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White Blood Cell Staining Methods

What are we distingushing by staining?

In the previous article, the donated white blood cell may cause the following potential risks; immune response, leading to fever, chills, and the reduction of the donated blood samples.

It is necessary to check whether white blood cells are present in the reserved blood packs.

Romanowsky stain

Types of white blood cells are died by romanowsky stain

Red blood cells (RBCs) and white blood cells (WBCs) can be distinguished by staining using a variety of techniques. One common method is to use a Romanowsky stain, such as Wright's stain or Giemsa stain. These stains bind to different components of the cells, causing them to appear different colors. RBCs will typically stain pink or red, while WBCs will stain blue or purple.

Fluorescent stain

PI only stains the non-viable cells which means that dead cells are permeable

Another method for distinguishing RBCs and WBCs is to use a fluorescent stain, such as acridine orange(AO) and pyridine iodide(PI). This stain binds to nucleic acids (ex. DNA), which are found in the nucleus of the cell. RBCs do not have a nucleus, so they will not stain with AO or PI. WBCs, on the other hand, will stain bright green or red depends on the staning materials.

Staining is a valuable tool for identifying and differentiating different types of cells. By using different staining techniques, it is possible to quickly and easily distinguish RBCs and WBCs, as well as other types of cells.

If you would like to know more about staining technique check this article:

What staining techniques should you choose?

There are different clinical findings you can earn by the two different staining methods.

Here are the common clinical findings using Romannowsky staining and Fluorescent stains on white blood cells:

  • To identify count different types of white blood cells.

  • To assess the morphology of white blood cells

  • To detect abnormalities in white blood cells, such as those seen in leukemia, anemia, and infections: Blood and bone marrow pathology, Detection of malaria and other parasites

  • To monitor the response to treatment for blood disorders.

Those two staining methods are valuable tools for diagnosing and monitoring blood disorders safely.

However, there are differences in the two methods. In this comparison table, PI is used for representing fluorescent stains.


Romanowsky stain

Pyridine iodide stain




Dyes used

Azure B and methylene blue

Pyridine iodide


More sensitive

Less sensitive


More time-consuming ( > 15 mins)

Faster ( < 5 mins)

Ease of preparation

More difficult


Preferred use

Diagnosing blood disorders

Rapid screening

Distinguishing WBCs and RBCs is vital for assessing risks and ensuring donated blood quality. Romanowsky stains differentiate cells by color, while fluorescent stains detect nucleic acids in WBCs. These techniques identify WBC types, assess morphology, detect abnormalities, and monitor treatment response in blood disorders. Both staining methods are valuable for diagnosis, but fluorescent stains have specific advantages. Overall, they are essential tools for safe diagnosis and monitoring of blood disorders.


  1. Horobin RW. How Romanowsky stains work and why they remain valuable - including a proposed universal Romanowsky staining mechanism and a rational troubleshooting scheme. Biotech Histochem. 2011 Feb;86(1):36-51. doi: 10.3109/10520295.2010.515491. PMID: 21235292.

  2. NanoEntek, Ch.4-2 Fluorescence dye solution (PI/AO/DAPI), Accessed Jun. 2023.


Residual Leukocyte Counter


Residual White Blood Cell Counter

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