Iron Stain

Iron Stain

Assay-kits

Product code: ACMBS-48
Specifications: 5Tests, 20Tests, 100Tests (Box)
Intended use: This kit is for staining bone marrow cell
Store temperature: 2℃～8℃
Expiry date: 12 Months

More Details

PrincipleMain componentsSample requirementTest procedureResult interpretationMethod limitationPrecautions

Principle

Normal bone marrow exists in a certain amount of stored iron, in the form of hemosiderin stored in tissue macrophages, available for nucleated red blood cells using synthetic hemoglobin, which exists in red blood cells outside the storage of iron known as extracellular iron, part in the late erythrocyte and a small number of mature red blood cells also contain iron particles, respectively, iron granule red blood cells and red iron cells, they belong to intracellular iron. Acid potassium ferrocyanide with the cells, the occurrence of Prussian blue iron reaction, the formation of blue ferrocyanide iron precipitation, located in the iron-containing parts.

Main components

Reagent composition	Main ingredient
1. Fixative	Formaldehyde
2. Potassium ferrocyanide solution (solution A)	Potassium ferrocyanide
3. Hydrochloric acid solution (solution B)	Hydrochloric acid
4. Safranin solution (solution C)	Safranin

Sample requirement

Fresh bone marrow cell smear.

Test procedure

A. Working solution preparation:

Dip stain working solution preparation (50ml volume dye vat: A dip stain working solution is 40ml, at the same time can place at least 8-10 smears.): Add each of 20 ml of A and B solution into dye vat (prepare dye vat by yourself), mix thoroughly.

Note: The above is the preparation amount of the dip staining working solution for the 50ml volume dyeing vat. Each laboratory can scale up or down according to the volume of its own dyeing vat.

Drops stain working solution preparation (A drops stain working solution is 2 ml): Add each of 1 ml of A and B solution into test tube, mix thoroughly.

B. Staining procedure:

1. Air dried bone marrow smears then fixed with fixative (Return to room temperature before use and shake well) for 30 to 60 seconds, rinse with distilled water, wait until dry or dry with filter paper.
2. Drop or dip into working solution on the slide for 60 minutes (if the temperature is low in winter ,it must be incubated in a 37℃ water bath). Rinse with distilled water for 5 minutes, air dry or dry with filter paper.
3. Counterstain with solution C (Be sure to shake well before use!) for 1～2 minutes. Rinse with distilled water. Dry the smear for microscopic examination.

Result interpretation

Iron can be stained as blue granules, small deposits or small masses.

Extracellular iron: Observe the folded marrow granules under low power magnification before using immersion oil lens for examination.

-: No blue iron granules are found.
+: Few iron granules are found, or only few iron deposits are found.
2+: Lots of iron granules and iron deposits.
3+: Lots of iron granules or deposits and a few small masses.
4+: Lots of iron granules, iron deposits, and many small masses.
Intracellular iron: Count 100 nucleated erythrocytes and record the percentage of positive cells which are found as blue granules in cytoplasm. Find the numbers of iron granules in cells, sizes, colors and if siderocytes INS-CB010-EN-D0 in ring shape are present.

Reference value:

The amount of iron granule and iron deposit in extracellular iron of normal human bone marrow varies in different degrees. Most people are "+", few are "2+". Positive rate of normoblast containing iron granule is 12～14% which are mainly composed by polychromatophilic normoblast and orthochromatic normoblast. Most positive normoblasts contain 1～2 small and irregular iron granules. Few of them contain 3～5. In general, normoblast and ringed sideroblast containing more than 5 iron granules are hardly found.

Method limitation

Only use for morphological staining to observe.

Precautions

1. 5 Tests only use on drops stain. 20 Tests and 100 Tests can be used for drip staining and dip staining.
2. Return to room temperature before use and shake well. Please thoroughly mixed A and B solution, all containers must be clean.
3. Prepare working solution prior to use.
4. Slides and apparatus should be clean and are not contaminated by iron. (By iron removal pretreated in advance, especially the glass slides.)
5. Tightly cap Potassium ferrocyanide in a brown bottle. Potassium ferrocyanide will be denatured easily while exposure to air or sunlight.
6. Select and stain bone marrow smear that contains more marrow granules. Observe extracellular iron and intracellular iron in the same smear.
7. Iron Stain can be used in smear that is well stained with Wright Stain and with no sediment. Counterstain for these slides are unnecessary.
8. Rinse the smear thoroughly before counterstain, otherwise lots of acicular crystals shall produce.
9. Tightly cap the reagent bottle immediately after use to avoid evaporation.
10. This kit must be used by professionals and the interpretation of results.
11. Should carefully read the instruction manual before using. Can only be used before expiration date, and good personal hygiene protection.
12. After use, disposal of waste should comply with the hospital or the environmental protection department requirements.
13. Production lot number and expiration date on the package.

References

1. The PRC Ministry of Health Medical Administrative Department. National Clinical Laboratory Procedures (M) version 3. Nanjing: Southeast University Press, 2006.

2. National Health and Family Planning Commission of the People's Republic of China Medical Administration Hospital Authority. National Clinical Laboratory Procedures [M]. 4th ed. Beijing: People's Medical Publishing House, 2014.

3. Gradwohl's Clinical Laboratory Methods And Diagnosis, 8th, ed.

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