# US 10,481,054 B2 · Method and automatic machine for embedding a tissue sample into an embedding medium > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 10,481,054 B2 - Document ID: US10481054B2 - Status: Granted - Kind: B2 - Application number: 15/254,211 - Filed: 2016-09-01 - Published: 2017-03-09 - Granted: 2019-11-19 - Earliest claimed: 2015-09-04 - Assignees: Leica Biosystems Nussloch GmbH - Applicants: Leica Biosystems Nussloch GmbH - Inventors: Florian Böhl; Ralf Eckert; Karin Flieger; Markus Berberich; Hermann Ulbrich; Stella Knorr; Andrew Guy - Web page: https://patentgrep.com/patent/US10481054B2 ## CPC - G01N 1/36: Embedding or analogous mounting of samples - G01N 2001/364: {using resins, epoxy} - G01N 2001/366: {Moulds; Demoulding} - G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q) ## Abstract The invention relates to a method for embedding a tissue sample into an embedding medium. In a first step the tissue sample is held in an intended orientation in a container by means of a holding element that presses the tissue sample against a base of the container. In a further step a liquid embedding medium that has a temperature above 64 degrees Celsius, in particular in the range from 65 degrees Celsius to 67 degrees Celsius, or a temperature of 66 degrees Celsius, is poured into the container, and the base of the container is cooled. In a further step a separating motion is executed by way of which the tissue sample and the holding element move away from each other, those layers of the embedding medium through which the holding element moves during the separating motion having, during the separating motion, temperatures in the range from 54 degrees Celsius to 64 degrees Celsius, in particular of 60 degrees Celsius, and/or those portions of the embedding medium which are directly adjacent to the holding element having, during the separating motion, a temperature in the range from 54 degrees Celsius to 64 degrees Celsius, in particular of 60 degrees Celsius. ## Claims ### Claim 1 A method for embedding a tissue sample (1) into an embedding medium, comprising: a. holding the tissue sample (1) in an intended orientation in a container (2) by means of a holding element (3) that presses the tissue sample (1) against a base (4) of the container (2); b. pouring into the container (2) a liquid embedding medium (5) that has a temperature above 64 degrees Celsius; c. cooling the base (4) of the container (2); d. executing a separating motion by way of which the tissue sample (1) and the holding element (3) move away from each other, the embedding medium (5) through which the holding element (3) moves during the separating motion having, during the separating motion, temperatures in the range from 54 degrees Celsius to 64 degrees Celsius, and/or those portions of the embedding medium (5) which are directly adjacent to the holding element (3) having, during the separating motion, a temperature in the range from 54 degrees Celsius to 64 degrees Celsius; wherein e. the separating motion begins automatically when a predefined or predefinable activation temperature of the embedding medium (5) or of the container (2) or of the holding element (3) is reached: and/or f. the separating motion is automatically activated when a predefined or predefinable activation temperature of the embedding medium (5) or of the container (2) or of the holding element (3) is reached. ### Claim 2 (depends on 1) The method according to claim 1, wherein the embedding medium (5), upon pouring, has a temperature in the range from 65 degrees Celsius to 67 degrees Celsius. ### Claim 3 (depends on 2) The method according to claim 2, wherein the embedding medium (5), upon pouring, has a temperature of 66 degrees Celsius. ### Claim 4 (depends on 1) The method according to claim 1, wherein those layers of the embedding medium (5) through which the holding element (3) moves during the separating motion have, during the separating motion, a temperature of 60 degrees Celsius, and/or those portions of the embedding medium (5) which are directly adjacent to the holding element (3) have, during the separating motion, a temperature of 60 degrees Celsius. ### Claim 5 (depends on 1) The method according to claim 1, wherein the embedding medium (5) has, during execution of the separating motion, an average temperature in the range from 54degrees Celsius to 64 degrees Celsius. ### Claim 6 (depends on 5) The method according to claim 5, wherein the embedding medium (5) has, during execution of the separating motion, an average temperature of 60 degrees Celsius. ### Claim 7 (depends on 1) The method according to claim 1, wherein the embedding medium (5) contains paraffin. ### Claim 8 (depends on 7) The method according to claim 7, wherein the embedding medium (5) contains a greater amount of paraffin than any other ingredient. ### Claim 9 (depends on 8) The method according to claim 8, wherein the embedding medium (5) is paraffin. ### Claim 10 (depends on 1) The method according to claim 1, wherein the embedding medium (5), upon pouring, has a temperature that is at least 4 degrees Celsius above a drop point of the embedding medium. ### Claim 11 (depends on 10) The method according to claim 10, wherein the embedding medium (5), upon pouring, has a temperature that is 5 to 7 degrees Celsius above the drop point of the embedding medium. ### Claim 12 (depends on 11) The method according to claim 11, wherein the embedding medium (5), upon pouring, has a temperature that is 6 degrees Celsius above the drop point of the embedding medium. ### Claim 13 (depends on 1) The method according to claim 1, wherein a. the temperature of the embedding medium (5) or of the container (2) or of the holding element (3) is measured with a temperature sensor; or b. the temperature of the embedding medium (5) or of the container (2) or of the holding element (3) is measured with a temperature sensor and the separating motion is activated automatically as soon as the measured temperature reaches the activation temperature. ### Claim 14 (depends on 1) The method according to claim 1, wherein a holding mechanism that moves the holding element (3) comprises at least one shape memory component. ### Claim 15 (depends on 1) The method according to claim 1, wherein the separating motion occurs within a predefined or predefinable time window and/or lasts for a predefined or predefinable time period (t2). ### Claim 16 (depends on 15) The method according to claim 15, wherein the time window is 17 to 20seconds long; and/or the time period (t2) lasts 17 to 20 seconds. ### Claim 17 (depends on 1) The method according to claim 1, wherein the container (2) is cooled exclusively at the base (4); and/or the base (4) is cooled by the fact that the container (2) is brought into thermally conductive contact with a cooling element. ### Claim 18 A method for embedding a tissue sample (1) into an embedding medium, comprising: a. holding the tissue sample (1) in an intended orientation in a container (2) by means of a holding element (3) that presses the tissue sample (1) against a base (4) of the container (2); b. pouring into the container (2) a liquid embedding medium (5) that has a temperature above 64 degrees Celsius; c. cooling the base (4) of the container (2); d. executing a separating motion by way of which the tissue sample (1) and the holding element (3) move away from each other, the embedding medium (5) through which the holding element (3) moves during the separating motion having, during the separating motion, temperatures in the range from 54 degrees Celsius to 64 degrees Celsius, and/or those portions of the embedding medium (5) which are directly adjacent to the holding element (3) having, during the separating motion, a temperature in the range from 54 degrees Celsius to 64 degrees Celsius, wherein e. the separating motion begins immediately after expiration of a predefined or predefinable time period (t1) after the beginning of cooling; and/or f. the separating motion is automatically started immediately after expiration of a predefined or predefinable time period (t1) after the beginning of cooling. ### Claim 19 (depends on 18) The method according to claim 18, wherein the time period between the beginning of cooling and the beginning of the separating motion is in the range from 1second to 4 seconds. ### Claim 20 (depends on 19) The method according to claim 19, wherein the time period between the beginning of cooling and the beginning of the separating motion is in the range from 2seconds to 3 seconds. ### Claim 21 A method for embedding a tissue sample (1) into an embedding medium, comprising: a. holding the tissue sample (1) in an intended orientation in a container (2) by means of a holding element (3) that presses the tissue sample (1) against a base (4) of the container (2); b. pouring into the container (2) a liquid embedding medium (5) that has a temperature above 64 degrees Celsius; c. cooling the base (4) of the container (2); d. executing a separating motion by way of which the tissue sample (1) and the holding element (3) move away from each other, the embedding medium (5) through which the holding element (3) moves during the separating motion having, during the separating motion, temperatures in the range from 54 degrees Celsius to 64 degrees Celsius, and/or those portions of the embedding medium (5) which are directly adjacent to the holding element (3) having, during the separating motion, a temperature in the range from 54 degrees Celsius to 64 degrees Celsius, wherein the separating motion ends while the holding element (3) is still located inside the embedding medium; and/or the holding element (3) is embedded together with the tissue sample (1). ### Claim 22 A method for embedding a tissue sample (1) into an embedding medium, comprising: a. holding the tissue sample (1) in an intended orientation in a container (2) by means of a holding element (3) that presses the tissue sample (1) against a base (4) of the container (2), wherein the container (2) is part of a tissue cassette, and the tissue cassette comprises the holding element (3) that presses the tissue sample (1) against the base (4) of the container (2); b. pouring into the container (2) a liquid embedding medium (5) that has a temperature above 64 degrees Celsius; c. cooling the base (4) of the container (2); d. executing a separating motion by way of which the tissue sample (1) and the holding element (3) move away from each other, the embedding medium (5) through which the holding element (3) moves during the separating motion having, during the separating motion, temperatures in the range from 54 degrees Celsius to 64 degrees Celsius, and/or those portions of the embedding medium (5) which are directly adjacent to the holding element (3) having, during the separating motion, a temperature in the range from 54 degrees Celsius to 64 degrees Celsius. ### Claim 23 A method for processing a tissue sample (1), comprising: introducing the tissue sample (1) into a container (2); fixing the tissue sample (1) with at least one fixing agent, and/or infiltrating the tissue sample (1) with an infiltration medium while the tissue sample (1) is located in the container (2); keeping the tissue sample (1) in the container (2) after the fixing and/or infiltration; and block-embedding the tissue sample (1) into an embedding medium while the tissue sample (1) remains in the container (2) after the fixing and/or infiltration, wherein the block-embedding comprises: a. holding the tissue sample (1) in an intended orientation in a container (2) by means of a holding element (3) that presses the tissue sample (1) against a base (4) of the container (2); b. pouring into the container (2) a liquid embedding medium (5) that has a temperature above 64 degrees Celsius; c. cooling the base (4) of the container (2); and d. executing a separating motion by way of which the tissue sample(1) and the holding element (3) move away from each other, the embedding medium (5) through which the holding element (3) moves during the separating motion having, during the separating motion, temperatures in the range from 54 degrees Celsius to 64 degrees Celsius, and/or those portions of the embedding medium (5) which are directly adjacent to the holding element (3) having, during the separating motion, a temperature in the range from 54 degrees Celsius to 64 degrees Celsius. ### Claim 24 (depends on 23) The method according to claim 23, wherein the infiltration medium is paraffin. ### Claim 25 (depends on 23) The method according to claim 23, wherein the container (2) is part of a tissue cassette and the container (2) has a base (4), and the tissue cassette comprises a holding element (3) that presses the tissue sample (1) against the base (4) of the container (2).