US20100087333A1 - Process of fabricating tissue array block, process of fabricating tissue array sheet, tissue array block, tissue array chip, system of fabricating tissue array block and system of fabricating tissue array sheet - Google Patents

Process of fabricating tissue array block, process of fabricating tissue array sheet, tissue array block, tissue array chip, system of fabricating tissue array block and system of fabricating tissue array sheet Download PDF

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US20100087333A1
US20100087333A1 US12/449,865 US44986508A US2010087333A1 US 20100087333 A1 US20100087333 A1 US 20100087333A1 US 44986508 A US44986508 A US 44986508A US 2010087333 A1 US2010087333 A1 US 2010087333A1
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Prior art keywords
tissue
block
pieces
fabricating
tissue array
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English (en)
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Junya Fukuoka
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University of Toyama NUC
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University of Toyama NUC
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Assigned to NATIONAL UNIVERSITY CORPORATION UNIVER OF TOYAMA reassignment NATIONAL UNIVERSITY CORPORATION UNIVER OF TOYAMA ASSIGNMENT OF ASSIGNORS INTEREST (SEE DOCUMENT FOR DETAILS). Assignors: FUKUOKA, JYUNYA
Assigned to NATIONAL UNIVERSITY CORPORATION UNIVERSITY OF TOYAMA reassignment NATIONAL UNIVERSITY CORPORATION UNIVERSITY OF TOYAMA CORRECTIVE ASSIGNMENT TO CORRECT THE ASSIGNEE'S NAME PREVIOUSLY RECORDED ON REEL 023413, FRAME 0326. Assignors: FUKUOKA, JYUNYA
Publication of US20100087333A1 publication Critical patent/US20100087333A1/en
Priority to US13/845,195 priority Critical patent/US8551922B1/en
Abandoned legal-status Critical Current

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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00—Sampling; Preparing specimens for investigation
    • G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
    • G01N1/36—Embedding or analogous mounting of samples
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00—Sampling; Preparing specimens for investigation
    • G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
    • G01N1/286—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q involving mechanical work, e.g. chopping, disintegrating, compacting, homogenising
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00—Sampling; Preparing specimens for investigation
    • G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
    • G01N1/286—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q involving mechanical work, e.g. chopping, disintegrating, compacting, homogenising
    • G01N2001/2873—Cutting or cleaving

Definitions

  • the present invention relates to a process of fabricating a tissue array block; a process of fabricating a tissue array sheet; a tissue array block; a tissue array chip; a system of fabricating a tissue array block; and a system of producing a tissue array sheet that are used when examining or analyzing body tissue.
  • a tissue array chip having pieces of tissue disposed on a substrate has heretofore been used for the examination or analysis of body tissue.
  • the tissue array chip is used for the examination of presence or absence of diseased tissue, the analysis of a gene or protein, screening, etc. through application of a stain solution for specifically staining a test substance to the test substance.
  • FIG. 11 is an explanatory view illustrating a conventional process of fabricating a tissue array chip.
  • body tissue is formed into a tissue block that is then punched to collect cores.
  • the collected cores are inserted into holes arrayed on and formed in a base block.
  • the surface of the base block having the cores inserted into the holes is sliced on the order of several ⁇ m to fabricate tissue array sheets each having pieces of tissue disposed thereon.
  • a tissue array sheet is mounted on a substrate to fabricate a tissue array chip. Steps (c) and (d) are taken repeatedly to fabricate plural tissue array chips.
  • Patent Document 1 discloses a device equipped with a punch for a base block and a punch for a tissue block. The device uses the punch for the base block to form holes in the base block, punches the tissue block using the punch for the tissue block to collect cores and inserts the collected cores into the holes in the base block, thereby fabricating a tissue array block.
  • Patent Document 1 JP-A 2004-215667
  • tissue blocks have a variety of sizes and possibly have a small thickness from the beginning of collection. Furthermore, since the tissue blocks are used while being chipped off a number of times during the course of the examination or analysis thereof, they are gradually reduced in thickness. For these reasons, in fabricating a tissue array chip, there is a case where cores having a sufficient length cannot be obtained, resulting in extreme reduction in number of tissue array chips capable of being fabricated or failure to fabricate a tissue array chip.
  • the tissue blocks have different degrees of hardness depending on the kind of tissue. For this reason, situations have arisen, in which the insertion of a punch fails to core the tissue blocks and in which the punch is damaged. A technique of stably collecting cores from the tissue blocks with the punch requires a lot of skill.
  • FIG. 12 is a schematic cross section of a tissue block illustrating cores collected when exemplifying a paraffin-embedded tissue block, (b) is a schematic cross section of a tissue array block having the cores embedded therein, and (c) includes schematic plan views of tissue array chips fabricated from the tissue array block.
  • the tissue block is amorphous, cores collected from plural tissue blocks or from different places of a single tissue block have portions of tissue t irregular in length.
  • FIG. 13 is a schematic cross section of the tissue block illustrating cores collected when using the site of interests a as the target, (b) is a schematic cross section of a tissue array block having the cores embedded therein, and (c) includes schematic plan views of tissue array chips fabricated from the tissue array block.
  • the site of interests a contained in the tissue block is amorphous and, even when it is included in the surface layers of the cores, there is a case where it is not included in the lower layers of the cores.
  • FIG. 14 is an explanatory view illustrating the positional relationship between a site of interests a contained in the core on the tissue array chip and a site of interests a on the surface of the tissue block.
  • the present invention has been proposed in view of the aforementioned state of affairs and the object thereof is to provide a process of fabricating a tissue array block; a process of fabricating a tissue array sheet; a tissue array block and a tissue array chip; and a system of fabricating a tissue array block and a system of fabricating a tissue array sheet used for these, in which the tissue array block can be fabricated even in the case of that having a small thickness, is little affected by tissue hardness and, when having been processed into a tissue array chip, does not suffer any defect in a piece of tissue or at a site of interests, enables the collection of pieces of tissue from an area dispersed more than conventionally, also enables the establishment of positional relationship between the inside of a tissue piece and the inside of a tissue block, and scarcely affects the utilization of the tissue block remaining after collecting the pieces of tissue.
  • a process of fabricating a tissue array block according to the present invention comprises the steps of slicing a tissue block to obtain roll-shaped pieces of tissue and inserting the roll-shaped pieces of tissue in an axial direction into holes arrayed in a base block.
  • a process of fabricating a tissue array block according to the present invention comprises the steps of slicing a tissue block to obtain sheet-like pieces of tissue, coiling and rolling the sheet-like pieces of tissue to form roll-shaped pieces of tissue, and inserting the roll-shaped pieces of tissue in an axial direction into holes arrayed in a base block.
  • the tissue block is a paraffin-embedded tissue block or a frozen tissue block and may be any of tissue blocks insofar as it is capable of being sliced to obtain the aforementioned pieces of tissue.
  • the sheet-like pieces of tissue may be those obtained by slicing the entire surface of the tissue block or those obtained by slicing only a part of diseased tissue contained in the tissue block.
  • the maximum length of the fabricated roll-shaped pieces of tissue can be made equal to the width of the tissue block.
  • the thickness of the tissue block is small, it is possible to fabricate pieces of tissue having a sufficient length insofar as the width of the tissue block can be secured and to fabricate a considerable number of tissue array chips. Since the surface of the tissue block is sliced, the force required for the slicing step is smaller than that of the punching step concentrated at a single point, and the tissue block is little affected by tissue hardness. Since the pieces of tissue are collected in a small amount all over, the tissue block scarcely affects the utilization of the tissue block remaining after collecting the pieces of tissue.
  • the tissue array block is fabricated either by inserting in the axial direction the roll-shaped pieces of tissue after being cut into a predetermined length into the holes of the base block or by cutting the roll-shaped pieces of tissue into a predetermined length after being inserted in the axial direction into the holes arrayed in the base block. Therefore, in each of these processes, the pieces of tissue arrayed in the tissue array block are brought to a state in which they have the predetermined length. In addition, it is also possible to insert roll-shaped pieces tissue only existing in the site of interests. Furthermore, it is possible to increase the number of tissue array chips to be fabricated relative to the amount of pieces of tissue collected from the tissue block in comparison with the case where cylindrical cores are collected by punching in order to embed pieces of tissue sliced in a small thickness.
  • a process of fabricating a tissue array sheet according to the present invention is characterized in that it includes the step of slicing the tissue array block fabricated by the process of fabricating a tissue array block in a direction in which the pieces of tissue inserted into the holes have a spiral shape in cross section.
  • the tissue array sheet is fabricated by slicing the tissue array block fabricated by the aforementioned process in the direction in which the pieces of tissue inserted into the holes have the spiral shape in cross section, the tissue or site of interests of the tissue array sheet to be fabricated suffer no defect even when slicing the tissue array block from any position. Since the spiral pieces of tissue arrayed in the tissue array sheet are collected from a region moving down through the surface of the tissue block in a coiling and rolling direction, they are to be collected from the regions very dispersed in comparison with the cores collected by punching from respective single points.
  • a tissue array block of the present invention is characterized in that in the tissue array block having plural pieces of tissue retained on the base block, the pieces of tissue are roll-shaped.
  • the roll-shaped pieces of tissue are used, it is possible to fabricate the same by slicing the tissue block even when the tissue block has a small thickness.
  • By adjusting the length of the roll-shaped pieces of tissue it is possible to make the lengths of the retained pieces of tissue equal to one another.
  • a tissue array chip of the present invention is characterized in that in the tissue array chip having plural pieces of tissue arrayed on the substrate, the pieces of tissue have a spiral (helical) shape.
  • the spiral (helical) pieces of tissue are to be collected from regions dispersed from respective starting points of spirals to respective terminal points thereof.
  • a system of fabricating a piece of tissue according to the present invention comprises means for slicing a tissue array block to obtain a sheet-like piece of tissue and means for coiling and rolling the sheet-like piece of tissue obtained by the slicing means to obtain a roll-shaped piece of tissue.
  • tissue block is sliced by the slicing means to form a sheet-like piece of tissue and since the sheet-like piece of tissue is coiled and rolled by the coiling and rolling means to form a roll-shaped piece of tissue, roll-shaped pieces of tissue can be fabricated from the tissue block.
  • a system of fabricating a tissue array block comprises means for slicing a tissue block to obtain roll-shaped pieces of tissue, means for cutting the roll-shaped pieces of tissue at predetermined positions and means for inserting the roll-shaped pieces of tissue in an axial direction into holes arrayed in a base block.
  • the positions, at which the roll-shaped pieces of tissue are cut by the cutting means can be set.
  • the position of cutting by the cutting means is set to be a position at which the roll-shaped pieces of tissue are cut in the axial direction in a predetermined length and, after the roll-shaped pieces of tissue are cut in the predetermined length by the cutting means, the pieces of tissue are inserted by the inserting means into the holes of the base block, thereby fabricating the tissue array block of the present invention.
  • the tissue array block of the present invention can be fabricated.
  • the cutting positions may be controlled with a position sensor, by an action stored in advance in a microcomputer, or manually.
  • a system of fabricating a tissue array sheet according to the present invention comprises the aforementioned system of fabricating a tissue array block and means for slicing the tissue array block fabricated by the system of fabricating a tissue array block in a direction in which the pieces of tissue have a spiral shape in cross section.
  • the tissue array block is fabricated by the aforementioned tissue array block-fabricating system and since the tissue array block can be sliced by the slicing means in the direction in which the pieces of tissue have a spiral (helix) shape in cross section, it is possible to fabricate a tissue array sheet having spirally shaped pieces of tissue arrayed.
  • the maximum length of pieces of tissue to be inserted into holes of the tissue block can be made equal to the width of the tissue block, insofar as the tissue block has a sufficiently large width, even when the thickness of the tissue block has a small thickness from the beginning or the thickness of the remaining tissue has been reduced during the course of the examination or analysis, it is possible to fabricate a considerable number of tissue array chips. Since the pieces of tissue are fabricated by the step of slicing the surface of the tissue block, the tissue block is little affected by tissue hardness in comparison with the punching step requiring a force concentrated at a single point. In comparison with the punching step requiring subtle adjustment of a force depending on the tissue hardness, a technique requiring a lot of skill is unnecessary to conduct, and the pieces of tissue can be fabricated by a comparatively simple operation.
  • the lengths of the pieces of tissue arrayed on the tissue array block can be made equal. It is also possible to insert roll-shaped pieces of tissue composed only of the site of interests. Since the sheet-like pieces of tissue are embedded in a state of having been rolled, the number of tissue array chips to be fabricated relative to the amount of tissue collected from the tissue block can be increased in comparison with the conventional punching step.
  • the process of the present invention for fabricating a tissue array sheet since the lengths of the pieces of tissue retained on the tissue block are made equal, even when the tissue array block is sliced from any position, the tissue or site of interests of the tissue array sheet does not suffer any defect. As a result, it is possible to prevent a situation constituting an obstacle to the examination or analysis and enhance the yield of the tissue array chips to be fabricated. Since the spiral pieces of tissue arrayed on the tissue array sheet are collected from the dispersed regions moving down through the surface of the tissue block in a coiling and rolling direction, the reliability of the examination or precision of the analysis is heightened.
  • tissue array block of the present invention it is possible to fabricate a tissue array block using the sheet-like pieces of tissue obtained through slicing the tissue block. It is possible to fabricate a considerable number of tissue array chips using the tissue block even when the tissue block has a small thickness. By adjusting the lengths of the roll-shaped pieces of tissue, it is possible to make the lengths of the retained pieces of tissue equal, thereby fabricating tissue array chips having the piece of tissue or site of interests suffering no defect.
  • the spiral pieces of tissue are to be collected from the dispersed regions corresponding to those from the starting point to the terminal point of the spiral.
  • the roll-shaped pieces of tissue can be fabricated and inserted into the tissue block to make it possible to fabricate the tissue array block of the present invention.
  • the system of the present invention for fabricating a tissue array sheet it is possible to fabricate a tissue array sheet having spiral pieces of tissue arrayed.
  • the tissue array chips of the present invention can be fabricated using the tissue array sheet.
  • a tissue block B 1 is a paraffin-embedded tissue block or frozen tissue block, for example, and is not limitative insofar as a block can be sliced into sheets.
  • a description will be made using a tissue block B 1 embedded in the neighborhood of the center of paraffin as an example. However, the same description will be applicable to the case using frozen tissue.
  • the tissue block B 1 is sliced to fabricate a piece of tissue t 2 having a sheet-like piece of tissue coiled and rolled into a roll shape ( FIGS. 1 ( a ) and ( b )).
  • the surface of the tissue block B 1 is sliced together with the surrounding paraffin using slicing means, such as a cutter.
  • a portion of the tissue block sliced and substantially simultaneously formed into a sheet is coiled and rolled ( FIG. 1( a )), with the front surface thereof as an inside surface, and upon completion of the slicing step, formed into a roll shape ( FIG. 1( b )).
  • tissue block when a tissue block is sliced in a small thickness, the portion thereof sliced is sequentially curved, with the front surface as an inside surface and, when being continuously sliced as it is, is coiled and rolled into a roll shape.
  • the present embodiment utilizes this feature.
  • the piece of tissue t 2 has a hollow core and assumes a substantially cylindrical shape, a step of densely coiling and rolling step may be added in order to heighten the integration degree of tissue.
  • the roll shape used herein indicates a shape of the sheet-like piece of tissue t 1 coiled, with one end thereof as an axis, and though the piece of tissue t 2 preferably has a spiral (helical) cross section orthogonal to the axis thereof over the entire length thereof, it may have at least part thereof spiral in cross section.
  • the thickness of the piece of tissue T 1 is not particularly limited insofar as the thickness is capable of coiling and rolling the piece of tissue as by desiccation into a roll shape.
  • the thickness of the piece of tissue t 1 is 200 ⁇ m or less (excluding 0 ⁇ m), preferably 30 ⁇ m to 200 ⁇ m, more preferably 50 ⁇ m to 100 ⁇ m.
  • the thickness is larger than 200 ⁇ m, the piece of tissue t 1 induces a trace of crack to possibly mar the shape thereof, whereas when it is less than 30 ⁇ m, the piece of tissue t 1 induces a defect when being sliced and, when having been formed into a tissue array chip, the user-friendliness in an examination is possibly hindered.
  • tissue block B 1 is sliced at the slicing step.
  • the sheet-like piece of tissue t 1 obtained at the slicing step is coiled and rolled into a roll shape at a forming step.
  • the roll-shaped piece of tissue t 2 obtained at the slicing step of the first process may be stretched and then densely coiled and rolled into a roll shape.
  • the integration degree of tissue can be heightened.
  • a piece of tissue sliced in a thickness to an extent incapable of being coiled and rolled to assume a planar shape or a slightly curved shape may be formed into a roll shape.
  • a force required and an influence by tissue hardness are smaller in comparison with the case of punching requiring a force to be concentrated in a single point.
  • the slicing means is seldom damaged by and seldom fails to slice a very hard tissue block, and does not necessitates considering a technique requiring a lot of skill as has been required in the punching step requiring the adjustment of a force depending on the hardness of the tissue block.
  • tissue block is used for various examinations and analyses after the pieces of tissue t 1 are collected from it, since very thin sheet-like pieces of tissue t 1 are collected from the surface of the tissue block B 1 over all, there is little influence on the utilization of the tissue block from which the pieces of tissue have been collected, as compared with the case of the punching step of punching a single point in the width direction.
  • the roll-shaped piece of tissue t 2 obtained by the piece of tissue-fabricating process has a maximum length in the direction of an axis b that becomes equal to the width of the tissue block B 1 .
  • the tissue block B 1 has a sufficient width even when it has a thin thickness, it is possible to obtain a piece of tissue t 2 having a length capable of fabricating a considerable number of tissue array chips.
  • a pre-step of inserting a cut into the tissue block B 1 so as to surround the site of interests and a post-step of slicing may be performed to fabricate a piece of tissue t 2 composed only of the site of interests.
  • the tissue array block-fabricating process P 2 is divided into two.
  • the first process P 2 - 1 includes a cutting step ((c 1 ) in FIG. 1 ) and an inserting step ((d 1 ) in FIG. 1 ) as post-steps of the above piece of tissue-fabricating process P 1 .
  • the roll-shaped piece of tissue t 2 is cut in the direction of the axis b so as to have a predetermined length ((c 1 ) in FIG. 1 ).
  • the cutting position may be adjusted so that the piece of tissue t 3 may have a desired length and site.
  • the length of the pieces of tissue t 3 cut is made equal to or larger than the axial length of holes h arrayed in a base block B 2 so that the pieces of tissue t 3 may be filled in the holes h without any gap.
  • the piece of tissue t 2 includes the site of interests, it may be cut to extract the site of interests. In this case, the site of interests on the surface of the tissue block B 1 is first specified and the piece of tissue t 2 is then cut so as to extract a site corresponding to the specified site of interests.
  • the base block B 2 is a block having plural cylindrical holes h arrayed therein. These holes h are formed as by punching at a separate step.
  • the base block B 2 may be formed of a material having a surface sliceable into a sheet and, for example, is a paraffin block.
  • the pieces of tissue t 3 obtained from different tissue blocks B 1 or the same tissue block B 1 may be inserted into the holes h, respectively.
  • the second process P 2 - 2 includes, as post-steps of the piece of tissue-fabricating process P 1 , a pre-step ((c 2 ) in FIG. 1 ), a subsequent inserting step ((d 2 ) in FIG. 1 ) and a cutting step ((e 2 ) in FIG. 1 . Only points of the second process different from the first tissue array block-fabricating process will be described hereinafter.
  • the pieces of tissue t 3 are inserted in the direction of the axis b into the holes h of the base block B 2 ((d 2 ) in FIG. 1 ).
  • the pieces of tissue t 3 inserted into the holes h are cut in the direction of the axis b to have the predetermined length ((e 2 ) in FIG. 1 ).
  • the cutting position is set to be the front surface position of the base block B 2 , and the pieces of tissue t 3 cut are preferably filled in the holes h.
  • the pieces of tissue t 3 are inserted into the base block B 2 , they are preferably fixed to the base block B 2 .
  • a step of heating the base block B 2 to dissolve paraffin or filling a paraffin melt into the holes h and a subsequent step of solidifying the paraffin are taken.
  • a tissue array block B 3 is fabricated by the first tissue array block-fabricating process or the second tissue array block-fabricating process ( FIG. 1( f )).
  • the tissue array block B 3 has the roll-shaped pieces of tissue t 3 , which are obtained by coiling and rolling the sheet-like pieces of tissue, retained in the holes h, . . . , h arrayed in the base block B 2 . Since the pieces of tissue t 3 are cut to the predetermined length at the cutting step, the pieces of tissue t 3 arrayed in the tissue array block B 3 are adjusted to have an equal length.
  • the pieces of tissue t 3 are cut in a length equal to or larger than the depth of the holes h is further preferred because the pieces of tissue t 3 are filled in the holes h.
  • the very thin sheet-like pieces of tissue t 1 are embedded in the form of the roll shape, it is possible to increase the number of tissue array chips C 1 produced relative to the amount of the pieces of tissue collected from the tissue block B 1 in comparison with the case where cylindrical cores are collected by punching.
  • FIG. 2 includes explanatory views illustrating a tissue array sheet-fabricating process P 3 and a tissue array chip-fabricating process P 4 of the present embodiment.
  • the array sheet-fabricating process P 3 includes, as a post-step of the tissue array block-fabricating process, a tissue array block-slicing step ( FIG. 2( a )).
  • the tissue array block B 3 fabricated by the tissue array block-fabricating process P 2 - 1 or P 2 - 2 is sliced in a direction in which the pieces of tissue t 3 are spiral (helical) in cross section, i.e. in a direction intersecting the axis b of the pieces of tissue t 3 ( FIG. 2( a )).
  • a tissue array sheet S 1 having plural spiral (helical) pieces of tissue t 4 arrayed thereon is fabricated ( FIG. 2( b )).
  • a tissue array chip-fabricating process P 4 includes a step of mounting the tissue array sheet S 1 on a substrate of a glass slide or nylon film ( FIG. 2( c )). As a result, a tissue array chip C 1 having the spiral pieces of tissue t 4 arrayed thereon is fabricated.
  • FIG. 3( a ) is an explanatory view illustrating the comparison between the state in which tissue array chips C have been fabricated by the conventional punching step from a tissue array block B and the state in which tissue array chips C 1 have been fabricated from the tissue array block B 3 of the present embodiment, and (b) an explanatory view illustrating the comparison between the states in which the tissue array chips C and C 1 composed of sites of interests a have been fabricated respectively from the tissue array blocks B and B 3
  • the portions of tissue t and sites of interests a contained in the cores have different lengths, there are defects of the pieces of tissue or sites of interests in the tissue array chip C depending on positions L 1 to L 3 at which the slicing step is taken.
  • tissue array block B 3 of the present embodiment since the lengths of the pieces of tissue t 3 or sites of interests a are made equal, there is no defect of the pieces of tissue or sites of interests in the tissue array chip C 1 even when the slicing step is taken at any of the positions L 1 to L 3 .
  • FIG. 4( a ) is a diagram illustrating the positional relationship between a core on the tissue array chip C and a collection region d of the tissue block B 1 according to the conventional punching step, and (b) a diagram showing the positional relationship between a spiral piece of tissue t 4 on the tissue array chip C 1 and a collection region d of the tissue block according to the present embodiment.
  • the core is collected from the very lopsided region d.
  • the spiral piece of tissue t 4 is collected from the dispersed region d transecting the tissue block B 1 , it is possible to heighten the reliability in an examination or the precision of analysis in comparison with the conventional technique of single-point punching.
  • the positional relationship between the inside position of the core and the inside position of the collecting region d of the tissue block B 1 is unclear. For this reason, even when the site of interests a exists in the core, for example, it is difficult to specify where the site of interests a is positioned in the tissue block B 1 .
  • the positional relationship among the piece of tissue t 4 on the tissue array chip, the piece of tissue t 3 after the cutting step, the piece of tissue t 2 before the cutting step and the tissue block B 1 is grasped.
  • An arrow A in the figure indicates the coiling and rolling direction at the roll-forming step, and the inside end d (start) of the piece of tissue t 4 is on the coiling and rolling starting side and the outside end d (end) thereof is on the coiling and rolling terminating side.
  • a tissue array chip C 1 is fabricated by any of the processes described above using a tissue block B 1 constituting a test object. Since a piece of tissue t 2 contained in the tissue block B 1 is collected from a dispersed region from one end to the other end of the tissue block, the reliability in the examination can be heightened in comparison with the prior art using the single-point punching step.
  • a site of interests has been found in a piece of tissue t 4 contained in the tissue array chip C 1 , by causing the piece of tissue t 4 and tissue block B 1 to correspond in position to each other, it is possible to specify the position of the site of interests in the tissue block B 1 .
  • a pretreatment comprises using the piece of tissue obtained by slicing the surface of the tissue block B 1 to fabricate a tissue chip and staining the site of interests to specify the position thereof.
  • the tissue chip and tissue block B 1 are contrasted with each other to specify a region corresponding to the site of interests in the tissue block B 1 , and a cut is inserted so as to surround the specified region.
  • the tissue block-slicing step and roll-forming step are then taken to fabricate a roll-shaped piece of tissue t 2 .
  • the piece of tissue t 2 is composed only of the site of interests.
  • the cutting step and inserting step are taken using the piece of tissue t 2 to fabricate a tissue array block B 3 . Since the tissue array block B 3 has filled therein a piece of tissue t 3 composed of the site of interests, it is possible to prevent a defect of the site of interests when fabricating a tissue array chip C 1 . It is also possible to cause the inside position of the site of interests in the tissue block B 1 to correspond to the inside position of a piece of tissue t 4 . Since the site of interests is collected from the dispersed region, the heterogeneity thereof is covered to enhance the degree of analysis.
  • the procedure thereof may comprise causing the tissue chip and roll-shaped piece of tissue 12 at the pretreatment to correspond to each other and cutting the piece of tissue t 2 so as to extract a site corresponding to the site of interests.
  • FIG. 5 is an explanatory view illustrating a piece of tissue-fabricating system Sys 1 according to the present embodiment.
  • the piece of tissue-fabricating system Sys 1 is suitable for fabricating the roll-shaped piece of tissue t 2 .
  • the piece of tissue-fabricating system Sys 1 is provided with at least slicing means 1 and roll-forming means 2 .
  • the slicing means 1 has a function to slice the surface of the tissue block B 1 to fabricate the roll-shaped piece of tissue t 1 and is a cutter, for example.
  • the slicing means 1 is configured so that the surface of the tissue block B 1 may be sliced in a desired thickness in accordance with a relative change in position of the tissue block B 1 .
  • the slicing means 1 is attached and fixed to a fixing member 1 a and, by adjusting the fixing position or fixing angle, it is possible to adjust the thickness of the roll-shaped piece of tissue t 1 .
  • the tissue block B 1 is mounted on conveying means 3 for conveying the tissue block B 1 and, by sliding the tissue block B 1 relative to the slicing means 1 , the surface of the tissue block B 1 is sliced.
  • the slicing means 1 may be slid, with the tissue block B 1 fixed.
  • tissue block B 1 since a paraffin-embedded tissue block is used as the tissue block B 1 , when the tissue block B 1 is sliced with the slicing means 1 , the sheet-like piece of tissue t 1 is coiled and rolled, with the front surface thereof as an inside surface.
  • a rod-shaped guide member 4 is disposed in the vicinity of the slice means 1 and along the axial direction of the slice means 1 , and the sheet-like piece of tissue formed by slicing is allowed to sequentially twine around the guide member 4 .
  • the guide member 4 is fixed with a fixing member not shown so as to make the relative position thereof constant relative to the slice means 1 .
  • roll-forming means may be further provided for densely coiling and rolling the piece of tissue U.
  • the roll-forming means is materialized, for example, through making it possible to rotate the guide member 4 and adjust the temperature of the guide member.
  • the piece of tissue t 1 is adhered to the guide member 4 by elevating the temperature of the guide member upon the end of the sheet-like piece of tissue t 1 coming into contact with the guide member to dissolve paraffin and then lowering the temperature to solidify the paraffin and, in that state, the guide member 4 is rotated to densely coil and roll the piece of tissue t 1 .
  • It may also be adopted to elevate and lower the temperature of the guide member again at the time of the completion of coiling and rolling to dissolve and solidify the contained paraffin, thereby making the roll shape stable.
  • FIG. 6 includes explanatory view illustrating the movement of the piece of tissue-fabricating system Sys 1 .
  • the conveying means 3 conveys the tissue block mounted thereon to the side of the cutting means 1 ( FIG. 6( a )).
  • the cutting means 1 slices the conveyed tissue block B 1 to fabricate the sheet-like piece of tissue t 1 ( FIG. 6( b )).
  • the slicing means 1 causes the fabricated piece of tissue t 1 to sequentially twine around the guide member 4 to fabricate the roll-shaped piece of tissue t 2 , with the guide member 4 as an axis ( FIG. 6( c )). Thereafter, the piece of tissue may be densely coiled and rolled.
  • FIG. 7 is an explanatory view illustrating a tissue array block-fabricating system Sys 2 according to the present embodiment.
  • the tissue array block-fabricating system Sys 2 includes cutting means 5 and inserting means 6 in addition to the means in the piece of tissue-fabricating system Sys 1 .
  • the cutting means 5 has a function to cut the roll-shaped piece of tissue t 2 at a set position.
  • the cutting means 5 is a cutter having a cutting edge on the side of the guide member 4 .
  • the guide member 4 slides in a direction intersecting the axial direction of the guide member 4 so as to be movable in the axial direction of the guide member 4 and can be controlled in position based on information or a program from a sensor.
  • the inserting means 6 has a function to insert the roll-shaped pieces of tissue t 2 in the axial direction into the holes h arrayed on the base block B 2 .
  • the inserting means 6 comprises slide means 6 a for sliding along the guide member 4 the roll-shaped pieces of tissue t 2 coiled and rolled on the guide member 4 and positioning means 6 b for retaining the base block B 2 and positioning the same in three directions.
  • the slide means 6 a is a cylindrical member into which the guide member 4 is inserted and can be controlled so as to slide to a predetermined position along the guide member 4 .
  • the positioning means 6 b can fix the base block B 2 to a surface on the side of the guide member 4 and has a function to position the base block B 2 in the X-, Y- and Z-directions based on the information or program from the sensor.
  • FIG. 8 includes explanatory views illustrating how the tissue array block-fabricating system Sys 2 moves.
  • the piece of tissue t 2 is brought to a state in which it has been coiled and rolled on the guide member 4 ( FIG. 8( a )).
  • the sliding means 6 a slides along the guide member 4 to move the piece of tissue t 2 to a set position by means of a surface 6 c in contact with the piece of tissue t 2 ( FIG. 8( b )).
  • the set position is a position at which a paraffin portion at the end of the piece of tissue t 2 projects from the distal end of the guide member 4 .
  • the cutting means 5 is positioned and then slid to cut the projecting portion of the piece of tissue t 2 ( FIG. 8( c )). As a result, the paraffin portions are cut off to align the ends of the pieces of tissue t 2 .
  • the positioning means 6 b causes a desirable hole h to face the guide member 4 and moves in the direction of the guide member 4 to insert the piece of tissue t 2 into the desirable hole h ( FIG. 8( d )).
  • the cutting means 5 is slid to cut the piece of tissue t 2 ( FIG. 8( e )). As a result, a state is obtained in which the cut piece of tissue t 3 is inserted into the hole h ( FIG. 8( f )).
  • tissue array block-fabricating systems Sys 2 are disposed in parallel, and a single positioning means 4 b is prepared for the tissue array block-fabricating systems Sys 2 .
  • the positioning means 4 b is used to position the base blocks B 2 relative to the tissue array block-fabricating systems Sys 2 , thereby causing the tissue array block-fabricating systems Sys 2 to function in the same manner as described above.
  • tissue array block-fabricating system Sys 2 is for realizing the second tissue array block-fabricating process, is may be used as a system for realizing the first tissue array block-fabricating process.
  • the system is further provided with grasping means for grasping the piece of tissue t 3 cut by the cutting means 5 and, after the piece of tissue t 2 is cut by the cutting means in a predetermined length, the cut piece of tissue grasped by the grasping means is to be inserted into the base block B 2 .
  • a separate punching mechanism is used to form holes h in the base block B 2
  • the positioning means 6 b is used to position the holes h and the pieces of tissue t 3 .
  • the positioning step may be simplified.
  • the tissue array block-fabricating system Sys 2 is preferably provided with fixing means for fixing the pieces of tissue t 3 having been inserted into the holes h to the base block B 2 .
  • the fixing means has a function to heat and cool the base block B 2 , with the pieces of tissue t 3 inserted into the holes, or a function to fill and solidify molten paraffin, for example.
  • the positioning means 6 b is provided with a temperature adjusting function to adjust the temperature in a state wherein the base block B 2 is retained so as to bring the positioning means into contact with one surface of the base block B 2 , thereby heating and cooling the base block B 2 .
  • a tissue array sheet-fabricating system Sys 3 is provided with slicing means 7 for slicing the tissue array block B 3 in addition to the means of the tissue array block-fabricating system 2 .
  • FIG. 9 includes explanatory views functionally illustrating the tissue array sheet-fabricating system Sys 3 .
  • the slicing means has a function to slice the tissue array block B 3 in the direction in which the pieces of tissue t 3 having been inserted into the holes have a spiral shape in cross section.
  • the slicing means 7 is a cutter, for example, having a cutting edge of a width substantially the same as the width of the tissue array block B 3 .
  • the position or angle at which the slicing means comes into contact with the tissue array block B 3 is adjustable and, by adjusting it, it is possible to adjust the thickness of the tissue array sheet t 4 .
  • the tissue array block B 3 Upon completion of the fabrication of the tissue array block by the tissue array block-fabricating system sys 2 , the tissue array block B 3 is in a state in which it has been mounted on the positioning means 7 ( FIG. 9( a )).
  • the positioning means 7 By positioning the pieces of tissue t 3 in the tissue array block B 3 by the positioning means 7 so as to bring the surfaces of the pieces of tissue arrayed into contact with the slicing means 7 and sliding the sliding means to the side of the slicing means 7 , the surfaces of the pieces of tissue t 3 arrayed on the tissue array block B 3 are sliced ( FIG. 9( b ).
  • it is possible to fabricate a tissue array sheet S 1 FIG. 9( c )).
  • a tissue array chip C 1 is fabricated.
  • the fabricating method of the above embodiment was performed to fabricate a tissue array chip.
  • the tissue used was mesothelial tumor tissue and was embedded in paraffin to fabricate a paraffin-embedded tissue block that was used as a tissue block.
  • a paraffin block was formed therein with holes h to use the resultant block as a base block.
  • a tissue block was sliced by a cutter to fabricate a sheet-like piece of tissue t 1 and, at the same time, the piece of tissue t 1 was coiled and rolled to fabricate a roll-shaped piece of tissue t 2 .
  • the thickness of the sheet-like piece of tissue t 1 was set to be about 50 to 100 ⁇ m in view of the easiness of formation and observation, and the diameter of the roll-shaped piece of tissue t 2 was set to be about 1 mm to 2.5 mm from the standpoint of the integration degree in the tissue array chip.
  • the roll-shaped piece of tissue t 2 was cut in a length equal to the depth of the holes to fabricate a piece of tissue t 3 .
  • tissue array block having plural pieces of tissue t 3 collected from the different tissue blocks.
  • the tissue array block was mounted on a glass slide to form tissue array chip that was then subjected to immunostaining.
  • FIG. 10( a ) is a diagram showing a tissue array chip according to the present example, and ( b ) a diagram showing a tissue array chip obtained by the conventional punching step.
  • the pieces of tissue had to possess an array configuration in the conventional tissue array chip, there were very many defects of pieces of tissue in portions surrounded by undulating lines in the figure.
  • the pieces of tissue had to possess a circular shape, there were many pieces of tissue partially broken as shown by an arrow.
  • a tissue array chip that had plural spiral pieces of tissue t 4 having an array configuration was obtained.
  • the pieces of tissue t 4 emerged in the tissue array chip without being substantially broken and, therefore, a very high effect was obtained as compared with the conventional process of collecting cores by punching.
  • FIG. 10( c ) is an enlarged view of pieces of tissue t 4 on the tissue array chip fabricated by the present example, and ( d ) an enlarged view of the pieces of tissue on the tissue array sheet fabricated by the conventional process of collecting cores by punching.
  • the conventional pieces of tissue had a circular shape and were collected from lopsided regions.
  • the pieces of tissue in the present example had a spiral shape and were collected widely from dispersed regions.
  • FIG. 10( e ) is a partially enlarged view of the spiral piece of tissue t 4 emerging in the tissue array chip of the present example.
  • a site of interests was contained in the piece of tissue at the position indicated by an arrow. According to the conventional collection of cores by punching, such a site of interests could not be found unless it should be contained in a single point to be punched.
  • the pieces of tissue t 4 of the present example had a spiral shape and were collected from a region more dispersed than ever to enable the probability of finding the site of interests to be heightened. Thus, a very high effect could be obtained.
  • the tissue block to be used is not limited thereto.
  • a frozen tissue block having tissue frozen may be used.
  • an embedding agent called a compound is used instead of paraffin. It is preferred that the tissue is frozen in a state embedded in the compound and that the aforementioned processes are performed in a cooled environment capable of maintaining the frozen state.
  • the tissue to be used is not limited insofar as it is body tissue.
  • the aforementioned processes and systems may appropriately be modified within the object of the present invention.
  • FIG. 1 includes explanatory views illustrating a process of fabricating pieces of tissue, a process of fabricating a tissue block and a tissue array block according to an embodiment of the present invention.
  • FIG. 2 includes explanatory views illustrating a process of fabricating a tissue array sheet according to the present embodiment.
  • FIG. 3 includes explanatory views illustrating the difference between a conventional tissue array chip and a tissue array chip according to the present embodiment.
  • FIG. 4 includes explanatory views illustrating the difference between the conventional tissue array chip and the tissue array chip according to the present embodiment.
  • FIG. 5 is an explanatory view illustrating a system of fabricating a piece of tissue according to the present embodiment.
  • FIG. 6 includes explanatory views illustrating the operation of the system of fabricating the piece of tissue according to the present embodiment.
  • FIG. 7 is an explanatory view illustrating a system of fabricating a tissue array block according to the present embodiment.
  • FIG. 8 includes explanatory views illustrating the operation of the system of fabricating the tissue array block according to the present embodiment.
  • FIG. 9 includes explanatory views functionally illustrating a system of fabricating a tissue array sheet according to the present embodiment.
  • FIG. 10 includes photographs illustrating the difference between the tissue array chip according to the present embodiment and the conventional tissue array chip.
  • FIG. 11 includes explanatory views illustrating a process of fabricating the conventional array chip.
  • FIG. 12 includes explanatory views illustrating a conventional tissue block, a conventional tissue array block and conventional tissue array chips.
  • FIG. 13 includes explanatory views illustrating a conventional tissue block, a conventional tissue array block and conventional tissue array chips assumed when a conventional site of interests has been used as a target.
  • FIG. 14 includes explanatory views illustrating positional relationship between a site of interests included in a core and a site of interests formed on a tissue block surface.

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US12/449,865 2007-03-07 2008-03-05 Process of fabricating tissue array block, process of fabricating tissue array sheet, tissue array block, tissue array chip, system of fabricating tissue array block and system of fabricating tissue array sheet Abandoned US20100087333A1 (en)

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JP5629939B2 (ja) * 2010-09-09 2014-11-26 株式会社パソロジー研究所 パイルアップ組織アレイ
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JP6367741B2 (ja) * 2015-03-12 2018-08-01 学校法人北里研究所 生体組織標本アレイ切片、生体組織標本アレイ、シート状生体組織標本アレイ、生体組織標本ブロック及びそれらの製造方法
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US8551922B1 (en) 2013-10-08
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US20130281322A1 (en) 2013-10-24
EP2116610A1 (en) 2009-11-11

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