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Hematoxylin staining method

A technology of hematoxylin and hematin, applied in the field of biological sample staining

Inactive Publication Date: 2013-11-27
VENTANA MEDICAL SYST INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

This is especially problematic for automated systems that contain very small opening diameter metal parts such as nozzles and sprinklers that can become clogged with sediment

Method used

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  • Hematoxylin staining method
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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Embodiment 1: the preparation of solution

[0030] Prepare the following solutions:

[0031]

[0032]

[0033] For solutions A and B, the propylene glycol was heated on a hot plate at 60 °C. The phthalic acid is added, and the composition is heated until the solids dissolve. Turn off the heat and add potassium hydrogen phthalate in DI H to the propylene glycol solution with stirring. 2 solution in O. Add solid potassium aluminum sulfate to solution A. Solution A was then stirred overnight and then filtered using 25 micron fluted filter paper. Hematoxylin was added to Solution B, and the composition was stirred for about 15 minutes. Solid sodium iodate was then added and the solution was stirred overnight, then filtered using 25 micron grooved filter paper.

Embodiment 2

[0034] Example 2: Staining of Biological Samples

[0035] Material: KAl(SO 4 ) 2 Buffered solution (double strength; 2x solution A); hematoxylin buffered solution (double strength; 2x solution B), 50 / 50 propylene glycol / H with 1% Merpol wash 2O; 50 / 500.1 Tris blue dye (blueing); 95% propylene glycol; transfer fluid (Transfer fluid); 3X3MTB slides.

[0036] Use the following protocol on thick film (open) cassettes. Slides were manually deparaffinized, lightly blotted and placed in boxes. Dispense 400 μL of 2X Solution A onto the tissue, followed immediately by dispensing 400 μL of 2X Solution B on the tissue. Slides were then incubated for 3 minutes. Slides were then rinsed twice for 10 s with 800 μL of 1% aqueous Merpol. Slides were then treated with 800 µL of 95% propylene glycol for 10 s. Slides were then rinsed 3 times for 10 seconds each with 800 [mu]L of transfer fluid. Coverslips were then applied on a Sakkura TissueTek instrument.

[0037] Use this protocol on ...

Embodiment 3

[0038] Example 3: Preparation of two-part staining solution

[0039] Solution A: 0.75M phthalate-buffered KAl(SO 4 ) 2 . To 250ml of propylene glycol was added 8.97g of phthalic acid and the mixture was stirred at 60°C until the solids dissolved. Add potassium hydrogen phthalate solution (4.3 g in 750 mL DI H 2 O), and remove the solution from the heat source. Then add 12.0g KAl (SO 4 ) 2 12H 2 O, the mixture was stirred until it was homogeneous. The solution is then filtered. The pH is 2.59.

[0040] Solution B: 0.75M Phthalate Buffered Hematine. To 250 mL of propylene glycol was added 8.97 g of phthalic acid, and the mixture was stirred at 60°C until the solids dissolved. Add potassium hydrogen phthalate solution (4.3 g in 750 mL DI H 2 O), and remove the solution from the heat source. Add 12.0 g of hematoxylin with stirring. After 15-20 minutes, 1.2 g of sodium iodate was added and the mixture was stirred overnight. The solution is then filtered. The pH is 2...

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Abstract

The present invention relates to processes for staining biological samples, and in particular to automated processes for staining biological sample with hematoxylin stains. In the processes and systems of the invention, separate hematein and mordant solutions are provided which may be premixed prior to application to a biological sample. This method prevents precipitation common in hematein staining solutions and which fouls automated slide / sample processing equipment.

Description

field of invention [0001] The present invention relates to methods for staining biological samples, and in particular to automated methods for staining biological samples with hematoxylin stain. Background of the invention [0002] Several histochemical staining protocols, including hematoxylin and eosin (H&E) and Papanicolaou (PAP) staining, rely on the dye hematoxylin to stain cytological and tissue samples. Specifically, staining of nuclei with hematoxylin is used by pathologists to detect the presence of malignant and / or metastatic cells in tumor biopsy samples. [0003] Hematoxylin is a naturally occurring compound found in the red heartwood of trees of the genus Hematoxylon. Hematoxylin itself is colorless in aqueous solution and is not the active ingredient for staining tissue components. Instead, hematoxylin, the oxidation product of hematoxylin, becomes the reactive dyeing component of hematoxylin dye solutions, especially when complexed with mordants. Sumac occu...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N1/30G01N1/31
CPCG01N1/30G01N1/31G01N1/312G01N33/483
Inventor B.W.巴恩斯C.H.韦德纳
Owner VENTANA MEDICAL SYST INC
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