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Immunohistochemical hematoxylin-toluidine blue double-redyeing method for melanocyte

A technique of toluidine blue and immunohistochemistry, applied in the field of detection, to achieve the effect of consistent color, simple and convenient steps, and obvious color difference

Inactive Publication Date: 2015-10-21
CHUGOKU IGAKU KAGAKUIN HIFUBIYOU KENKYUSHO
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Toluidine blue stock solution is more simple and convenient to prepare than Giemsa stock solution

Method used

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  • Immunohistochemical hematoxylin-toluidine blue double-redyeing method for melanocyte
  • Immunohistochemical hematoxylin-toluidine blue double-redyeing method for melanocyte
  • Immunohistochemical hematoxylin-toluidine blue double-redyeing method for melanocyte

Examples

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Embodiment 1

[0074] Example 1, the method of hematoxylin-toluidine blue double staining of melanin tissue immunohistochemistry, the steps are as follows:

[0075] ⑴. Routine xylene dewaxing of slices, gradient alcohol dehydration:

[0076] Xylene I and II each for 30 minutes, absolute ethanol, 95%, 90%, and 85% gradient alcohol for 3 minutes each, and washed with tap water.

[0077] ⑵. Block and inactivate endogenous peroxidase:

[0078] 3%H 2 o 2 Incubate at 37°C for 10 min, wash with PBS buffer for 5 min x 3 times;

[0079] ⑶. Antigen retrieval (routine operation);

[0080] ⑷. Serum blocking: Incubate goat serum at 37°C for 10 minutes, pour it out and do not wash;

[0081] ⑸. Add the primary antibody dropwise, PBS instead of the primary antibody as a negative control, and incubate overnight at 4°C;

[0082] ⑹. Wash with PBS buffer for 5min×3 times;

[0083] ⑺. Add the secondary antibody dropwise and incubate at room temperature for 20 minutes in the dark;

[0084] ⑻. Wash wi...

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Abstract

The invention provides an immunohistochemical hematoxylin-toluidine blue double-redyeing method for melanocyte. The method comprises the following steps: (1), carrying out conventional xylene dewaxing on a slice and carrying out gradient alcohol dehydration; (2), blocking and inactivating endogenous peroxide enzyme; (3), repairing an antigen; (4), carrying out serum sealing; (5), dropwise adding primary antibodies; (6) dropwise adding secondary antibodies; (7), carrying out DAB developing; (8), carrying out hematoxylin redyeing and hydrochloric acid alcohol differentiation; (9), redyeing by a toluidine blue working solution and carrying out glacial acetic acid differentiation; (10), dehydrating to be transparent and sealing the slice. According to the invention, positive granules are still brownish yellow after redyeing, and original melanin granules in the tissue become deep green, so that distinction is obvious, the melanin granules are easily distinguished from brown granules, the developing of hematoxylin is not influenced, cell nucleuses of background cells are clearly visible, the color difference of immunohistochemical positive brownish yellow granules, deep green melanin granules and blue cell nucleus is obvious and intuitive, and distinguishing is convenient. Toluidine blue dyeing liquid is prepared simply and conveniently.

Description

technical field [0001] The invention relates to a detection technology, in particular to a hematoxylin-toluidine blue double superstaining method for immunohistochemistry of melanocytes and their proliferative lesions. Background technique [0002] In the pathological research or clinical research of skin diseases, the research of human melanocytes and their proliferative lesions, especially malignant melanoma is one of the very active fields. The study of melanocytes with their own melanin granules must use immunohistochemical staining techniques for melanin tissue. The experimental steps of routine clinical non-melanin tissue immunohistochemical staining technique (ABC method) are as follows: [0003] ⑴. Routine xylene dewaxing of slices, gradient alcohol dehydration: [0004] Xylene I and II each for 30 minutes, absolute ethanol, 95%, 90%, and 85% gradient alcohol for 3 minutes each, and washed with tap water. [0005] ⑵. Block and inactivate endogenous peroxidase: ...

Claims

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Application Information

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IPC IPC(8): G01N1/30
Inventor 姜祎群吴琼孙建方李阿梅邵雪宝
Owner CHUGOKU IGAKU KAGAKUIN HIFUBIYOU KENKYUSHO
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