JP2010185673A - Tissue microarray preparation method - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a tissue microarray preparation method not including a defect of a conventional technology. <P>SOLUTION: This method includes: a processes for heating an embedding dish and a mold, pouring paraffin having a melting point higher than a donor block as much as at least 1°C into the embedding dish, and setting and cooling the heated mold to solidify the paraffin; a process for extracting the mold from the solidified paraffin, and preparing a recipient block for tissue microarray preparation; a process for extracting a cylindrical tissue from the donor block, and inserting the extracted tissue into a cylindrical hole on the recipient block for tissue microarray preparation; and a process for pouring heated and dissolved paraffin having the same melting point as the donor block into the recipient block for tissue microarray preparation into which the tissue from the donor block is inserted, and incubating the recipient block at a temperature which is higher than the melting point of the donor block and lower than the melting point of the recipient block, to thereby dissolve the paraffin in the donor block. <P>COPYRIGHT: (C)2010,JPO&INPIT

Description

  The present invention relates to a recipient block for producing a tissue microarray, a sword mountain mold for molding a recipient block, and a method for producing a tissue microarray using them.

  A tissue microarray is a large number of tumor tissues and tumor cell lines arranged in an array on a single glass slide, and can be immunostained and in situ hybridized under the same conditions at once. Since then, it has come to be used in the fields of biology, medicine and pharmacy.

  The tissue microarray is produced by the following method. First, a paraffin block for embedding a tissue sample called a recipient block is prepared, a hole is made in the recipient block with a hollow needle, and then a sample is taken from a predetermined part of the paraffin block (donor block) containing the tissue sample Then, the array block is produced by repeating the operation of embedding the collected sample in the recipient block. Thereafter, the array block is sliced and fixed to the slide glass (Patent Document 1).

  In the conventional method, since the columnar tissue cut out from the donor block was only inserted into the hole formed in the recipient block, the inserted columnar tissue and the paraffin constituting the wall surface of the hole in the recipient block were used. Are not fused, and the columnar tissue was in a movable state in a hole drilled in the recipient block, so that the tissue might fall off or be damaged when sliced. For this reason, it was necessary to use the tape cutting system which coat | covered the adhesive agent (patent document 1, FIG. 4). Furthermore, in the widely used paraffin tape transfer system (Instrumedics Inc.), it is necessary to slice and paste using a dedicated tape and a dedicated slide, and to fix the adhesive on the slide by UV irradiation.

Special table 2002-505431

  An object of the present invention is to provide a method for producing a tissue microarray that does not include the above-described drawbacks of the prior art.

  As a result of intensive studies to solve the above-mentioned problems, the present inventors have made a difference in the melting points of the donor block and the recipient block, and when slicing without using a dedicated tape or a dedicated slide. As a result, the present invention was completed. That is, the present invention is as follows.

[1] A recipient block for preparing a tissue microarray having a melting point that is at least 1 ° C. higher than that of the donor block.
[2] The recipient block according to [1], having cylindrical holes arranged in an array for embedding a cylindrical tissue from the donor block.
[3] A sword mountain-shaped form for molding a recipient block according to [2], wherein the cylinders are arranged in an array.
[4] The mold according to [3], wherein the cylinder is made of Teflon (registered trademark) with an iron material.
[5] The following steps:
Heat the embedding dish and the form described in [3] or [4], pour paraffin having a melting point higher by at least 1 ° C. than the donor block into the embedding dish, and set the heated form. Cooling and solidifying the paraffin;
Extracting the formwork from the solidified paraffin, and producing a recipient block for producing a tissue microarray according to [2],
Extracting the cylindrical tissue from the donor block and inserting the extracted tissue into a cylindrical hole on the recipient block for tissue microarray preparation;
Pour heated and dissolved paraffin having the same melting point as the donor block into the tissue microarray preparation recipient block into which the tissue from the donor block has been inserted, and the recipient block is higher than the melting point of the donor block but the recipient block. A method for producing a tissue microarray, comprising the step of incubating at a temperature lower than the melting point of the solution to dissolve the paraffin of the donor block.

  The tissue microarray fused with the tissue by using the recipient block for producing the tissue microarray of the present invention, the sword mountain mold for molding the recipient block, and the method for producing the tissue microarray using them, and a special device or instrument. It is possible to easily and easily manufacture the device without using the material, and it is possible to reduce the working time and the cost.

  The “donor block” of the present invention is a paraffin-embedded tissue specimen prepared according to a conventional method. The melting point of paraffin for the “donor block” of the present invention is 55-65 ° C., preferably 55-60 ° C., more preferably 57-60 ° C. The paraffin can include beeswax if desired. Examples include tissue-tech paraffin wax II60 (Sakura Seiki, melting point 57-60 ° C), histoparaffin (Merck, melting point 56-58 ° C), paraffin (pellet type) for histology (Merck, melting point 56-58 ° C). be able to

  The “recipient block for preparing a tissue microarray” of the present invention has a higher melting point than the donor block. The difference in melting point between the recipient block for tissue microarray preparation and the donor block is at least 1 ° C, preferably at least 3 ° C, more preferably at least 6 ° C. When the melting point of the paraffin for each block is indicated in a numerical range, the difference between the highest temperature in the numerical range of the melting point of the donor block and the lowest temperature in the numerical range of the melting point of the recipient block is indicated. For example, when the numerical range of the melting point of the donor block is 57-60 ° C and the numerical range of the melting point of the recipient block is 66-68 ° C, the difference in melting point is 6 ° C. The recipient block paraffin can optionally include beeswax.

  The “recipient block for producing a tissue microarray” of the present invention can have cylindrical holes arranged in an array for embedding a cylindrical tissue from a donor block. The diameter of the holes and the arrangement in the array (the number of holes and the arrangement in the vertical and horizontal directions) can be appropriately set as needed by those skilled in the art.

  The “sword mountain-shaped form for molding a recipient block for producing a tissue microarray” of the present invention is a sword-shaped form for forming a recipient block in which cylinders are arranged in an array. For example, a cylindrical needle is arranged on a flat plate. The diameter of the holes and the arrangement in the array (the number of holes and the arrangement in the vertical and horizontal directions) can be appropriately set as needed by those skilled in the art. The cylinder can be Teflon (registered trademark) processed into an iron material.

  In the method for producing the tissue microarray of the present invention, the temperature higher than the melting point of the donor block but lower than the melting point of the recipient block is a temperature at least 1 ° C. higher than the melting point of the donor block but at least 1 ° C. lower than the melting point of the recipient block. Preferably, at least 3 ° C. above the melting point of the donor block but at least 1 ° C. below the melting point of the recipient block, more preferably at least 5 ° C. above the melting point of the donor block but at least 1 ° C. below the melting point of the recipient block. It is. When the melting point of the paraffin for each block is indicated in a numerical range, the highest temperature in the numerical range of the melting point of the donor block and the lowest temperature in the numerical range of the melting point of the recipient block are indicated. For example, when the numerical range of the melting point of the donor block is 57 to 60 ° C and the numerical range of the melting point of the recipient block is 66 to 68 ° C, the temperature is higher than 60 ° C but lower than 66 ° C.

  EXAMPLES Hereinafter, although an Example is given and this invention is demonstrated further in detail, this invention is not limited to these.

  A Kenyama-shaped formwork made of Teflon (registered trademark) on an iron material (the diameter and number of cylindrical needles are 228 of 12 × 19 when the diameter is 1 mm, and 8 × when the diameter is 2 mm. In the case where the diameter of 13 is 104 and the diameter is 3 mm, the number is 54 of 6 × 9.).

  Paraffin (melting point 66-68 ° C., paraffin (Code NO. 24-0450, Sigma-Aldrich Japan) dissolved in a gas burner in an embedding dish preheated to 65 ° C. with a heat block: beeswax (Code No. 03 -0670-5, Sigma-Aldrich Japan) = 9: 1) was poured, and a mold that had been heated in the same manner was set and cooled to room temperature. After the paraffin has solidified, the heat block is heated to 65 ° C. with the mold facing down, the mold frame is pulled out from the paraffin block, and the cylindrical holes are arranged in an array defined by the mold A recipient block was prepared.

  Next, using a paraffin block dedicated arrayer (KIN-1 type: Toya Medical Instrument) from a tissue donor block prepared according to a standard method, the tissue is hollowed out in a cylindrical shape at a desired position, and the hollowed-out tissue is used as a recipe. The paraffin having a melting point of 57-60 ° C. was poured into the hole of the ent block. This recipient block was incubated at 65 ° C. to dissolve paraffin with a melting point of 57-60 ° C. and then cooled to room temperature to solidify the paraffin.

  When sliced and stained by a conventional method, a good tissue microarray could be obtained.

  The recipient block for preparing a tissue microarray of the present invention, the Kenyama-shaped form for forming the recipient block, and the tissue microarray preparation method using the same allow processing of multiple specimens of pathological specimens. Useful for throughput analysis.

Claims (5)

A recipient block for preparing a tissue microarray, having a melting point that is at least 1 ° C. higher than the donor block. 2. The recipient block of claim 1 having cylindrical holes arranged in an array for embedding cylindrical tissue from a donor block. The sword mountain-shaped form for forming a recipient block according to claim 2, wherein the cylinders are arranged in an array. The mold according to claim 3, wherein the cylinder is made of Teflon (registered trademark) with an iron material. The following process:
Heating the embedding dish and the mold according to claim 3 or claim 4, pouring paraffin having a melting point at least 1 ° C. higher than that of the donor block into the embedding dish, setting the heated mold; Cooling and solidifying the paraffin;
Extracting the formwork from the solidified paraffin, and producing a tissue microarray recipient block according to claim 2,
Extracting the cylindrical tissue from the donor block and inserting the extracted tissue into a cylindrical hole on the recipient block for tissue microarray preparation;
Pour heated and dissolved paraffin having the same melting point as the donor block into the tissue microarray preparation recipient block into which the tissue from the donor block has been inserted, and the recipient block is higher than the melting point of the donor block but the recipient block. A method for producing a tissue microarray, comprising the step of incubating at a temperature lower than the melting point of the solution to dissolve the paraffin of the donor block.