TW463042B - System for processing and imaging - Google Patents

Abstract

The invention features a system for processing a sample, including: (a) a container for storing and transporting the sample; (b) means for infiltrating and embedding the sample to form a block in the container; (c) means for sectioning the sample in the block; (d) means for imaging the sample in the block; and (e) means for identifying the sample during the processing of the sample.

Description

463042 V. Description of the invention (1) Background of the invention The present invention relates to histology for visible light and electronic display materials. The production of tissue samples and other materials of the machine, and transmission microscopy, have undergone a series of chemical treatments, resulting in—'normally by cultivating the sample ', the sample is embedded therein. The solid matrix (b 1 ck) is produced in the conventional method of tissue preparation. Aldehyde or other materials can be used as preservation samples. Avoid iron by first chemically fixing with formalin and glutaric acid to make the sample firm and select 2 = 5 (self Decomposition) of the material solution. The chemical fixation of the amphoteric nature, thereby enhancing the subsequent increase in the concentration of the solvent and the gradual replacement, is designed to melt the stone by using the solvent used, such as the alcohol in the product, to increase the penetration After that, followed by the external tissue, "," and then the sample is cooled to room temperature to solidify. This method penetrates the hard gel polymer and then heat, ultraviolet or other. This hardened, penetrating The tissue is then placed in a mold bl0Cl〇 ^ surrounded by paraffin or plastic to produce a tissue matrix (tissue f conventional tissue preparation methods used for standard optical and electron microscopy 7 ° σ thin sections are taken from the substrate Cut out from the block and transfer to glass slides or other supports & symbols. Stain the tissue to improve the contrast of the image, and then use the microscope clock T & Dimensional (χγ) image. Information about the one-dimensional structure in the tissue can be obtained by examining successive sections of the tissue. US Patent No. 4,960, 330 (herein incorporated by reference) illustrates a basic Block slicing Image acquiring system 'wherein Serial sections from 4 63 D42

The substrate is removed and the newly generated section of the substrate is microscopically developed. This method provides fast recording and storage of structured data: time and errors in individual tissue sectioning operations. Surface development can provide the advantages of three-dimensional reconstruction, and the production of two-dimensional images of χζ & γζ can also be easily obtained. Summary and simplification of the invention. (Client) The client's digital image can be modified by the client (that is, the invention takes into account the strength of the operation and development of tissue samples. In the system of the invention, clinicians and scientists send tissue samples to the processing center, which Obtain a slice-like screen hardcopy. In this system, a new digital image is generated from a digital image, or even a digital image from a data volume provided by the processing center. Therefore, including the penetration and placing of samples in the bag; and the hair set, and the method of infiltrating the hair, including the container in the container to form the first king of the present invention, the system that bites the silver U)-storage and transmission Sample container; (b) device buried in the container to form a base block in the container. Device for slicing the sample port; (d) showing sample 2 in the base block (e) identifying the sample during its processing Dress. 'The system may include a computer program for generating and storing digital samples for developing digital images. : Before loading and embedding, the sample is chemically fixed. The second form, which can also be described, provides a method for developing the sample: (a) providing a container suitable for storing the sample and fixing the sample; (0 infiltrating and embedding the sample in a base block; 切片 sectioning the sample in the base block;

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5. Description of the invention (3) An image of the sample is generated in the base block. Preferably, the method includes step (f) storing the image as a digital image; and step (g) developing the digital image. The term "sample 11" as used herein refers to the tissue or other material to be stained and embedded. The thickness of the sample can be greater than 200 microns, 1 mm, 1 cm or more. Immobilization refers to treatment with a chemical solution Tissue sample, which can be stored, hardened, and permeated. &Quot; Infiltration " refers to the treatment of tissue with one or a series of liquids that penetrate the entire tissue to the molecular level and then transform into solids so that The sample hardened. Embedding or embedding "refers to placing the infiltrated tissue in a mold and surrounding it with a substance (usually the same substance used for infiltrating tissue), and then the substance hardens to form a package. Of the block. So the role of the 'embedding substance' is to provide solid support and to make the sectioning process smooth. "Sectioning" refers to a thin slice cut from the substrate. "Staining" refers to the treatment of materials with colored or fluorescent substances. 'This colored or fluorescent substance can be combined with the material at the molecular level. "" Fluorescent C fluorescence) π or "darkfield (darkfi eld)" staining is by non-binding dyes (ie dyes that can exhibit natural fluorescence when stimulated by light of an appropriate wavelength), or binding dyes (I.e. molecules attached to a sample and attached directly or indirectly to a fluorescein). Examples of binding dyes include 'but not limited to: (i) primary antibodies linked to an antigen and comprising

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Molecules that are linked to and covalently bonded to the two primary antibodies of the primary antibody. The glory image, while the "standard," or "bright field," is tinted to create an image with a bright or white background. For fluorescent and standard microscopy. Surface development provides a wide range of power reductions and provides more data in conjunction with the cost of fixing materials. The method of this document (digital image format), this. Antibodies; and (ii) fluorescently stained black backgrounds are often treated and shaped with non-fluorescent dyes. It should be understood that the same dyes can automatically section and develop. Labor 'can reduce costs or provide a means for accumulating greater numbers without increasing materials and labor for the same invention as described and apparent from the following preferred embodiments. The features and advantages of the present invention can be more detailed in the patent scope. It has been found that the process of processing tissues and developing tissue samples is caused by the use of (i) a single container, which indicates the purpose and phase of the identification. Content: 个 of processing samples; IX and ⑴)-basic slice and image acquisition, the present invention provides a highly automated tissue processing and development. The system of the present invention includes the following general elements: a container for storing and transferring a sample; a device for infiltrating and embedding the sample to form a base block in the container, a device for slicing the sample in the base block; Means for developing the sample; and means for identifying the sample during its processing. Preferably, the system also includes a device for chemically fixing the sample before infiltration and embedding, although this is not necessary. In a particularly preferred embodiment, the system also includes: generating and storing digital images of the sample Devices; and

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463042 V. Description of the invention (5) Computer program for digital image development. The system of the invention and its compatibility with microscopy. The method of two microscopic examination of the base block 'lies on the base block surface, which can inspect the image, and the elimination of other material requirements can make the increase in the number of extra sections of slave disease collected from each sample. In addition, it promotes computer-assisted analysis of images and three-dimensional reproduction. The method used is a microscope that is superior to a standard bright field microscope for microscopy, especially for microscopes that produce high-quality biological tissues. It is not necessary to make glass tissue slides. The processing of this principle is fully automated, reducing the number of copies of each result. The image can be easily stored in digital format, such as the measurement of area or volume, or in the microscopy of the substrate surface. The unmodified bit-virtual slice captured is a dark field image derived from the colors emitted by the fluorescently stained sample. This sample contrasts with a black background, which is an opaque polymer that penetrates and embeds the sample. In contrast, in most surgical == laboratories and other medical-related diagnostic facilities based on microscopy ' The traditional transmission light microscope method is used to generate bright field images. 'Because thin sections of tissue and other materials are stained with standard non-fluorescence, and then illuminated with white or near-white light sources, the background is more than that of tissue. The image 'bright' is not darker. The original dark field image captured by the base surface can be converted and displayed into the image produced by the more familiar bright field microscopy. Such an image conversion method is described in, for example, USSN 09 / 311,789, which is incorporated herein by reference. Component description Container:

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4 63 042 V. Description of the invention (6) Place the tissue samples of interest in a container. The tissue processing steps are compatible, including storage, transmission, embedding, sectioning and development. For example, this container is used to prepare samples for sectioning and inspection. The shape of the matrix in which the tissue sample is embedded. Defining an extended cavity ' in the more mold ' includes--introduction of tissue samples outside of the daughter. This proximal end defines a suitable shape. The tip of the mold to one end defines a suitable polyhedron; this polyhedron is related to both the test shape and the image range and the tissue sample. Therefore, the end of the mold is preferably close to the image skirt and shape. This device It can be used to observe the new cut plane of this mold, so that the cross-section of the sample chamber transverse to the exposed surface of the sample chamber has a size or shape close to or similar to the field used to observe the surface. The area on the device where surface-related data can be recorded. The observed exposed surface of the block is not significantly larger than the shadow and is smaller than the imaging area. The imaging device provides this surface, that is, the true proportion of the imaged area is not dominated by the scanning magnification optics. Therefore, the size of the imaging field is large, and the shape of the imaging field is determined by the array, which is preferably fixed with the input, chemically fixed, and infiltrated. It can be used as a mold. Therefore, this mold will be a good tool. The openings are used for slicing the cavity to the size and shape of the slice to engage the safety device. The size and shape of the mold are compatible. The size of the imaging field is large. The length of the prepared tissue base block, or the imaging collar of the imaging device. The imaging field is in the image. What is desired is that, in the image field, there is no visible microscope image, and the array is also dominated by the array. Instead, it is dominated by the small and magnifying optical column design. It should be understood that the 'suitable container shape is selected based on the size of the tissue sample and the optimal imaging geometry. For example, a hole-like epidermal biopsy can produce a round epidermal sample with a diameter of approximately 4 mm. With 4. 5 X 4. 5 flat

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V. Description of the invention (7) The container of the square millimeter sample chamber can easily accommodate such a sample, which leads to a basic block having an image substantially corresponding to the imaging field. When the sample has an irregular shape, the shape of the sample chamber can be selected accordingly. For example, prostate needle specimens produce samples of approximately 1 mm in diameter and 17 mm in length. A preferred container for such a tissue sample is a sample chamber having the same opening > shape, for example, a rectangular cross section having a width of about 2 mm and a length of 20 mm and a length of 20 meters. In such an example, a linear CCD scan ^ can be used to generate a rectangular image area.

Although the quality of the section cut from such a long and thin substrate may be less suitable for inspection with traditional microscopy techniques, the quality of the exposed section is not affected. Therefore, when the mold base is used to connect with the automatic image recording device of U.S. Patent No. 4'9_60, 330, various container shapes can be used, not just the shape suitable for tissue development previously considered to obtain High quality development area. This container is made of polysiloxane or a container which is made of elasticity, including the destruction of the container in the slicing block. The method can include polypropylene, resulting in itself being made of any polystyrene. influences. Container materials are prepared during operation and removed containers are easily broken. Cut the desired volume in the mold. The choice of suitable materials can be reused ‘for example, gathered in the mold base. The cutting step on the material to be torn or torn from the container can be prepared from hard materials, for example, partly or discarded. Heavy siloxane. Can be removed from the container 'example. In the example of this example, the material is ‘polypropylene’, which is rejected by the solvent to be reused, and the material which is suitable for deafness (which is made of more expensive materials).

4 63 G, 12 V. Description of the invention (幻 '--- # In the preferred embodiment, the container is engraved with a thread, so that the container can be closed tightly. In addition, it can also be used removable (snap_ 〇n) lid or other leak-proof lid. Device for identifying samples during processing: The container is preferably labeled so that it can be clearly identified. There are a number of devices that can clearly distinguish the container from all other containers Come out; any device can be used in the present invention. In one embodiment, each container is marked with a unique barcode. Such container marking can be used to process a large number of samples and determine the sample. For chemical fixation, penetration and embedding Sample device: A standard tissue embedding method is used to embed a tissue sample in a mold base. Therefore, for example, the tissue is first chemically solidified with a standard fixing liquid to avoid autolysis (self-decomposition) of the sample, so that Tough and increase the permeability, thereby enhancing the penetration of subsequent solutions. Thereafter, water in the sample = is removed by gradually setting up with increasing concentrations of solvents (such as alcohol and xylene). Then The melted paraffin or plastic polymer is treated to infiltrate and produce a hardened perforated fabric. The previous step is to place the hardened infiltrated tissue in a sample chamber of the container after the container has been inserted. Slicing device: A strip knife blade typically includes a reciprocating discard rack supporting a tissue substrate bracket (and a knife bracket for a doctor blade and a reciprocating blade. A reciprocating strip Move the tissue mold base block: to the wedge base block safety device) move up and down, and move 'to remove the section of the base block. When the mold base block worships

V. Description of the invention (9) When connected with the automatic image recording device of US Patent No. 4,960, 330 (the complete content is incorporated herein as reference material), the tissue section cut from the base block is discarded or left behind To facilitate future use, as described in US Patent Nos. 5,7 4 6, 8 5 5 (which is incorporated herein by reference) and to view the storm surface of the block. With the microtome at the bottom, it can also be used to support an imaging device to observe the exposed side of the substrate. Developing device: A typical developing device that can be used in the mold base of the present invention, including zone arrays and linear charge linking devices (CCDs). CCD is a type of digital image detector that can provide °° -bitscape; digit image detector. It can be operated by computer and can be modified or copied electronically. The electronic storage and editing of images offers significant advantages over traditional photographic methods. The observation area of the area array scanner is a straight line area defined by a two-dimensional pixel array. The larger the number of pixels that define the image, the greater the resolution. Low-resolution area array CCDs can have 300x600 pixels, while high-resolution CCDs have 2000 × 2000-level pixels. Beta linear CCD scanners have only one or a few pixels . The complete image is obtained with a linear CCD (or sample) moving forward, as if the scanner was moving across the width of the sample. In both examples, the developed area defines a straight line area. More preferably, non-linear & CCD can be used. The selection of the size and shape of the sample chamber is determined based on the type of developing device of the image captured on the exposed surface of the base block. Therefore, the shape of the sample chamber is selected 'to avoid cavities in the development area or extension of the sample outside the development area. Avoid these possible situations because they reduce

ϋ63 042 V. Description of the invention (10) The amount of tissue samples (compared to the f-bit data (pixels) of the cavity should be taught. The larger the definition is, the less the number of pixels, the more The greater the resolution. The factor. The background in the heart area can improve the development of the sample. The device that generates and stores digital images: Format uses any storage medium and is stored in a digital output medium. Transmission 'For example, # by mail, such as, by ㈣ϋ electronically sent to the client's medium, such as sample: physical properties f: any type of sample, whether biological and autopsy :: two = Examples of samples include biopsies = industry; =: ;;;), = 1: mineral samples obtained, and polymer and plastic samples used in the automotive industry). It will be further explained in detail through the following examples. The example of bismuth is only used as an example, and is not intended to limit the examples of the present invention. Collect hole-shaped epidermal biopsies and place them in a container. Jf empty tkr ^ ^ ^, iiU ^ Dagger 4 shows no barcode and contains chemical solids. The container and fixative are provided by the processor. Provided (will: shadow person or company). Preferably, the processor also provides a chemical tincture, which can be provided in this container or in a second container

4 63 Ο 4 · 2. 5. Description of the invention (11) (and therefore transferred by the client to the first container). After placing it in the container, the client places a threaded lid on the container and inserts the number of bar codes and the sample content for subsequent identification and entrusts the sample to the processor. After that, scan the barcode into the computer with a suitable processor, and the computer will close the + digital image. Place the container in an automated sample processor (丨,

Leica LYNX automatic tissue processor; Wetzlar, Germany) Via a series of processing solutions (eg, dyes, alcohols that increase concentration, sample σ absorption, and / or dimethylbenzyl or other organic solvents) and combinations containing optical additives (Eg, opaque). 'Place the sample in the container' end sample chamber before the polymer hardens into a solid matrix. This can be easily achieved, for example, by centrifugation, and the orientation of the two samples relative to the slice plane is not important. Because the directivity of digital images can be presented by a computer during development. The polymer can be hardened by any suitable method (for example, heat, UV, time, etc.). Once hardened, the sample and substrate are fixed. On a microtome for slicing +. In a specific embodiment, the substrate is left in the container and the container is sliced along the embedded sample. In addition, the container may be completely or Partially (for example, 'a container with only adjacent tissue chambers) is removed. Then the sample is sliced and developed. Digital images of the sample are stored in the computer. After sectioning, the substrate can be discarded, or' if Leave the sectioned sample in the base block 'to store the base block for future use. The section itself can also be discarded or left behind (for example, for archival purposes). One method of collecting the section is described in the US patent 5, 746, 855.

Five 'invention description (12) In all places corresponding to the digital image of the sample, using analogue) and sent to electronically can press the computer file Yes, the person with the digital image can observe the image. Observe the image. Such a dimension, XZ or YZ other specific embodiments of the present invention. For example, this publication 0 9/1 54, 430 (the agent and the embedded polymer system can be integrated with Ren Ming Ke and the complete internal. A suitable client on the processing step is Is. Transfer to the contraction for more The content format is not because of this, the software is also flat. You need to cut the sample number of the bar code and the bar code to cut the image, in order to-in the better second medium disc, optical disc or _ Λ 〇, square to go, Line the digital image α as 'through the Internet', and send the digital image f. It should be understood that = ', which provides a commission with a device, can also provide client computer software to: provide further information (eg, three images, etc.). He Xizhi's Microscope Tissue Preparation Techniques Summary

The USSN is hereby incorporated in the United States as a reference material for reference. The present invention has been referred to the preferred specific embodiments, and is shown and described in detail. It is known to those skilled in the art that In addition to the spirit and scope of the invention as defined by the scope of the additional patent application, various changes can be made in form or details. Those skilled in the art will perceive or discover many specific aspects specific to the present invention using only routine experimental methods. Examples are the same type of examples. These same types of specific embodiments are included in the scope of patent applicationsa

Claims (1)

4 63 C: VI. Patent application scope L A system for processing samples, including: (a) —a container for storing and transferring the sample; a device for infiltrating and embedding the sample to form in the container—a block (c) a device for slicing the sample at the base; (d) a farm for developing the sample in the base; and (e) a device for identifying the sample during processing of the sample. And 2. The system as described in item 范围 of the patent application scope, wherein when the and the base block are cut > {, the base block is in the container. Wherein the system is more 9 Wherein the system is more 3 'The system as described in the scope of the patent application item 4 includes a device 4 for generating and storing digital images of the sample, as described in the patent application park item 3 The system includes a computer program for developing the digital image. 5. The system described in item i of the scope of patent application: on, including a device for chemically fixing the sample before infiltration and embedding. 6-A method for developing a sample, the method comprising: (a) providing a A container suitable for holding the sample; (b) chemically fixing the sample in the container; (c) infiltrating and embedding the sample to form a base block in the container; (d) placing the sample in the base block (E) Produce the sample in the base block. 7. As described in item 6 of the scope of patent application-Office 1 豕, ^ and the base block is sliced, the base block is in the method, where when the sample Among the plagues. 1084-3375-PF.ptd Page 16 6. Scope of Patent Application _ 8. The method described in item 6 of the scope of patent application, wherein the method further includes (f) storing the image as a digital image. 9. The method according to item 8 of the scope of patent application, wherein the method further comprises (g) developing the digital image. I 1084-3375-PF.ptd Page 17