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Chloroacetate AS-D naphythol AS-D chloroacetate esterase (AS-DNCE) staining solution (chemical coloring process)

A technology of AS-D and chlorinated acetic acid, which is applied in the field of medical in vitro diagnosis, can solve the problems of unfavorable observation of granulocyte line positive cells, affecting the diagnosis and typing of leukemia diseases, and smears cannot be stored for a long time, and the dyeing effect is not easy to decolorize. , The dyeing effect is stable and the dyeing effect is lasting

Active Publication Date: 2014-06-25
SHANGHAI SUNBIO TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] At present, with the traditional AS-DNCE staining method, the stained smears cannot be stored for a long time, and must be observed in time after staining, otherwise the pigmentation particles will become light or even fade, which is not conducive to the observation of granulocyte-positive cells, thus affecting leukemia diseases diagnostic classification

Method used

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  • Chloroacetate AS-D naphythol AS-D chloroacetate esterase (AS-DNCE) staining solution (chemical coloring process)
  • Chloroacetate AS-D naphythol AS-D chloroacetate esterase (AS-DNCE) staining solution (chemical coloring process)
  • Chloroacetate AS-D naphythol AS-D chloroacetate esterase (AS-DNCE) staining solution (chemical coloring process)

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Embodiment 1: the composition and preparation method of kit of the present invention

[0042] Fixative solution: take 10mL of 40% formaldehyde, weigh disodium hydrogen phosphate (Na 2 HPO 4 ) 0.65g, sodium dihydrogen phosphate (NaH 2 PO 4 ) 0.4g, add 90mL of distilled water, the pH value is 7.3, pack in 2.5mL per bottle, seal, and store in a refrigerator at 4°C.

[0043] Incubation Solution A (AS-D Naphthol Sodium Chloroacetate Solution): Weigh 50mg AS-D Naphthol Sodium Chloroacetate and dissolve it in 5mL N,N-Dimethylformamide (DMF). pack, seal.

[0044] Incubation solution B (diazo agent-fast red B): weigh 30mg fast red B powder and put it in a reagent bottle.

[0045] Incubation solution C (buffer solution containing saponin): weigh disodium hydrogen phosphate (Na 2 HPO 4 ) 8.8g, potassium dihydrogen phosphate (KH 2 PO 4 ) 0.9g, saponin (Saponin, purchased from Sigma) 0.1g, sodium azide 1g, dissolved in 1000mL of distilled water, pH value is 7.6, divided int...

Embodiment 2

[0047] Embodiment 2: the staining method of kit of the present invention

[0048] 1. Prefabricated dyeing incubation solution: take 1 bottle of incubation solution A, incubation solution B and incubation solution C prepared in Example 1, mix them, shake gently and mix well, and then make the dyeing incubation solution;

[0049] 2. Add the fixative prepared in Example 1 pre-cooled at 4°C to the fresh bone marrow cell smear or blood cell smear dropwise to cover the smear, rinse with water 3 times after 30 seconds, and dry the smear naturally;

[0050] 3. Pour the dyeing incubation solution in step 1 into the dyeing tank, place the smear in the dyeing incubation solution, and place it at 37°C for 30 minutes, then rinse the tank with running water for several minutes, and dry the smear naturally;

[0051] 4. Add the counterstain solution prepared in Example 1 dropwise to the dried smear to cover the smear, rinse with water for 3 times after 1-2 minutes, dry the smear naturally, an...

Embodiment 3

[0052] Embodiment 3: normal human blood cell smear detection

[0053] Get normal person's peripheral blood to make blood smear, use kit of the present invention to carry out staining to blood smear according to the method for embodiment 2, blood smear after dyeing observe blood image and take pictures with microscope oil lens, as figure 1 .

[0054] Judgment of staining results: Negative result is colorless cytoplasm; positive result is red precipitate in cytoplasm: cytoplasm is light red (+), bright red precipitate is full of cytoplasm (++), dark red precipitate is full of cytoplasm ( +++).

[0055] Dyeing result: by figure 1 The results showed that normal mature neutrophils were positive, and monocytes were negative.

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Abstract

The invention belongs to the field of medical in vitro diagnosis and discloses a chloroacetate AS-D naphythol AS-D chloroacetate esterase enzyme staining kit of which a component of a buffer solution comprises saponin. When the kit is used for staining, blood cells are incubated by the saponin-containing buffer solution, so that pigments can enter the blood cells through holes formed in a cell membrane through the pigments and are combined with the chloroacetate AS-D naphythol AS-D chloroacetate esterase for developing, the cells are closed again after washing, the pigmentation particles are closed in the blood cells, the chloroacetate AS-D naphythol AS-D chloroacetate esterase does not fade for a long time after staining, and staining groups are protected. The kit is lasting in staining effect, difficult to discolor, simple in composition and convenient to use and is widely applied to staining the chloroacetate AS-D naphythol AS-D chloroacetate esterase in the blood cells, and standard operation is easily realized.

Description

technical field [0001] The invention belongs to the field of medical in vitro diagnosis, in particular to a staining kit for chlorinated AS-D naphthol acetate esterase, in particular to a chemical staining method for detecting specific esterase chlorinated AS-D naphthol acetate esterase in blood cells Staining kit. Background technique [0002] Naphthol AS-D chloroacetate esterase (AS-DNCE) is a granulocyte-specific esterase that usually exists in the granulocyte lineage. At present, chemical staining is often used to detect specific esterases. The detection principle is that AS-DNCE in blood cells hydrolyzes AS-D naphthol chloride acetate to produce naphthol AS-D, which is coupled with a diazo agent to form a colored Precipitate, the color depth of the precipitate is directly proportional to the activity of AS-DNCE. [0003] AS-DNCE is almost always present in granulocytes, while monocytic cells are mostly negative for AS-DNCE staining, only a few are weakly positive. AS...

Claims

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Application Information

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IPC IPC(8): G01N1/30
Inventor 谢永华朱美萍许付丁婧姝
Owner SHANGHAI SUNBIO TECH
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