding tissue section has been placed on the slide, labeled, and optionally verified to be associated with the tissue block. In some embodiments, real-time updates are communicated to the device software via a laboratory information management system while also enabling real-time tape marking of barcode data.

[0208] In some embodiments, the present disclosure closes a loop for tracking a cut tissue sample as it passes between the block face and the destination of the glass microscope slide, providing real-time updatable tracking and identification of the tissue section location in the tissue processing device relative to the LIMS data. In some embodiments, the transfer system is labeled to associate the tissue sections placed thereon with the tissue block from which the tissue sample is cut. In some embodiments, the scanned barcode data is tracked from the time the section is placed on the tape, which includes steps of replicating the information by a tape marking / printing mechanism, replicating the barcode data by just-in-time digital printing of the label on the glass slide, transferring the tissue to the glass slide, scanning the printed barcode on the glass slide, then verifying the exact correspondence between the barcode data on the block, the printed tape, and the printed slide, and optionally communicating a summary report to the LIMS.

[0209] The present disclosure is directed to the tracking of tissue sections cut from a sample block and the just-in-time printing of glass slide identification labels for the cut tissue sections in an automated tissue transfer apparatus. Information from the tissue block is transferred to the slide in real time to ensure accurate one-to-one tracking and labeling of the tissue sections. The present disclosure overcomes the problems and deficiencies of the prior art by implementing a just-in-time glass slide labeling printing protocol. In particular, one or more tissue sections from the same tissue block are cut before the glass slide is actually printed. The glass slide can then be barcoded or labeled by a just-in-time digital printer using a barcode derived from the tissue block that has just been sectioned in the microtome. The freshly cut tissue section is then placed on the newly printed barcoded slide.

[0210] The tracking and identification of tissue sections can be accomplished by several integrated sub-assemblies and mechanisms, including, as a non-limiting example, a tape marking or tape printing device that replicates barcode data associated with the incoming tissue block, captured by a barcode reader, integrated within an automated tissue sectioning machine. The barcode data generated by laboratory information management system (LIMS) software is embodied by a printed adhesive label affixed thereon to a plastic cassette holding the tissue block. In some embodiments, the tissue transfer medium may have location markings or barcodes printed thereon before it is used for tissue transfer. When the tissue transfer medium is an adhesive tape, the location markings can be placed on the tape during a tape conversion operation.

[0211] Typically, a tissue sample (also referred to as a tissue block or sample block) is provided within a plastic cassette and embedded within paraffin wax or a similar material. The plastic cassette provides features for the sample block to be held within a microtome clamp. Once the sample block is secured within the microtome clamp for cutting (sectioning), the new block first undergoes sectioning with relatively thick sections, removing a 0.1 mm to 1 mm layer of paraffin wax over the tissue sample to expose the tissue sample. After removal of the paraffin layer on this surface, once the complete contour of the tissue sample is exposed, the block is in a state where it can be sectioned. This process of removing the paraffin layer and exposing a large cross-section of the tissue is referred to as block cross-sectioning. Once this paraffin layer is removed, in clinical and research settings, the tissue is typically sectioned to a thickness of 3 μm to 5 μm. That is, once sufficient paraffin is removed (the block is said to be “cross-sectioned”), subsequent block sectioning provides tissue sections for placement onto glass slides for analysis (to be further processed for evaluation). Tissue sections cut from the sample block can be transferred to the slides, for example, using a tape transfer mechanism. In some embodiments, the process can be automated, as disclosed, for example, in U.S. Publication No. 2017 / 0205317 by the same applicant. Other examples of automated devices and variations thereof are disclosed in U.S. Publication No. 2017 / 0003309 and U.S. Publication No. 2017 / 0328818. The entire contents of these three publications are incorporated herein by reference. It should be understood that an automated tissue device provides an example of an automated device, as a lighting / imaging system and a quality control system can be used in combination with other automated devices. Also, as discussed herein, a section tracking system can be used in combination with manual systems and methods.

[0212] Referring to FIGS. 17A and 17B, the incoming tissue block includes a label attached to the plastic cassette, i.e., a barcode number. The barcode data generated by the Laboratory Information Management System (LIMS) software provides information about the source of the tissue sample. For example, the barcode information includes a receipt number and a block ID. In some embodiments, this information can also include the patient name and the date the sample was obtained. Depending on the laboratory, additional information may be included. In addition to the barcode, the label or etching on the block can include a human-readable alphanumeric version of the data. In some embodiments, the microtome device communicates with the LIMS and enables real-time LIMS updates to be correctly matched to the appropriate tissue block from the initial pick-up by the robotic arm to the actual tissue sectioning and delivery of the tissue sections to the slides. The barcode information on the tissue block, which is optionally updated, is also printed on the slides such that there is a one-to-one correspondence between the tissue sections from the tissue block and the slides. As described in more detail below, the label for the slide can be printed after the tissue section is cut from the tissue block.

[0213] Referring to FIG. 21A, a barcode reader is provided to scan a barcode associated with an incoming tissue block. The scanning can be performed at the time of tissue sectioning. The barcode information is used to query data from the LIMS to determine the number of sections that need to be cut, the thickness of the sections, and other processing parameters. Next, one or more tissue samples can be cut by a microtome, transferred to slides, and also labeled using barcode data associated with the tissue block to create a one-to-one association between the tissue block metadata and the tissue sections on the slides. In some embodiments, the slides are labeled based on the tissue block barcode with iterative variations. For example, if the block barcode is 12345, the barcode for the first slide could be 12345-a, the second could be 12345-b, and so on. The slide label is printed just-in-time before tissue transfer to the slide.

[0214] In some embodiments, the tissue sections are transferred to the slides using tape. Note that transfer media other than tape (also referred to as transport media) can also be utilized. Thus, since the systems and methods disclosed herein are fully applicable to other transfer media as well as tape, the reference to tape in this specification is used for convenience.

[0215] In some embodiments, the tape transfer system is configured to enable a tracking and identification system. The tape transfer system can be marked using information that can be associated with the tissue block. Such marking can be done after the tissue block is received within the device or can be pre-printed on the tape. In some embodiments, the transfer medium (tape) may include location marking. The block ID assigned by the laboratory can then be associated with this location marking when the tissue section is picked up by the transfer medium, and the two IDs can be associated using software tools. Also, although there are no physical markings on the tape, virtual markings can be inferred based on the position encoders in the device, and it is conceivable that this can track the absolute position on the tape and associate this absolute position, explicitly or implicitly, with the section via a suitable control algorithm.

[0216] FIG. 21A is a schematic view of some embodiments of a device with an automated tape transfer apparatus (system) 300. In some embodiments, the tape transfer system is configured to enable a tracking and identification system. FIG. 21A illustrates the path of a tape 302 for transporting cut tissue sections after the block has been fully sectioned. FIG. 21A shows a microtome 304 that holds a sample block with a tissue sample encapsulated within a support block of an embedding material such as paraffin wax. The microtome 304 includes a blade (not shown) that is aligned to cut slices (or sections) from the face of the tissue block. Once the tissue samples are cut from the tissue block, they are mounted onto the tape that is to be transported to the slide.

[0217] The automated microtome method device can also include a tape marking system 306 that communicates with the barcode scanning and reading system 308. The tape marking system 306 can be used to mark the adhesive tape using barcode information captured from a label attached to a plastic cassette holding the tissue block. Various printing methods, including thermal or continuous inkjet printing techniques, may be used within the automated tissue sectioning device for this purpose. Additionally, a thermal transfer printing unit may also be used to generate on the tape, in situ, identification information that tracks the incoming tissue block to the cut sections on the tape, this information then enabling the correspondence between the sections on the tape and those on the slide and ensuring that the integrity of the sample is maintained.

[0218] The tape transports tissue sections from the microtome 304 for sectioning to the slide station 310. In some embodiments, the device can also include a glass slide printing system 312 and a barcode reading system 314. The printing system 312 prints a label for the slide that associates the tissue section to be placed on the slide with the tissue block from which the tissue was cut. In some embodiments, the automation system can associate a barcode identifier on the tissue block with a marking on the tape transfer medium and then associate the marking on the transfer medium with a barcode printed on the slide glass as required. This is in the context of a fully automated tissue sectioning device and provides just-in-time printing of real-time LIMS data onto the glass slide. Note that other transport devices / systems can also be used. Tissues within / on these other transport systems can be tracked according to the tape printing system described herein. Thus, the systems described herein, e.g., the tape printing system, the slide printing system, etc., are fully applicable to sections (slices) on various transport systems. Multiple levels of tracking are provided due to the tracking of the tissue at different stages in the device, e.g., new slices from the block, slices during transport, slices on the slide. In some embodiments, the label may link the slide to relevant LIMS-based information such as the originating sample tissue block and the sectioning date. The tissue block may be labeled as well. To accommodate pre-labeled blocks, an optical reader such as a barcode reader may be used to read the block label and generate the associated slide label.

[0219] In some embodiments, the print head of the tape marking device may be installed in the tape path at some arbitrary point prior to the transfer of tissue from the tissue block to the tissue transfer medium. A barcode reader adjacent to the block plastic cassette reads the barcode data on the plastic cassette so that barcode data or other alphanumeric data is replicated onto the tape transfer medium at the time of sectioning. In some embodiments, the markings on the transfer medium may not be a replication of the barcode on the block cassette, but the two markings may be associated through a software construct. In some embodiments, the information printed / etched on the tape can enable tissue tracking inside the microtome device. The match between the block barcode and the tape mark (inside the device) can be ensured by the mechanical operation of the device or, for example, by scanning the sections along with the tape mark using a camera. The glass slide is printed using a barcode associated with the barcode on the tissue cassette barcode and then finally scanned, thereby ensuring a one-to-one mapping or correspondence between the barcode data associated with the incoming block and the barcode data printed on the glass slide label, and thus ensuring tissue sample tracking, which is an important aspect of regulatory quality assurance.

[0220] In some embodiments, the match between the tissue within the block and the tissue section on the slide can be verified based on the camera image and image processing. In some embodiments, an imaging or manipulation device can be employed to check that the label on the slide is printed correctly such that the tissue section on the slide is associated with the correct tissue block. At each point in time, a physical replication or printing of the markings on the tissue transport container (block cassette, transfer medium, or glass slide) is recorded and cross-checked for proper tracking of the tissue. This provides situational awareness of how the tissue and cut tissue sections traverse through the tissue processing apparatus. In some embodiments, the disclosed methods and systems can utilize a quality control imaging system as disclosed, for example, in co-pending U.S. Application No. 62 / 980,203, filed on February 22, 2020, which is incorporated herein by reference in its entirety.

[0221] The scanned barcode data is tracked from the time the section is placed on the tape, which includes steps of replicating the information by a tape marking / printing mechanism, replicating the barcode data by just-in-time printing of the label on the glass slide, transferring the tissue to the glass slide, scanning the printed barcode on the glass slide, and then verifying the exact correspondence between the block face, tape, and slide barcode data, and optionally communicating a summary report to the LIMS.

[0222] In some embodiments, the disclosure also provides a system for checking the tissue on the tape (or other transport medium) as an intermediate quality control check, for example, for tracking purposes or for tissue integrity purposes (under appropriate conditions for analysis).

[0223] Referring to FIG. 21B, in some embodiments, the tape can be pre-printed. The transfer medium (tape) may be pre-printed with location markings thereon by a conversion process outside of the device. The block ID assigned by the laboratory can then be associated with this location marking when the tissue section is picked up by the transfer medium, and the two IDs can be associated using software tools such as creating corresponding entries in a database table.

[0224] Referring to the tracking flowchart of FIG. 22, a barcode associated with an incoming tissue block is scanned at the time of tissue sectioning using an integrated barcode reader (step 320). In some embodiments, to duplicate the LIMS-based barcode data associated with the incoming tissue block, a section transfer system (such as a tape) is marked with identification information (322), which may be the same as the barcode or include any arbitrary representation of the LIMS data. Additionally or alternatively, the physical location of the tissue section on the tape is tracked by tracking the length of the tape from a reference mark such as the start of the tape. Next, the tissue section is cut from the tissue block and placed on the tape in association with the marking on the tape (steps 324 and 326), which also corresponds to the barcode information on the tissue block. Once the tissue sample is cut from the block, the glass slide is labeled using the same barcode data associated with the tissue block by a just-in-time digital printer (step 328). The tissue section is transported from the microtome to the glass slide with the printed label and placed thereon (step 330). Optionally, a comparison can be made at step 331 between the baseline image of the tissue block and the image of the tissue section. At this point, an error-free association is established between the barcode of the block from which the section was cut and the label on the slide containing the section. Finally, the barcode data on the glass slide can be scanned to verify the slide barcode against the block barcode (step 332). In some embodiments, the block face LIMS data, the transfer media location marking, and the slide barcode data are associated (step 334) to ensure that the three pieces of information match precisely, thus ensuring the tracking of the excised tissue sample within the automated tissue sectioning device and guaranteeing regulatory quality compliance.

[0225] System Implementation

[0226] The quality control analysis described above can be achieved using an automated apparatus for the automated transfer of tissue sections from a sample block to a transfer medium such as tape and from the transfer medium to a slide. In some embodiments, an automated tape transfer system is provided that includes a controller, a support for holding a sample block of tissue embedded within an embedding medium, a cutting device configured to cut tissue sections from the sample block, and a transfer medium for transporting the cut tissue sections from the sample block. The quality control system includes one or more imaging devices configured to capture at least a first image of the sample block and at least a second image of the cut tissue section, and the first and second images are compared to confirm that the cut tissue section corresponds to the pre-sectioned tissue of the sample block, e.g., within a preset criterion.

[0227] In some embodiments, an automated method is provided for transferring cut tissue sections from a tissue sample block and providing quality control. The method includes the following steps. a) Advancing a transport medium in an automated system; b) Cutting a first tissue section of the sample block; c) Transporting the first tissue section away from the sample block, the cutting exposing a next cutting plane of the sample block; d) Transferring the first tissue section to a first slide; and e) Comparing the first tissue section to the sample block to determine if a correspondence exists.

[0228] In some embodiments, the automated method includes transporting the cut tissue section to a slide station containing the first slide for transfer to the first slide, and an image of the first cut tissue section is captured on the first slide.

[0229] Figure 23 provides a workflow diagram for section tracking. In step 340, the workflow involves a step of scanning the tissue block and a step of determining whether the sample block ID is readable using a scanning device (step 342). If not applicable, in step 344, a determination is made as to whether the block ID is readable by a histotechnologist. If not applicable, in step 346, the block is flagged, and if applicable, in step 348, the block ID is manually typed on the system. Once the block ID becomes readable either by scanning or manual entry (step 350), the tissue is sectioned to create slides as described herein (step 352). If a determination is made that the tissue on the slide has chip-outs of the blade, air bubbles, missing parts, and / or other unacceptable features as described above (step 354), additional sections are cut. If the tissue on the slide is acceptable, in step 356, a printed label for the slide is requested. In step 358, a determination is made regarding the functionality of the slide label printer. If the slide label printer does not function, the laboratory needs to execute a process to obtain a label in an alternative method (step 360). If the printer functions, in step 362, the label is printed and attached to the slide. Next, in step 364, the tissue and block face on the slide are compared (using the method / process / system described herein) to determine whether there is a match. If not applicable, in step 366, a determination is made as to whether any other processed block tissue shape matches the tissue on the slide. If not applicable, in step 368, the slide is flagged, and if applicable, in step 370, a check is made to confirm that the slide and sample block barcodes match. If not applicable, in step 372, the slide is flagged, and if applicable, the slide is acceptable for processing (step 374).

[0230] Referring to FIG. 24, the vision system of the present disclosure can be part of an automated microtome method device. In some embodiments, the automated microtome method device 400 can include a combination of mechanisms for receiving a sample block, cutting a sample / slice from the sample block, and transferring the sample cut from the block onto a tape to be transferred onto a slide for analysis. The combination of mechanisms can include at least one microtome 404, a tape transfer device 406, a slide adhesive coater 412, a slide printer 414, a slide input rack 416, a slide singulator 418 for selecting a slide from a stack of slides, and a slide output rack 420. This combination of mechanisms functions together to prepare the sample on the slide and to prepare the slide itself.

[0231] FIG. 25 is a schematic diagram of an exemplary embodiment of an automated tape transfer device (system) 430 that includes a visualization system having an illumination system and an image system. Note that other automated devices can also be utilized, and device 430 is shown as an example. FIG. 25 illustrates the path of a tape 432 for transporting a cut tissue slice after the block has been fully sectioned. FIG. 25 shows a microtome 434 that holds a sample block that includes a tissue sample encapsulated within a support block of an embedding material such as paraffin wax. The microtome 434 includes a blade (not shown) that is aligned to cut a slice (or section) from the face of the tissue block.

[0232] In addition to the take-up tape 432 and the microtome 434, the automated tape transfer device 430 of FIG. 25 includes an electric feed mechanism 436, a tape applicator 438, a slide station 440, and a take-up mechanism 442. An illumination system 444 and an imaging system 446 for the block face are shown (schematically) in the drawing. The same or different illumination and imaging systems (not shown) can be utilized for the tissue sections on the tape. An illumination system 448 and an imaging system 450 for the slides are also shown (schematically) in the drawing. The path of the tape 432 begins at the feed mechanism 436 and proceeds toward the applicator end of the microtome 434 and the tape applicator 438. The tape 432 then leaves the microtome and proceeds toward the slide station 440 and is finally stored on the take-up mechanism 442. Note that the details of the device / system 430 are described in U.S. Publication Nos. 2017 / 0205317 and 2017 / 0328818, the entire contents of which are incorporated herein by reference. The electric reel advances the adhesive tape such that the portion of the adhesive tape containing the cut section moves away from the microtome and the sample block, and a new portion of the adhesive tape is positioned and adhered to the cut surface for the new section to be cut by the microtome and transferred to the adhesive tape.

[0233] FIG. 26 shows the tape applicator when the cycle is initiated. The tape applicator moves toward the cut surface of the sample block of tissue. This presses the adhesive side of the tape, e.g., an adhesive tape, against the cut surface using the roller member of the tape applicator to adhere the tape and cover the entire cut surface with the tape. The tape applicator is then retracted in the opposite direction to reset the roller member to its original position where the roller member is away from the cut surface. In some embodiments, the cut tissue section is moved into contact with the tape after sectioning by the microtome.

[0234] FIG. 26 shows the slide station 440 of the automated tape transfer device 430 in more detail. The slide station 440 can be a UV station for transferring tissue sections on a tape to a microscope slide 460 that is pre-coated with a UV curable adhesive. A roller can then press the section on the adhesive tape onto the slide. The system of FIG. 25 includes a slide station for transfer to a slide, but it should be understood that in some embodiments, the present system does not include a slide station and after transfer of the cut sections to the tape and movement of the tape from the microtome area, the sections can be transferred from the tape to the slide by other means, such as manual transfer or storage on the tape.

[0235] The slide station 440 has a lower portion 462 with a spacer that creates a slide slot, a support section 464, a UV source 466, and a motor 468. The slide slot created by the spacer and the support section 464 hold the slide 460. The motor 468 is used to translate or move the lower portion of the slide station 440 to adjust the section location on the slide 460 so that the exact location where the sample section from the tape is deposited on the slide 460 can be controlled. The illumination and imaging system can be provided within or adjacent to the slide station to illuminate the tissue section on the slide and take an image thereof for quality control, e.g., comparison with a base image of the tissue prior to transfer to the tape. The imaging system can also be utilized to assess the condition of the tissue section on the slide for a tissue integrity check. FIG. 27 illustrates an exemplary schematic showing the tape 432 prior to being applied to the surface 470 of the sample block 472.

[0236] As described above, the illumination and imaging systems disclosed herein can be utilized with other automated devices, tapes other than adhesive tapes, and devices that do not have an automated slide station, as well as in manual systems.

[0237] The automated system provides for the use of adhesive tape or alternatively another transfer medium to support samples from tissue block sectioning. The automated system and method also provide for the automated subsequent transfer of samples from the adhesive tape to the slide. The system and method further provide improved quality control by providing methods and devices / systems for comparison of i) the tissue on the transfer medium and / or ii) the tissue on the glass slide with the tissue on the sample block or slices freshly cut from the block. This is in the context of a fully automated tissue sectioning device and provides automated quality control.

[0238] The system is described with the use of a continuous strip of adhesive tape, but it should be understood that other transfer media may also be utilized. The adhesive tape as disclosed herein adheres to the cut surface of the sample block prior to sectioning. Following the adhesion of the adhesive tape to the cut surface, the microtome initiates the cutting action. The adhesion of the adhesive tape to the cut surface supports the sections being cut by the microtome. Once the microtome has completed the cutting, the cut sections remain adhered to the adhesive tape. In an alternative embodiment, the sections can first be cut and subsequent adhesion to the transfer medium can follow.

[0239] Note that the tape provides an example of a transport device / system for tissue sections. Other transport systems such as robotic arms, a series of cups with water therein, etc. can also be utilized. The tissue within / on these other transport systems can be evaluated according to the quality control system described herein. Thus, the systems described herein, e.g., lighting systems, imaging systems, etc. are fully applicable to sections (slices) on various transport systems.

[0240] Multiple internal levels of quality control are provided due to the tracking of the tissue at different stages in the device, e.g., new slices from the block, slices during transport, slices on the slide.

[0241] In some embodiments of the quality control system described herein, slides in a slide station can be held stably (firmly). In some embodiments, the automation system further includes a support for stably holding a sample block and a support for stably holding a slide in front of one or more imaging devices.

[0242] It should be understood that the term "adhesive tape" as used herein refers to any type of bonding, including molecular bonding, mechanical bonding, etc., and may include a dry adhesive tape that provides bonding via van der Waals forces (molecular bonding), the tape peel force of which varies significantly depending on the peel angle, which minimizes section damage during peeling. The tape can be such that it leaves no residue, adheres when required, and peels off without damaging the tissue when required. Also, note that the term "continuous strip of adhesive tape" as used herein means that the tape is longer than the amount of adhesive tape used for a single section (a single sample of tissue cut from a tissue block). The adhesive area of the adhesive tape can be large enough to completely cover the cut surface of the sample block, i.e., to hold a complete section when it is sliced from the sample block.

[0243] Examples of automation devices are illustrated in FIG. 26 described above and further described in U.S. Patent Publication No. 2017 / 0205317 by the same applicant. Other examples and variations of automation devices are disclosed in U.S. Patent Publication No. 2017 / 0003309 and U.S. Patent Publication No. 2017 / 0328818. The entire contents of these three publications are incorporated herein by reference. It should be understood that these automation devices provide examples of automation devices because the illumination / imaging system and quality control system can be used in combination with other automation devices. Also, as discussed herein, the illumination / imaging system and section tracking and quality control system can be used in combination with manual systems and methods.

[0244] The automated tape transfer device may include a programmable digital controller, a processor, or another type of application-specific integrated circuit (ASIC) used to control the movement of the automated tape transfer device 1, communicate with the user of the automated tape transfer device 1, and / or communicate with the microtome 4 to which the automated tape transfer device 1 is connected. There are many movements that can be controlled within the automated tape transfer device 1. Examples of these movements include the movement of the feeding mechanism 3 and the winding mechanism 6, the movement of the lower portion 30 and the translational portion of the slide station 5, the movement of the linear actuator member, etc. The controller may also provide the user with information about the functions or conditions of the automated tape transfer device 1, such as the number of prepared slides, the number of transferred sections, the amount of tape remaining on the roll, etc. The controller is capable of receiving any type of input (e.g., mechanical, visual, electrical, etc.) to perform its control functions. The controller may also, in some embodiments, be able to control the quality control system described herein.

[0245] In some embodiments, the automated tape transfer device 1 further includes an optical device for inspecting the sample block. For example, the microtome 4 may store a plurality of sample blocks for sectioning. The optical device may be used to evaluate the condition of the cutting surface or to determine the location of the tissue within the embedding medium. In one example, a macro image of the cutting surface may enable more precise placement of the adhesive tape 2 on the cutting surface. Analysis of the cutting surface may facilitate automatic trimming of the cutting surface to expose the desired tissue for sectioning.

[0246] In some embodiments, one or more optical sensors may be used to provide feedback to the controller regarding the position and quality of the sections on the adhesive tape 2. For example, a luminance sensor approaching the backlit section of the adhesive tape 2 may distinguish between the empty portion of the adhesive tape 2 and the portion carrying the sections. This may provide the approximate location of the sections on the adhesive tape 2, which can be used as an input to the controller for various purposes such as motion control. A CCD imager or a similar device may be used to image the sections and provide feedback regarding the quality of transfer. These images may be used to check for errors in the process such as incomplete transfer of the sections, misalignment of the sections on the adhesive tape 2, the presence of section trimming waste on the tape, etc. In the case of these errors, additional sections may be taken to replace the defective sections.

[0247] Similar optical methods for inspecting sections on slides may also be used. The sensor system may provide feedback on the quality of section transfer onto the slide and alert the controller of errors in the process. The same or different optical sensors may be used for the inspection of both the tape and the slide.

[0248] The automated tape transfer device may also, in some embodiments, include an automated system for labeling the slides and sample blocks using barcodes or other markers for identification. Feasible slide labeling methods include steps of attaching an adhesive printed label, etching the label into the material, or printing the label on a dedicated location. The label may link the slide to relevant information such as the originating tissue block and the sectioning date. The sample blocks may be labeled similarly. To accommodate pre-labeled blocks, an optical reader such as a barcode reader may be used to read the block label and generate the associated slide label.

[0249] The system can also include an automated quality control system for comparing the cut tissue to the tissue on the sample block to ensure that the cut tissue is properly labeled on the slide to match the sample block.

[0250] Note that the use of the term tissue section or cut section in this specification assumes that the initial sections cut from the sample block may not contain much tissue because they may contain the material they are on top of, e.g., paraffin or other embedding media. However, what is important for histopathology are the sections of tissue, i.e., the tape areas containing sufficient tissue sections, which are one of those selected for transfer to the slide. Features for ensuring this can be provided in the manner described in this specification.

[0251] The tape transfer device (system) can include one or more automated imaging devices, such as a digital camera, for taking pictures between various stages of the automated tape feeding / advancing process. The pictures can be taken at the time of transfer of the cut section to the tape, at the time of transfer of the cut section to the slide, and / or at any other time during the process. Such pictures can provide visual / quality control as described in this specification.

[0252] Photographs of the sample block (block face) can also be taken. For example, a mismatch between the block face image and the section image on the tape is a cue regarding an error during sectioning. The macro image can be useful in the thumbnails within the database that enumerates the section images. This can be useful for roughly grasping when to begin transferring sections to the tape when cutting. These are various ways of imaging tissues within this system other than the digital camera. For example, micro-CT can be used to construct a 3D model of the tissue within the paraffin. If this system has a 3D model of the tissue within the block as input, this can use the information to determine when to stop trimming and sectioning.

[0253] In another aspect of visual / quality control, as the tape advances through the apparatus and sections are cut from the sample block by the microtome and adhered to the adhesive of the tape, a photograph (or other imaging technique) of each tape area containing the tissue sample (cut section) being transferred to the adhesive tape is taken, thereby enabling real-time analysis to ensure that the section is properly, i.e., completely, transferred to the tape. Using the same camera or imaging device, or alternatively, using another camera or imaging device, the tape with the adhered sections cut from the sample block advances to the slide station, and as the sections are transferred to the slide, a photograph (or other imaging technique) of each slide containing the sample is taken, enabling real-time analysis to ensure that the section is properly, i.e., completely, transferred to the slide. Thus, the process can be monitored to ensure that proper sections of the sample block are cut and transferred to the slide for pathology before the tape feed stops. In one embodiment, if an improper section is being transferred, the system is reversed and the tape is rewound in a direction opposite to the initial forward direction, and more sections (samples) can be collected and transferred from the sample block. Note also that multiple photographs of each tape area and each slide containing the cut sections can be taken for evaluation. The illumination and imaging systems described herein enhance this analysis.

[0254] Other information from the photographs can also be stored to identify the sample block, sections, etc., along with the marking and tracking of the blocks and sections.

[0255] In the implementation of the visual control system, a photograph of each tape area containing a section cut from a sample block by a microtome, e.g., a tissue section, is taken. The photograph is then analyzed to determine whether the cut section has been properly transferred to the tape. In further analysis, the photograph is evaluated to determine the end of sample block trimming (described below). Once the tissue section has been transferred to a slide within a slide station (downstream of the microtome), an image of the slide is also taken. The photograph of the slide is analyzed to determine whether the tissue section has been properly transferred to the slide. The photograph can also be analyzed to determine whether sufficient tissue section is contained on the slide. The photograph can also be utilized for matching to the sample block. If the section is not sufficient, e.g., if this results in containing insufficient tissue sample, for example, mostly containing paraffin, the section is not used for evaluation. The illumination and imaging systems described herein enhance these analyses. The illumination and imaging systems described herein enhance these analyses.

[0256] If further use and analysis are desired, the photograph can be stored in a database for future selection.

[0257] Figures 28A, 28B, 28C, 29, and 34 illustrate exemplary embodiments of an automated system for implementation of the methods described above. However, it should be noted that the methods and systems described above can be implemented in a manual microtome method process or other automated microtome method processes.

[0258] Referring to FIGS. 28A and 28B, in some embodiments, an automation system 500 is provided to enable automated tissue sample processing from blocks to slides. The system 500 can be designed to include a first section for cutting samples from tissue blocks. In some embodiments, for example, the first section as shown in FIG. 28B can include a block handler, at least one microtome 504, a transfer medium 506 (e.g., tape), a hydration chamber 508, and a block tray 510. The block handler, at least one microtome 504, the transfer medium 506 (e.g., tape), the hydration chamber 508, and the block tray can be designed to operate together using any combination of systems and methods to harvest, section, hydrate, slice, and transfer tissue sections to slides from tissue blocks.

[0259] In some embodiments, system 500 can include a transfer medium 506 (e.g., a tape) for receiving sample slices taken from a tissue block, for example, by a microtome 504. The transfer medium 506 can include any combination of materials or surfaces that can receive a sample sliced from the microtome 504 and transport the sliced sample to another location. In some embodiments, the transfer medium 506 can include at least one adhesive surface that can remove, receive, and / or transport a sample sliced from the microtome 504 after it is cut from the tissue block. For example, the transfer medium 506 can include any combination of tapes, such as, for example, a tape roll, a windowed tape, etc. The transfer medium 506 can include or otherwise be part of a larger mechanism for transporting the sliced sample. For example, the transfer medium 106 can be an adhesive tape wound across a combination of pulleys, wheels, spools, conveyors, etc., designed to allow the transfer medium 506 to move a sliced sample thereon from one location to another. Any other combination of transfer media can be used without departing from the present disclosure. For example, the transfer medium 506 can be a belt with ridges, depressions, etc., designed to grip and / or hold the sliced sample.

[0260] In some embodiments, the transfer medium 506 can transfer the sectioned sample from its surface to the offset assembly 522 to transfer the sample onto the slide. The offset assembly 522 can be designed to remove the sample adhered to the transfer medium 506 and place the sample on one or more slides. In some embodiments, the transfer by the offset assembly 522 can include the step of separating the actual tissue sample material to isolate the sample from non-sample material. The offset assembly 522 can use any combination of systems and methods for separating all or a portion of the biological sample from the surrounding paraffin material such that only the biological sample material is transferred to the slide. For example, the offset assembly 522 can core out the portion of the biological sample to be removed from the transfer medium 506. In some embodiments, the non-sample material (e.g., paraffin material) can remain on the transfer medium 506 to be discarded with the used transfer medium 106.

[0261] Continuing with FIGS. 28A - 28C, system 500 can also include a second section, such as shown in FIG. 28C, having a combination of mechanisms for preparing and providing slides for receiving biological samples cut from blocks (e.g., in a first section) from a transfer medium 506 (e.g., a tape), and processing the slides for analysis. In some embodiments, the combination of mechanisms for processing slides in the second section can include a slide adhesive coater 512, a slide printer 514, a slide input rack 516, a slide singulator 518, and a slide output rack 520. This combination of mechanisms can function together to prepare slides for receiving samples, affix the samples onto the slides, and deliver / compile the slides with the samples to a rack for later use. In some embodiments, an initial blank slide can be provided in a storage rack of a slide assembly for pre - processing. For example, the slide assembly can include one or more slide input racks 516 for storing a plurality of blank slides. The slide assembly can store and compile a large capacity of slides, e.g., 200 slides.

[0262] In some embodiments, the slide singulator 518 can be designed to grip slides from a stack of slides within the input rack 116. The slide singulator 518 can include any combination of mechanisms capable of picking up and transporting the slides. For example, the slide singulator 518 can be an actuated mechanical arm, a gantry, etc. Before being processed, the slide singulator 518 can provide the slides for quality control steps. During the quality control steps, an analysis can be performed on the slides to ensure that the slides are suitable for receiving a sample. For example, quality control can include the slide singulator 518 transporting the slide within the field of view of a camera to identify any potential problems associated with the slide and to provide image data for image processing to check the orientation of the slide, the condition of the slide, etc. If the slide fails the quality control inspection, it can be discarded, and if it passes, it can be transported within the system 500 and prepared to receive a sample. In some embodiments, the slides can be transported to the slide printer 514 to receive identification information printed thereon. For example, information about the sample type, sample origin, sample date, etc. can be printed on the slide. The identification information can include any combination of machine-readable and human-readable codes or text such that the slide and its contents can be properly identified and tracked. For example, the slide printer 514 can print a machine-readable barcode on the slide to identify the slide number, batch, contents, etc.

[0263] In some embodiments, the slides can be transported to the slide adhesive coater 512 to be coated by an adhesive material. For example, the slide adhesive coater 512 can spray a UV (ultraviolet) activated adhesive onto the slide, apply a UV activated adhesive tape, or perform any combination of adhesive systems or methods. In some embodiments, the adhesive can be applied in multiple layers. The multiple layers can be applied such that the slide receives a uniform coating of the adhesive and ensures clear visibility through the slide layer. In some embodiments, the slides can be inserted into a slide input rack 516 that has already been pre-treated or partially pre-treated.

[0264] Once the slide has been processed by the slide printer 514 and the slide adhesive coater 512, the slide can be transported to the transfer medium 106 to receive the sample from the transfer medium 506. For example, the slide can be transported to the offset assembly 522 to receive a sectioned tissue block sample from the transfer medium 506 (e.g., a tape mechanism). In some embodiments, prior to transferring the sample to the slide, the offset assembly 522 includes one or more cameras and can perform image processing to determine whether the sample on the transfer medium 506 is suitable for adhesion to the slide. For example, the image processing can inspect the sample to determine whether it is suitable for placement on the slide. If this is not suitable, the sample can be discarded and the transfer medium 506 can be advanced to the next sample. When the sample is suitable for placement on the slide, it can be applied to the slide. In some embodiments, the image processing can inspect the sample after it has been adhered to the slide to determine whether the placement of the sample is of sufficient quality. For example, the image processing can inspect the slide to determine whether the sample is cleanly adhered to the slide, whether there are bubbles, tears, paraffin residues, etc. If the slide is not suitable, the slide can be discarded instead of being placed in the slide output rack 520.

[0265] In some embodiments, the completed slide can be transported by the singulator 518 to be stored within the slide output rack 520. The slide can be stored within the slide output rack 520 in a predetermined order and / or arrangement such that the next step where the slide will be used can easily locate and remove the slide.

[0266] As described above, in some embodiments, system 500 can include a quality control imaging system as disclosed, for example, in co-pending U.S. Application No. 62 / 980,203, filed February 22, 2020, which is incorporated herein by reference in its entirety.

[0267] Referring to FIG. 29, in some embodiments, system 500 can be used to transfer samples from tissue blocks to slides according to the stages provided in automated process flow 600. FIG. 29 shows the process flow of steps from block to slide used in system 500 provided in FIGS. 28A-28C. In step 601, a sample tissue block can be loaded into system 500. For example, one or more tissue blocks, including tissue samples embedded in paraffin blocks, can be loaded into tray 510 and placed within system 500. In step 602, one of the sample tissue blocks can be moved from tray 510 to microtome 504 for sectioning. For example, the tissue block can be transported by a handler and placed within the chuck of sectioning microtome 504 to be sectioned. In step 603, the sectioned tissue block can be moved to hydration chamber 508 for hydration and cooling. For example, the tissue block can be transported by a handler and placed within hydration chamber 508 for a predetermined time period. After sufficient hydration is provided, in step 604, the tissue block can be moved to microtome 504 for sectioning. For example, the tissue block can be transported by a handler and placed within the chuck of sectioning microtome 504 for polishing and sectioning. The block can be provided to the same microtome 104 that performed the sectioning or a different microtome 504. Thereafter, each sectioned sample can be transferred to transfer medium 506. In step 605, the sectioned sample on transfer medium 506 can be transferred to a slide.

[0268] Simultaneously with, or subsequent to, steps 601 - 605, steps 606 - 608 can be performed to prepare one or more slides for combination with the sectioned samples from the tissue block. In step 606, a microscope slide can be selected and retrieved from a stack of new slides. For example, a slide singulator 518 can select and pull a slide from a stack of slides stored within a rack 516 of blank slides. In step 607, identification information can be printed on the selected slide. For example, the slide can be placed within a slide printer 514 for printing a machine-readable barcode thereon. In step 608, an adhesive material can be coated on the selected slide. For example, the slide can be placed within a slide adhesive coater 512 for spraying a UV-activated adhesive thereon. In step 609, the tissue sample can be transferred from the transfer medium 506 to the UV adhesive-coated slide. Additionally, during step 609, the slide can be imaged for on-board diagnostics, quality control, and sample tracking. For example, one or more cameras can be used to capture image data to be processed by an image processor with respect to a predetermined quality threshold. Once the slide passes quality control, in step 610, the completed tissue slide can be moved to an output rack 520 for storage for future analysis.

[0269] Algorithm

[0270] The flowchart of FIG. 30 illustrates the steps of a motor-controlled automated system for transferring tissue sections cut by a microtome to a tape and further transferring the tissue sections to a slide. For example, the automated system can include a movable tape or other support / conveyance medium, and the sections of the tissue are automatically transferred to the tape by the apparatus. Once the system, the imaging system, determines that the block is completely sectioned as described above so that the tissue sections can be transferred to the tape for later analysis, the feeding mechanism is automatically activated (alternatively, the apparatus can be designed such that the user will activate the feeding mechanism once the block is completely sectioned). Activation of the feeding mechanism advances the tape that is moved towards the cut surface of the sample block as described above. Next, a roller, for example, a roller member, presses the adhesive side of the tape, if an adhesive tape is utilized, onto the cut surface. The roller is then depressed so that the tape covers the entire cut surface. The linear actuator is retracted to its original position to reset the roller for subsequent application of the tape to another block for transfer of another cut section. The microtome then cuts the section covered by the tape (e.g., along a plane parallel or substantially parallel to the cut surface). The cut section conveyed by the tape is advanced to the slide station and aligned with the slide. After the cut section of the tissue is transferred to the tape by the automated apparatus, the tissue section is subsequently transported by the automated apparatus to a glass slide at the slide station and automatically transferred to the glass slide. The slide roller presses the section on the tape onto the slide, and the section can be laminated onto the slide. The slide roller is retracted to its original position, and the tape is advanced away from the slide, leaving the section on the slide. These steps of FIG. 30 are repeated until the desired number of sample sections are transferred to the tape, cut by the microtome, and transferred to the slide.

[0271] As shown in FIG. 30, at step 700, the feeding mechanism is activated to advance the tape. Next, at step 702, the linear actuator moves towards the cut surface of the sample block. At step 704, the roller member presses the adhesive side of the tape onto the cut surface. The roller member is then pressed down at step 706 to adhere the adhesive tape so as to cover the entire cut surface. At step 708, the linear actuator retracts to reset the roller member for subsequent application of the adhesive. At step 710, the microtome cuts a section covered by the adhesive tape, and the cut section advances to the slide station at step 712 and aligns with the slide. At step 714, the slide roller presses the section onto the slide, and the section is laminated onto the slide at step 716. Optionally, a comparison is made at step 717 between the baseline image of the tissue block and the image of the tissue section on the slide, which can ensure the match between the tissue block and the tissue section. At step 718, the slide roller retracts to its original position. Finally, at step 720, the tape advances away from the slide and is stored on the winding mechanism.

[0272] In some embodiments, a quality control system can be provided for checking the equipment or components of an automation system. More specifically, software algorithms can be utilized to determine whether the tissue transfer system is functioning according to manufacturing specifications based on variations in tissue images and projection patterns. This can be based, for example, on an image of the tissue on the tape. In an alternative variation, the landmark features of fixed mechanical components can be used as a reference instead of the projection pattern to determine tissue orientation variations. Such features can be useful for predictive maintenance of the device. The tissue is transferred to the tape at the same nominal location if all are functioning according to specifications. If the tissue transfer is to significantly different locations, this can alert the user that the mechanical components are out of alignment or not functioning properly. For example, if different locations are detected, this can mean that the rollers need to be aligned, the tape tension needs to be the same during transfer, for example, the tension sensor is off or the sensor is operating at different points, the applicator tension spring from the tape to the block does not still have the specified spring constant, the tape used for transfer does not have the same elastic constant and is not as specified, etc. It should be understood that while a single metric may not indicate any particular reason, this can alert the technician that there is a problem that needs to be corrected in the tissue transfer line. However, it is also envisioned that the system can provide algorithms that can more specifically detect the source of the misalignment, and thus the mechanical components that need adjustment can be identified.

[0273] The tissue imaging system is shown in combination with the tape transfer device (system) of FIG. 25, along with the flowchart of FIG. 31 that illustrates the steps of the automated tape transfer system. Note that a single imaging device, such as a digital camera, can be utilized to take a photograph adjacent to the transfer of the cut section to the adhesive of the tape. The same camera can be repositioned during the automated operation adjacent to the slide station to take a photograph of the slide after the transfer of the section to the slide. Alternatively, different imaging devices can be provided within or adjacent to the slide station to take a photograph after the transfer of the section to the slide. As described above, the apparatus of FIG. 25 can take a photograph of the cut section after the transfer of the cut section to the tape and after the transfer of the cut section from the tape to the slide, or alternatively, take a photograph only after the transfer to the tape or only after the transfer to the slide. Such photographs can be taken at the time of transfer, immediately after transfer, or downstream of the transfer (after the tape has advanced past the tape applicator or after it has advanced to the slide station). Photographs of the tape and / or the cut sections of the tape can also be taken at other times during the tape feed cycle, if desired.

[0274] First, the feeding mechanism is activated to advance the tape at step 730. The linear actuator moves towards the cut surface of the sample block at step 732. The roller member presses the adhesive side of the tape onto the cut surface at step 734, and the roller member is pressed down to adhere the adhesive tape so as to cover the entire cut surface at step 736. The linear actuator retracts at step 738 to reset the roller member for subsequent application of the adhesive. The microtome cuts the section covered by the adhesive tape at step 740. A photograph of the cut section covered by the tape is taken at step 742, and the cut section advances to the slide section at step 744 and aligns with the slide. Next, a determination regarding tissue adequacy is made at step 746. If the tissue section is not sufficient for transfer onto the slide, the section remains on the tape without being transferred to the slide at step 748. The tape then advances away from the slide at step 758 and is stored on the take-up mechanism. If the tissue section is sufficient for transfer onto the slide, the slide roller presses the section onto the slide at step 750, the section is laminated onto the slide at step 752, and a photograph of the section on the slide is taken at step 754. Finally, the slide roller retracts to its original position at step 756, and the tape advances away from the slide at step 758 and is stored on the take-up mechanism.

[0275] Referring to the flowchart of FIG. 31, after a tape, e.g., a tape cartridge, is loaded onto the feeding mechanism, the feeding mechanism 436 is activated to advance the tape, i.e., a continuous length of adhesive tape. The linear actuator member 438 is moved towards the cut surface of the sample block. Next, the roller member presses the adhesive side of the tape onto the cut surface. The roller member is then pressed down to adhere the adhesive tape so as to cover the entire cut surface. The linear actuator 438 is retracted to its original position to reset the roller for subsequent application of the adhesive tape to another sample. The microtome then cuts the section covered by the adhesive tape (along a plane parallel or substantially parallel to the cut surface). A photograph of the cut section is taken by a digital camera either at the time of transfer or immediately after transfer. The photograph is analyzed to confirm proper transfer to the tape. The cut section is advanced downstream to the slide station 440 and aligned with the slide. At this point, the photograph is analyzed to determine whether a sufficient section of the tissue has been cut from the sample block for transfer to the slide. If the section is not sufficient, e.g., if this results in containing not enough tissue sample, for example, mostly containing paraffin, the section is not transferred to the slide and remains on the tape. If the tape area contains a sufficient tissue section and is in a state where it can be transferred to the slide, the slide roller presses the section onto the slide, and the section is then laminated onto the slide by various methods described above. Photographs of the cut section and the slide are taken at the time of transfer to the slide or immediately after transfer. Note that the photograph can be taken before or after lamination onto the slide. Note that the slide from the slide machine has a barcode or other identification system corresponding to the barcode or other identifier on the sample block. The slide roller is retracted to its original position, the tape is advanced away from the slide, and is stored on the take-up reel of the tape cartridge mounted on the take-up mechanism. These steps of FIG. 31 are repeated until the desired number of sections from the sample block are cut by the microtome, transferred to the tape, and transferred to the slide.Photographs of each of these sections are taken as they are transferred to the tape and as they are transferred to the slide for analysis during the tape feeding operation (quality control).

[0276] The flowchart of FIG. 32 relates to a system similar to that of FIG. 31, except that a sample tape is removed from the carrier strip, as in the system of U.S. Publication No. 2017 / 0003309. The analysis of the photographs according to the flowcharts of FIGS. 31 and 32 is enhanced by the use of the illumination and visioning systems described herein.

[0277] As shown in FIG. 32, the feeding mechanism is activated in step 760 to advance a carrier strip that transports a patch of sample tape with an adhesive. Next, the sample tape is aligned in step 762 with the sample surface. In step 764, a roller moves to press the sample tape onto the sample surface, and the carrier strip guide is here in the application position. The roller retracts to its initial position in step 766. In step 768, the carrier strip guide moves to the removal position, moving the carrier strip out of the path and separating the carrier strip from the sample tape. A microtome cuts a section of the sample in step 770, and a photograph of the cut section on the tape is taken in step 772. Next, the sample tape with the attached section advances in step 774 to the slide station and aligns with the slide. Next, a determination of tissue adequacy is made in step 776. If the tissue section is not adequate for transfer onto the slide, the section remains on the sample tape in step 778 and is not transferred to the slide. If the tissue section is adequate for transfer onto the slide, a slide roller presses the section onto the slide in step 780, the sample tape is removed in step 782, and the section is laminated onto the slide in step 784. Finally, a photograph of the section on the slide is taken in step 786.

[0278] As discussed above, the automated system has an image-based quality control system for comparing the tissue on the block face of the sample block to the tissue section transferred to the glass slide. One or more imaging devices obtain a digital image of the block face, and one or more imaging devices obtain a digital image of the tissue section on the slide onto which the tissue section is transferred and immobilized, and the images of the sample block and the slide are compared to ensure that the items match. That is, the image from the sample block and the image from the slide containing the tissue section are compared to determine the presence or absence of a match. This provides a backup system to the barcodes provided on the cassette that transports the tissue block and on the slide. Thus, if the barcode match is confirmed, a double-check is performed by the automated device by image comparison. Therefore, quality control does not rely on human inspection.

[0279] To achieve such quality control, in some embodiments, three features are provided: 1) a series of image capture devices installed to capture the desired tissue image, 2) contrast is created on the digital image between the tissue and the paraffin (or other embedding medium) to improve the distinction and facilitate comparison / analysis, and 3) the tissue block and the slide are held steadily (firmly) to minimize or, in some embodiments, completely prevent movement of the block and the slide, enhancing imaging. Each of these features is discussed below.

[0280] The automated system can include a computer system for collecting images from a block-facing camera and a slide-facing camera, along with software for comparing the images from the block face and the slide. The software algorithm determines the outline of the tissue from each image and compares the two images. The images can be stored for later comparison if desired.

[0281] A computer system can have a decision algorithm for determining whether images from a block face and a slide match. The decision algorithm has knowledge of the identification of the sample block and the glass slide from which the images are then captured. The decision algorithm verifies that the sample also has a matching identification. The matching identification can be accompanied by a matching tissue contour as described herein. The matching identification can also be accompanied by matching barcodes on the sample block and on the slide, as shown, for example, in FIGS. 17A and 17B described herein.

[0282] Various types of imaging devices can be provided. The terms "imaging device" and "image capture device" are used synonymously herein and, for convenience, are discussed and shown from the perspective of a digital camera. However, it should be noted that various devices and methods for capturing images, including, for example, X-rays, infrared, tomography, micro-CT imaging, OCT cameras, etc., are also envisioned. A single image device can be provided, but alternatively, multiple imaging devices can be provided adjacent to the sample block and adjacent to the slide receiving the tissue section to enhance the image. It should be noted that an image capture device, such as a digital imaging device like a digital camera, can have an optical filter for incident light when the image capture device obtains an image of the block to enhance sharpness.

[0283] The imaging system can include a box containing the object to be imaged and the imaging hardware to prevent stray light from being captured by the camera.

[0284] In some embodiments, another aspect of having a sharp high-contrast image is to minimize the vibration between the image capture device and the object. The automation system can include a mechanism for stably holding a sample block of tissue to be sectioned in front of a plurality of cameras. This reduces the vibration of the sample block, thereby enhancing the image when using a camera sensitive to vibration, because such vibration blurs the image and makes tissue comparison more difficult, reducing the performance of the image post-processing tool. The sample block is held in a fixed position using a servo motor that monitors the position of the sample in real time. In one embodiment, the sample block is held in a fixed position using a high-inertia mounting bracket attached to the same reference frame as the image capture device. Other mechanisms for stably holding the sample block are also envisioned.

[0285] Similarly, to provide a sharp high-contrast image of the "second" image, i.e., the sectioned tissue section on the slide, the vibration between the image capture device and the slide is minimized. Thus, similar to stably holding the sample block as discussed above, the automation system can include a mechanism for stably (rigidly) holding the slide containing the sectioned tissue sample in front of a plurality of cameras. This stable holding reduces the vibration of the slide, thereby enhancing the image because such vibration blurs the image and makes tissue comparison more difficult, reducing the performance of the image post-processing tool. The slide can be rigidly held in a fixed position within the slide station by a slide holder during imaging. Alternatively, a mechanism such as a mounting bracket can be incorporated as part of the slide station.

[0286] The system and method of an automation system of one embodiment will now be described in conjunction with the flowchart of FIG. 33. The method described is an automated biological tissue sectioning device that processes biological tissue embedded with paraffin and produces thin sections on a glass substrate. The thin sections are suitable for microscopic analysis after further processing. In this method, the tissue shape on the block face is compared with the tissue contour on the glass slide. This is done in the context of a fully automated tissue sectioning device and provides an automated quality control capability. Thus, the present system provides an input and output product comparison for an automated biological tissue sectioning device, thereby providing quality control of the biological tissue that has been cut and placed on a glass slide.

[0287] As shown in FIG. 33, a photograph of the sample block is taken at step 790, and the feeder mechanism is activated at step 792 to advance the tape. The linear actuator moves toward the cut surface of the sample block at step 794. Next, the roller member presses the adhesive side of the tape onto the cut surface at step 796. The roller member is then depressed at step 798 to adhere the adhesive tape so as to cover the entire cut surface. The linear actuator retracts at step 800 to reset the roller member for subsequent application of the adhesive. The microtome cuts a section covered by the adhesive tape at step 802, the slide roller presses the section onto the slide at step 804, and the section is laminated onto the slide at step 806. An image of the section on the slide is taken at step 808. The slide roller retracts to its original position at step 810, and the tape advances at step 812 to move away from the slide for storage on the take-up mechanism. The section image is compared with the sample block image at step 814 to verify the match. Optionally, the comparison is made at step 815 between the baseline image of the tissue block and the section image, which can ensure the match between the tissue block and the tissue section. The barcode on the slide is compared with the barcode on the sample block at step 816 to verify the match.

[0288] More specifically, referring to the flowchart of FIG. 31, the steps are similar to the system described in conjunction with the flowchart of FIG. 31, except for the addition of quality control. The system of FIG. 31 also does not have a quality control step for determining whether the tissue should be transferred to the slide. However, the quality control system can be used in combination with a system that performs such steps. In the initial step of FIG. 31, one or more than one photograph of the sample block is taken by a digital camera. The initial photograph can be taken before or after activation of the feeding mechanism. The image is stored for comparison with an image taken after the process, i.e., after transfer to the slide (or transfer to the tape). Next, the method includes moving the tape towards the cutting surface of the sample block, pressing the adhesive tape onto the cutting surface to adhere the cutting surface to the surface (e.g., the entire surface of the cutting surface), cutting the tissue section through the microtome, advancing the cut tissue section on the tape to the slide station, pressing the tape section onto the slide, and stacking the sections on the slide. According to the quality control system, an image of the tissue section on the slide (before or after stacking) is taken by a digital camera. The sectioned image is compared with the sample block image to confirm matching. The barcode on the slide is also compared with the barcode on the sample block to confirm matching. This section comparison between the sample block and the tissue section on the slide is shown in FIG. 17A, and the photographed image of the upper part of the sample block containing the tissue embedded in the paraffin is compared with the tissue section on the slide. The barcode on each individual slide is compared with the barcode on the rear part of the sample block, as shown in FIG. 17B, which illustrates the slide and paraffin block identification comparison. Note that the barcode comparison can be performed before or after the tissue image comparison.

[0289] Note that the steps in the flowchart provide one embodiment for the use of a quality control system, and it is understood that the steps need not be performed in the exact order of the blocks in FIG. 33.

[0290] It should be understood that the quality control system described in conjunction with the method of FIG. 33 may include a quality control system that takes a photo of the tape when the tissue section is transferred thereto. The quality control system for identification matching for reliable verification can also be used without accompanying this tape section transfer photo check. The identification matching quality control system can be used in combination with any of the devices / systems and methods described herein.

[0291] It should be understood that a single camera or multiple cameras (or other imaging devices) can be used for sample blocks, cut sections, and / or slide images, and / or tape images. In addition, it is envisioned that either a single stationary or movable camera (or other imaging device) can be utilized to take images of the sample block, tape, and / or slide.

[0292] Note that multiple images of the sample block can be taken to provide a one-to-one comparison of the images of each slide. For example, before each section is cut, an image of the block face can be taken for comparison with the slide containing that specific cut section. Alternatively, only a single base image or some base images of the sample block can be taken for comparison with the images of each slide.

[0293] According to another aspect of the quality control system, the slide images are processed to check for bubbles or tears, i.e., to look for artifacts to ensure proper transfer of the cut tissue sections onto the slide. If bubbles are present, the slide can be discarded. At a second level of such an artifact quality control system, if an artifact is detected, the system will then determine whether the artifact is on the tissue or the paraffin. If it is on the tissue, the slide can be discarded; if it is on the paraffin, the slide can still be used as this will not affect the tissue analysis. In some embodiments, this artifact quality control system can be utilized in addition to the sample block / slide image comparison quality control system described herein.

[0294] As noted above, in various embodiments disclosed herein, it is envisioned that in one application, multiple sections can be transferred onto a single slide. Also, in some embodiments, it is envisioned that not all of the sections (or slides) will be stained. With regard to genetic analysis, since staining can destroy DNA, tissue tumor-specific sections are typically performed on blank or non-stained slides to preserve DNA. However, since non-stained slides are mostly transparent under the microscope, the contrast between normal tissue and tumor is very poor. In the system disclosed herein, the slide station can, in some embodiments, include alternating stained and non-stained slides. That is, a non-stained section (slide) is placed adjacent to a stained section (slide), and due to the tracking method disclosed herein, by detecting the positioning of the section on the tape and thus the slide, the non-stained slide can be genetically analyzed. Thus, the stained slide, which is nearly identical to the non-stained slide, will provide the region / coordinates for collecting material from the non-stained slide. This is achievable as typically the cut sections are 5 microns thick, which is about 1 / 2 the size of the cells.

[0295] As described herein, photographs are taken at various stages of the tape feed cycle for real-time analysis. Such photographs can be used in addition to, or as an alternative to, the optical sensors discussed above to provide feedback on the quality of the sections transferred to the tape and / or the quality of the sections transferred to the slide.

[0296] In some embodiments, any collected images and barcode associations can be synchronized with a Laboratory Information Management System (LIMS). When obtaining or making determinations of images / spectra, these images / spectra / determinations can enter the LIMS. Thus, the quality control system disclosed herein can facilitate such integration.

[0297] The automation system can include a computer system for collecting and analyzing the imaging data collected by the imaging system 2. The images can be stored for later analysis or comparison if desired. The computer system can have a decision algorithm for determining (in binary analysis) whether an image from a block face, tape, or slide indicates tissue abnormalities.

[0298] Any suitable computing system may be used to implement the computing devices and methods / functionality described herein, and as would be understood by one of ordinary skill in the art, may be transformed into a specific system for implementing the operations and features described herein through hardware, software, and modifications to the hardware in a manner that significantly exceeds merely the execution of software on a general purpose computing device. An exemplary embodiment of such a computing device 900 is depicted in FIG. 34. Computing device 900 is merely an exemplary embodiment of a suitable computing environment and in no way limits the scope of the invention. A "computing device" as represented by FIG. 34 may include, as would be understood by one of ordinary skill in the art, a "workstation", "server", "laptop", "desktop", "handheld device", "mobile device", "tablet computer", or other computing device. Assuming that computing device 900 is depicted for illustrative purposes, embodiments of the invention may utilize any number of different computing devices 900 in any number of different ways to implement a single embodiment of the invention. Thus, embodiments of the invention are not limited to a single computing device 900, as would be understood by one of ordinary skill in the art, nor are they limited to a single type of implementation or configuration of the exemplary computing device 900.

[0299] Computing device 900 can include a bus 910 that can be coupled directly or indirectly to one or more of the following exemplary components: a memory 912, one or more processors 914, one or more presentation components 916, input / output ports 918, input / output components 920, and a power supply 924. Those skilled in the art will understand that bus 910 can include one or more buses such as an address bus, a data bus, or any combination thereof. Those skilled in the art will also understand that, depending on the intended application and use of a particular embodiment, a plurality of these components can be implemented by a single device. Similarly, in some instances, a single component can be implemented by a plurality of devices. Accordingly, FIG. 34 is merely an illustration of an exemplary computing device that can be used to implement one or more embodiments of the present invention and is in no way intended to limit the present invention.

[0300] Computing device 900 can include or interact with various computer-readable media. For example, the computer-readable media can include random access memory (RAM), read-only memory (ROM), electrically erasable programmable read-only memory (EEPROM), flash memory or other memory technologies, CDROM, digital versatile disk (DVD), or other optical or holographic media, magnetic cassettes, magnetic tapes, magnetic disk storage devices, or other magnetic storage devices that can be used to encode information and can be accessed by computing device 900.

[0301] Memory 912 can include a computer storage medium in the form of volatile and / or non-volatile memory. Memory 912 can be removable, non-removable, or any combination thereof. Exemplary hardware devices are devices such as hard drives, solid state memories, optical disk drives, and the like. Computing device 900 can include one or more processors that read data from components such as memory 912 and various I / O components 920. Presentation component 916 presents data indications to the user or other devices. Exemplary presentation components include display devices, speakers, printing components, vibration components, and the like.

[0302] I / O port 918 can enable computing device 800 to be logically coupled to other devices such as I / O components 920. Some of the I / O components 920 can be built into computing device 900. Examples of such I / O components 920 include microphones, joysticks, recording devices, game pads, satellite television receiving antennas, printers, wireless devices, networking devices, and the like.

[0303] The above description contains many details, but those details should not be construed as limitations on the scope of the present disclosure, but rather should be construed merely as examples of its preferred embodiments. Those skilled in the art will envision many other possible variations that are within the scope and spirit of the present disclosure.

Claims

1. A method for sectioning a tissue block, comprising: imaging the tissue block to generate imaging data of the tissue block, wherein the tissue block comprises a tissue sample embedded in an embedding material; based on the imaging data, estimating a depth profile of the tissue block, the depth profile comprising a thickness of the embedding material to be removed to expose the tissue sample to a predetermined reference; the expected contour, expected size, and expected shape of the tissue sample after the thickness of the embedding material has been removed; and estimating; removing the thickness of the embedding material to expose the tissue sample to the predetermined reference; confirming, based on the imaging data, when the thickness of the embedding material has been removed to expose the tissue sample to the predetermined reference, by comparing the contour, size, or shape of the tissue sample in one or more sections cut from the tissue block with the expected contour, expected size, or expected shape of the sample estimated based on the imaging data; A method comprising.

2. progressively removing one or more sections from a tissue block comprising a tissue sample embedded in an embedding material; imaging the one or more sections to generate imaging data associated with the one or more sections; confirming, based on the imaging data, that the tissue sample is exposed to the predetermined reference; The method according to claim 1, further comprising.

3. The method according to claim 1, wherein the tissue block is imaged using structured light to determine the depth profile.

4. A method for sectioning a tissue block, comprising: prior to removing one or more sections from the tissue block comprising a tissue sample embedded in an embedding material, imaging the tissue block to generate baseline imaging data; estimating, from the baseline imaging data, the expected contour, expected size, or expected shape of the tissue sample at the expected exposure of the tissue sample to a predetermined reference; progressively removing one or more sections from the tissue block; Imaging one or more sections removed from the tissue block and generating imaging data associated with the one or more sections; Based on the imaging data, determining when a sufficient number of the one or more sections have been removed from the tissue block such that exposure of the tissue sample reaches the predetermined criteria; comprising; The determining includes comparing the contour, size, or shape of the tissue sample in the one or more sections with the expected contour, expected size, or expected shape of the tissue sample estimated from the baseline imaging data. A method.

5. The method according to claim 4, further comprising determining a depth profile of the embedding material from the baseline imaging data to remove a sufficient amount of the embedding material to expose the tissue sample to the predetermined criteria.

6. The method according to claim 4, further comprising illuminating the tissue block with structured light within the UV range.

7. The method according to claim 4, further comprising comparing imaging data of the section including the tissue sample with the baseline imaging data to determine when the tissue sample is sufficiently exposed.

8. The method according to claim 7, wherein the imaging data of the section comprises imaging data of the one or more sections on the tissue block, on a transfer medium, or on a slide.

9. The method according to claim 4, further comprising determining a depth profile by one or more of parallax, focus, or light field imaging.

10. The method according to claim 4, further comprising increasing the contrast between the tissue sample and the embedding material.

11. A method for sectioning a tissue block, comprising: Prior to removing the thickness of the embedding material from the tissue block comprising a tissue sample embedded in the embedding material, estimating the expected contour, expected size, or expected shape of the tissue sample when the tissue sample is exposed to a predetermined criterion; Removing the thickness of the embedding material configured to expose the tissue sample to the predetermined criterion from a tissue block comprising a tissue sample embedded in the embedding material; Subsequent to removing the thickness, progressively removing one or more sections from the tissue block; Based on a comparison of the contour, size, or shape of the tissue sample in the one or more sections with the expected contour, the expected size, or the expected shape of the tissue sample, confirming that the tissue sample is exposed to the predetermined standard; A method comprising.

12. A histological system, A microtome configured to progressively remove one or more sections from a tissue block, the tissue block comprising a tissue sample embedded in an embedding material; A vision system associated with the microtome, An illumination system configured to illuminate the tissue block comprising a tissue sample embedded in an embedding material; An imaging system configured to image the tissue block and generate imaging data associated with the tissue block; A processor in communication with the vision system, the processor being programmed to receive the imaging data and, prior to removing the one or more sections from the tissue block, determine the expected contour, size, or shape of the tissue sample from baseline imaging data generated by imaging the tissue block using structured light, and confirm that the tissue sample is exposed to a predetermined standard based on a comparison of the contour, size, or shape of the tissue sample in the one or more sections with the expected contour, the expected size, or the expected shape of the tissue sample; A vision system comprising A histological system comprising.

13. The histological system according to claim 12, wherein the processor is further programmed to determine when the tissue block is sufficiently sectioned by recognizing the amount of tissue sample exposed.

14. The histological system according to claim 12, wherein the illumination system is configured to illuminate the tissue block using structured light.

15. The histological system according to claim 12, further comprising a transfer medium for transferring one or more sections comprising a tissue sample from the tissue block to one or more slides, wherein the processor is further programmed to compare one or more sections on the tissue block, on the transfer medium, or on the one or more slides with baseline imaging data generated by imaging the tissue block using UV light prior to removing the one or more sections from the tissue block.

16. Estimating the expected contour, the expected size, or the expected shape of the tissue sample from the baseline imaging data includes determining the expected contour, the expected size, or the expected shape of the tissue sample at a plurality of depths of the tissue block, the method according to claim 4.

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