Method for tissue sample fixation

Patent metadata

Field	Value
Publication number	US-12158401-B2
Application number	US-202318110427-A
Country	US
Kind code	B2
Filing date	Feb 16, 2023
Priority date	Feb 17, 2011
Publication date	Dec 3, 2024
Grant date	Dec 3, 2024

Abstract

An aldehyde fixative solution at a first temperature is caused to contact a tissue sample for a first time period, additionally an aldehyde fixative solution is caused to contact the tissue sample at a second temperature higher than the first temperature for a second time period. The first time period typically ranges from about 15 minutes up to about 4 hours, and the first temperature typically is from greater than 0° C. to at least 15° C. The second temperature typically is from greater than about 22° C. to about 55° C., and the second time period ranges from about 1 hour to about 4 hours. Using this process, improved tissue morphology and IHC staining as well as superior preservation of post-translation modification signals have been accomplished in approximately 4 hours compared to 24 hours for room temperature protocols, and more even morphology and antigen preservation are observed.

First claim

The invention claimed is:

1. A method for fixing a tissue sample, comprising:
performing a cold immersion step on the tissue sample, the cold immersion step comprising immersing the tissue sample in a first aldehyde solution for a first time period sufficient to diffuse the first aldehyde solution into an interior region of the tissue sample, wherein the first aldehyde solution is in a first temperature range from a freezing point of the first aldehyde solution to about 10° C. during the first time period; and
performing a second immersion step on the tissue sample after the cold immersion step to obtain the fixed tissue sample, the second immersion step comprising exposing the tissue sample to a temperature in a second temperature range from about 22° C. to about 50° C. for a second time period,
wherein the tissue sample is not exposed to ultrasonic or electromagnetic irradiation during either (i) the cold immersion of the tissue sample, or (ii) the second immersion of the tissue sample.

2. The method of claim 1 , further comprising subjecting the fixed tissue sample to a series of alcohol immersions to obtain a dehydrated fixed tissue sample; subjecting the dehydrated tissue sample to an immersion in a clearing solution; embedding the dehydrated tissue sample in a wax block after the immersion in the clearing solution; sectioning the wax block to obtain a tissue section; and affixing the tissue section to a microscope slide.

3. The method of claim 2 , wherein the method further comprises: immunohistochemically staining the tissue section affixed to the slide for a protein or a post-translationally modified protein.

4. The method of claim 3 , wherein the post-translationally modified protein is a phosphorylated protein, and wherein the immunohistochemical stain specifically detects the phosphorylated protein, but not the non-phosphorylated protein.

5. The method of claim 2 , wherein the method further comprises: staining the tissue section affixed to the slide for a nucleic acid by in situ hybridization.

6. The method of claim 5 , wherein the nucleic acid is an RNA.

7. A method for processing a tissue sample, the method comprising:
immersing the tissue sample in a first solution at a first temperature for a first time period to diffuse the first solution into an interior region of the tissue sample; and
immersing the tissue sample in a second solution at a second temperature for a second time period,
wherein the tissue sample is not exposed to ultrasonic or electromagnetic irradiation during diffusion of the first solution into the interior region of the tissue sample or during immersion of the tissue sample in the second solution,
wherein at least one of the first or second time periods is a processing time of a tissue processing protocol derived from a time of flight curve for one or more of: (i) a tissue type that is about the same type as the tissue sample or of a similar type to the tissue sample; (ii) a tissue shape that is about the same shape as the tissue sample or of a similar shape to the tissue sample; or (iii) a tissue size that is about the same size as the tissue sample or of a similar size to that of the tissue sample.

8. The method of claim 7 , wherein first temperature ranges from between 0° C. to 10° C.

9. The method of claim 7 , wherein the second temperatures ranges from between 22° C. to 55° C.

10. The method of claim 7 , wherein the first and second solutions are the same.

11. The method of claim 7 , wherein the first and second solutions are different.

Assignees

• Ventana Med Syst Inc

Inventors

• Chafin David
• Pierson Theiss Abbey
• Otter Michael
• Roberts Esteban

Classifications

• G01N1/31

  Apparatus therefor · CPC title

• G01N1/30 Primary

  Staining; Impregnating {; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis} · CPC title