Leishman stain – principle, procedure, results, advantages & disadvantages

everything about Leishman stain
everything about Leishman stain

Reading this article, you can improve your knowledge about Leishman stain. As you already know, Leishman stain is a universally accepted cheap staining method under WHO. This stain is mostly used in hematology and parasitology works. In hematology, Leishman stain used to identify blood cell components, like red blood cells, white blood cells(Neutrophils, Eosinophils, Basophils), platelets. In parasitology, this stain used to identify malaria parasites.

principle of Leishman stain

Leishman stain is a differential stain which use to stain various components of cells. This stain contains methylene blue and eosin. Methylene blue is a basic dye that stains acidic component of cells and eosin is an acidic dye which stains basic components of cells.

The procedure of preparing Leishman stain (stock solution) – procedure, storage, labeling

Procedure of Leishman staining

  • Firstly you have to prepare a blood smear (It should not be too thick and thin)
  • Blood drop should not be very large. If it is a large drop, your blood smear becomes thick.
  • stain is poured on it (Cover the blood smear)
  • Dilute after 1 minute with buffer for 7 minutes
  • After that washed with de-ionized water and dried
  • Finally, observe the stained blood smear under oil immersion objective.

Results under the microscope

  • Red cells – red to yellowish-red
  • Neutrophils – dark purple nuclei, pale pink cytoplasm, and reddish granules
  • Eosinophils – blue nuclei, pale pink cytoplasm, and orange-red – large granules
  • Basophils – Purple to the dark blue nucleus, dark purple-black large granules
  • Lymphocytes – dark purple nuclei, light blue cytoplasm
  • Platelets – violet-purple

If you see abnormal results,

  • Rouleaux formation
  • Bursting of leucocytes
  • The unclear nucleus in the intact leucocytes
Possible causes are,
  • Rouleaux was actually agglutination from too thick a blood drop
  • Stain dye is poor and needs fresh preparation
  • Water contamination

Advantages and disadvantages of Leishman staining

Advantages of Leishman staining

  • Easily available
  • Simple to use at any time for urgency
  • Easy to prepare
  • Cheap in expense
  • Staining time below 10 minutes
  • Nucleus and neutrophilic granules stain in clear violet color
  • Malaria parasite very clear to identify
  • Very good in contrast
  • Impressive view in oil immersion objectives
  • Stain is well stained

Disadvantages of Leishman staining

  • Evaporation of methanol (More chance of precipitation occurs)
  • Leishman stain get precipitated (On storage-long term:- have to be re-filtered)
  • Filtration changes the original composition

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