Histological specimen treatment

• US 10,641,688 B2
• Filed: 02/23/2017
• Issued: 05/05/2020
• Est. Priority Date: 05/07/2015
• Status: Active Grant

• Current Assignee: Steven Wheeler
• Original Assignee: Steven Wheeler
• Inventors: Wheeler, Steven
• Primary Examiner(s): Kim, Taeyoon
• Assistant Examiner(s): Gough, Tiffany M
• Application Number:
  US15/440,102
• Publication Number:
  US 20170160172A1
• Time in Patent Office: 1,167 Days
• CPC Class Codes: C08F 220/20 of polyhydric alcohols or phenols , e.g. 2-hydroxyethyl (meth)acrylate or glycerol mono-(meth)acrylate
• CPC Class Codes: C08F 220/281 and containing only one oxygen, e.g. furfuryl (meth)acrylate or 2-methoxyethyl (meth)acrylate
• CPC Class Codes: C08F 222/102 of dialcohols, e.g. ethylene glycol di(meth)acrylate or 1,4-butanediol dimethacrylate
• CPC Class Codes: C08F 2800/20 as weight or mass percentages
• CPC Class Codes: C08K 5/0016 Plasticisers
• CPC Class Codes: C08K 5/12 of cyclic polycarboxylic acids
• CPC Class Codes: G01N 1/30 Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
• CPC Class Codes: G01N 2001/307 non-toxic, no Hg, no formaldehyde

Abstract

A single dissolving compound forms plural azeotropes, which can be azeotropically vaporized off at various stages of the treatment process, thus maintaining predictable concentrations of the chemicals present. The treatment process can be performed in the absence of formalin or related compounds which can interfere with the preservation of genetic material. A process for preserving a specimen includes using a dissolving compound that can form a plural number of azeotropes, at least one azeotrope being formed between one or more components of the dissolving compound and specimen-supplied water, and at least one azeotrope being formed between different components of the dissolving compound; successively and azeotropically vaporizing off formed azeotropes; and impregnating the specimen with a support medium.

Claims

1. A process for preserving a specimen comprising:

removing from the specimen cellular solutes that interfere with preservation and replacing the removed solutes with an impregnated support medium;

wherein the removing comprises extracting an amount of water supplied by the specimen;

wherein the extracting comprises;

dissolving the solutes including the amount of water using a dissolving compound;

forming an azeotropic water-containing mixture of the amount of water and a component of the dissolving compound;

azeotropically vaporizing off an amount of the azeotropic water-containing mixture;

forming a second azeotropic mixture of at least two components of the dissolving compound;

azeotropically vaporizing off an amount of the second azeotropic mixture;

forming a third azeotropic mixture of at least two components of the dissolving compound, the third azeotropic mixture being different from the second azeotropic mixture; and

azeotropically vaporizing off an amount of the third azeotropic mixture,

wherein, under pressure and temperature control, one of the azeotropic mixtures is formed and vaporized, followed by formation and vaporization of another of the azeotropic mixtures, followed by formation and vaporization of another of the azeotropic mixtures.

2. The process of claim 1, wherein the removing further comprises:

clearing an amount of the dissolving compound from the specimen;

wherein the clearing comprises;

forming one of the second and third azeotropic mixtures as an azeotropic fixitive-containing mixture of an amount of a fixitive agent component present in the dissolving compound and another component in the dissolving compound.

3. The process of claim 1, wherein the removing further comprises:

clearing an amount of the dissolving compound from the tissue;

wherein the clearing comprises;

forming one of the second and third azeotropic mixtures as an azeotropic dehydrant-containing mixture of an amount of a dehydrant agent component present in the dissolving compound and another component in the dissolving compound.

4. The process of claim 1, wherein the impregnated support medium comprises paraffin.

5. The process of claim 1, wherein the impregnated support medium is selected from the group consisting of:

paraffin, plastic polymers, polyvinyl alcohol, polyethylene glycols, waxes, cellulose derivatives, agars, gelations, and sugars.

6. The process of claim 1, wherein the dissolving compound comprises a single component or groups of components that is removed by vaporization from the specimen without damage to the specimen.

7. The process of claim 1, wherein the dissolving compound comprises a single component or groups of components that release formaldehyde from the specimen and convert it to a monomeric gas.

8. The process of claim 1, wherein the dissolving compound further comprises a support medium component.

9. The process of claim 1, wherein the dissolving compound comprises

a fixative component;

a dehydrant component; and

a clearing agent component.

10. The process of claim 9, wherein the dissolving compound further comprises a cation as a cellular anti-rupturing agent.

11. The process of claim 10, wherein the cation cellular anti-rupturing agent comprises a compound selected from the group consisting of:

zinc acetate, iron acetate, magnesium acetate, zinc citrate, iron citrate, and zinc malate.

12. The process of claim 9, wherein the fixative component is selected from the group consisting of:

formalin, glacial acetic acid, zinc chloride or zinc sulfate, chromic acid, ethanol, methanol, acetone, mercuric chloride, picric acid, and potassium dichromate.

13. The process of claim 9, wherein the dehydrant component is selected from the group consisting of:

alcohols, isopropyl alcohol, ethanol, denatured alcohols, butanols, ethers, ketones, dioxane, and acetone.

14. The process of claim 9, wherein the clearing agent component is selected from the group consisting of:

xylene, toluene, benzene, acetone, butanols, dioxane, methyl salicylate, cedarwood oil, hexane, heptane, octane, and decane.

15. The process of claim 9, wherein the dielectric constant of the clearing agent component is between 2.0 and 10.

16. The process of claim 1, wherein the dissolving compound comprises a mixture of between about 0 and 30% by weight alcohol, between about 45 and 99% by weight ether, and between about 0 and 20% by weight hydrocarbon.

17. The process of claim 1, further comprising prefixing the specimen using an amount of fixative comprising:

a mixture comprising between about 20 and 50% by weight methanol;

wherein the mixture is saturated with anhydrous ammonia;

wherein the mixture has a pH maintained between about 5.0 and 5.4 through the application of an effective amount of glacial acetic acid.

18. The process of claim 17, further comprising the mixture including:

an amount of vanadyl-ribonucleoside complex or RNAlater®

as a nucleic acid preservative.

19. The process of claim 1, further comprising prefixing the specimen using an amount of fixative comprising:

a mixture of between about 80 and 98% by weight methanol, between about 1 and 20% by weight acetonitrile, and between about 1 and 5% by weight EDTA;

wherein the mixture has a pH maintained between about 5.0 and 5.4 through the application of an effective amount of glacial acetic acid.

20. The process of claim 17, further comprising the mixture including:

between about 20 and 50% by weight ethanol.