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Cytochemical staining

Cytochemical staining is a science formed by combining cytology and chemistry. It is based on cell morphology and uses the principles of chemical reactions to detect various chemical components (such as enzymes, esters, iron, proteins) in blood cells. , nucleic acids, etc.) for qualitative, localization, and semi-quantitative analysis.

Edited at 2023-10-21 11:36:00

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Cytochemical staining

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Outline

Cytochemical staining

Overview of cytochemical staining

Definition: Cytochemical staining is a science formed by combining cytology and chemistry. It is based on cell morphology and uses the principles of chemical reactions to detect various chemical components (such as enzymes, esters, iron, etc.) in blood cells. , proteins, nucleic acids, etc.) for qualitative, localization, and semi-quantitative analysis.

Clinical application

Assist in determining the cell type of acute leukemia

Assist in the diagnosis and differential diagnosis of diseases such as the blood system

Observe disease efficacy and prognosis

Explore the pathogenesis

Steps: ① Fixation ② Display ③ Counterstaining

Commonly used cytochemical staining methods

Peroxidase staining (POX)

Peroxidase is widely present in living organisms. The peroxidase contained in blood cells is mainly myeloperoxidase, which is the most abundant protein in human neutrophils.

ICSH recommended methods: diaminobenzidine method DAB (common clinical method) benzidine dihydrochloride method, peroxidase amino-methylcarbazole staining method

staining reaction

Granulocyte cell line: mostly ( )

blasts

Poor differentiation (-)

Well differentiated ( )

Mature

Neutral lobulated granulocytes ( ) or strong

Eosinophils ( ) and the strongest

Basophils (-)

Mononuclear cell line: mostly weak Yang Yang

primitive monocytes (-)

Naive and mature monocytes are mostly weakly positive

Lymphocyte lineage, plasma cells, red blood cells, megakaryocytes (-)

clinical significance

The preferred and most important cytochemical staining method to assist in the diagnosis of acute leukemia types.

In acute myeloid leukemia, blasts are negative/positive, mostly positive

In acute lymphoblastic leukemia, both primitive lymphocytes and immature lymphocytes are negative, and the positive rate is <3%.

In acute monocytic leukemia, most primitive monocytes and immature monocytes are negative or weakly positive.

Sudan Black B stain (SBB)

Sudan black is a fat-soluble diazo dye that can dissolve in intracellular lipid-containing structures (such as neutral fats, phospholipids, glycolipids and steroids), making the lipids appear brown-black or dark black. Lipids are abundant in granulocytes and in small amounts in monocytes.

clinical significance

Basically similar to POX staining, the difference is that

Specificity: POX staining is higher than SBB staining Sensitivity: SBB staining is higher than POX staining

There are different requirements for the freshness of smears: SBB staining can use old smears, while POX staining requires fresh smears.

Esterase staining

specific esterase

Chloroacetate AS-D phenol esterase (NAS-DCE)

Non-specific esterase (different according to pH value)

Acid α-acetonaphthol esterase (ANAE)

neutral esterase

a-acetate esterase (a-NAE)

AS-D acetate esterase (NAS-DAE)

Alkaline esterase (a-butyric acid esterase; a-NBE)

staining reaction

NAS-DCE (particle, specificity)

granulocyte system

blasts

Well differentiated ( )

Poor differentiation (-)

Promyelocytes ( )

mature granulocytes

Neutrophils ( )

Eosinophils ( )/(-)

Basophils ( )

Many other cells (-)

α-NAE (Chinese, African, Special)/α-NBE/ANAE

monocyte system

primitive monocytes

Well differentiated ( )

Poor differentiation (-)

Naive monocytes, mature monocytes ( )

Positive reactions can be inhibited by sodium fluoride

Granulocyte system, lymphocyte system, other systems (-)/( ) Positive reactions are not inhibited by sodium fluoride

clinical significance

When the α-NAE staining of leukemia cells shows an obvious positive reaction, and the positive product can be inhibited by sodium fluoride, it should be considered as an emergency order.

Acid phosphatase staining (ACP)

Hairy cells ( ) are resistant to L-tartaric acid, and their positivity cannot be inhibited by L-tartaric acid

Gaucher cells ( ) tartaric acid-resistant, Niemann-Pick cells (-)

Clinical significance: Diagnosis of hairy cell leukemia, differentiation of Gaucher cells and Niemann-Pick cells

Glycogen staining (PAS), also known as periodic acid-Schiff reaction

staining reaction

granulocyte system

blasts

Well differentiated ( )

Poor differentiation (-)

mature granulocytes

Neutrophils ( )

Eosinophils ( ) between granules

Basophils ( ) on granules

Red blood cell system: Uniform (-)

Other departments (-)/( )

Judgment of results: Positive results are hemorrhagic red granules, lumps or diffuse red in the cytoplasm.

clinical significance

red blood cell system

( ): Erythemia, erythroleukemia, myelodysplastic syndrome, severe iron deficiency anemia, thalassemia major

(-): Megaloblastic anemia, aplastic anemia, other hemolytic anemias

Neutrophil alkaline phosphatase staining (NAP)

NAP mainly exists in neutrophilic mature granulocytes, so neutrophilic mature granulocytes are positive and other cells are basically negative. Positive reactions are manifested by the appearance of brown-black or brown-red particles in the cytoplasm.

Positive rate: Continuously observe 100 mature neutrophils under an oil microscope and record the percentage of positive cells. Positive rate＜40%

All positive reaction cells are graded ( ) to ( ) one by one according to their reaction intensity. Multiply the percentage of each grade by the series and then add them up to get the integral value 30-130 points.

clinical significance

1. Chronic granuloma ↓/0 Leukocytosis ↑ 2. PNH activity ↓ Aplastic activity ↑ 3. Acute granuloma ↓ Urgent lymphoma ↓ /Normal Acute lymphadenitis ↑ 4. Bacterial infection ↑ Viral infection (-) No change

Increased NAP points: seen in bacterial infections, leukemia-like reactions, aplastic anemia, certain myeloproliferative diseases, and chronic myelogenous leukemia

Decreased NAP score: seen in the chronic phase of chronic myelogenous leukemia, paroxysmal nocturnal hemoglobinuria, myelodysplastic syndrome, malignant histiocytosis, etc.

Iron staining

Extracellular iron is mainly present in macrophages of bone marrow granules, and intracellular iron is mainly present in intermediate erythrocytes, late erythrocytes and erythrocytes.

extracellular iron

Divided into type I, type II, type III, type IV and ring sideroblast erythrocytes

Ring sideroblasts refer to erythrocytes containing ≥6 iron particles in the cytoplasm and arranged more than 1/3 of the way around the nucleus.

Reference interval: extracellular iron ( ) ~ ( ), intracellular iron positive rate 12% ~ 44%, mainly type I, a few type II, no ring sideroblast red blood cells and sideroblast red blood cells

clinical significance

lack of poverty

vagina

Inside fe ↓/0

Fe young and poor

外fe↑

Inside fe↑ appears ring iron red

NPS

外fe↑

内fe↑