Stains In Focus: The Development of the Romanowsky Technique

Introduction to Romanowsky Staining

The Romanowsky Type Stain, named after Dmitri Romanowsky, are used worldwide to examine blood and bone marrow specimens. By combining basic and acidic dyes, they give a rich spectrum of colours to cellular components. This “Romanowsky effect” of vivid blues, purples and pinks makes nuclei, cytoplasm, and granules easily distinguishable, providing excellent diagnostic detail.

In this blog, Atom Scientific will cover the history of the Romanowsky Stains, and the development of stains related or derived from this principle.

The History of Romanowsky Staining

The Romanowsky stain originated in 1891 when Russian physician Dmitri Romanowsky introduced a staining method using a mixture of eosin and a partially oxidised methylene blue. This combination unexpectedly produced a striking polychromatic effect —where nuclei stained blue to purple and cytoplasm appeared pink, laying the foundation for what would become known as the "Romanowsky effect".

Subsequent refinements by pioneers such as Leishman, Giemsa, Wright, and May-Grünwald focused on stabilising the dye mixture and enhancing reproducibility. A key development was the use of azure dyes—oxidation products of methylene blue—which improved contrast and consistency. The combination of eosin Y with azure B became essential to the formulation, enabling precise differential staining.

Refining Romanowsky’s Technique

Following Romanowsky’s initial discovery, further progress was driven by the need for greater stability, clarity, and reproducibility in staining. Pioneers such as Leishman, Giemsa, Wright, and May-Grünwald sought to overcome these limitations by refining compositions.

Giemsa Stain

The development of the Giemsa stain goes back to Germany in 1867, where Gustav Giemsa, a German chemist and pharmacist, refined earlier Romanowsky-type formulations. Giemsa’s key innovation was using glycerol as a stabilising agent, allowing for a more consistent and longer-lasting staining solution. The final formulation included methylene blue, eosin Y, and azure B dissolved in a methanol / glycerol mixture producing a stain that could deliver high-contrast, polychromatic results.

Fields Stain

Field’s stain was developed by British pathologist John William Field in the early 20th century as a rapid Romanowsky-type stain. It was designed specifically for quick field diagnosis of malaria and other blood parasites in military and tropical environments.

The method uses two solutions: Field’s Stain A (methylene blue and azure) and Field’s Stain B (eosin). It is one of the original Romanowsky stains, for the detection of malarial parasites in blood films, it can also be further modified for the rapid staining of thin blood films.

Jenner Stain

Introduced by London physician Leonard Jenner in the late 19th century, the Jenner stain was one of the earliest Romanowsky-type formulations. It consisted of eosin and methylene blue dissolved in alcohol, creating a stable compound that provided good nuclear and cytoplasmic detail. Although now largely superseded by more refined stains, Jenner’s work laid important groundwork for subsequent innovations like Leishman and Giemsa stains.

Leishman Stain

Developed by Scottish pathologist William Boog Leishman in 1903, the Leishman stain was a modification of the Romanowsky method, designed for quicker and more efficient blood film preparation.

By dissolving methylene blue and eosin in methanol,Leishman created a rapid, effective stain that provides excellent stain quality and is used to differentiate and identify leucocytes, malaria parasites, and trypanosomes.

May-Grunwald-Giemsa Stain

The May-Grunwald-Giemsa (MGG) stain evolved through contributions from Richard May, Ludwig Grünwald, and Giemsa. May introduced a methylene blue and eosin-based stain in 1902 to improve white blood cell visualisation. Grünwald enhanced the method for broader diagnostic use, refining staining times and dye balance, leading to the May-Grünwald stain. Giemsa further developed a formulation with azure B and eosin Y, stabilised in glycerol / methanol. The combination of May-Grünwald and Giemsa stains created the MGG technique, offering enhanced diagnostic clarity.

Wright and Wright Giemsa Stains

The Wright stain was developed in 1902 by James Homer Wright, an American pathologist seeking to improve the Romanowsky staining method for blood and bone marrow smears. Wright’s key modification was fixing and staining cells simultaneously

by dissolving a mixture of methylene blue and eosin in methanol. This simplified preparation and significantly reduced staining time, making it a popular choice for clinical haematology.

Atom Scientific’s Romanowsky Stains

Atom Scientific is proud to be the only UK manufacturer offering IVDR-registered ISO13485-compliant stains and stain kits. We provide the most comprehensive range of UK-manufactured, pre-validated, IVDR-certified products for diagnostic use, all from a single trusted source. Choosing Atom Scientific means choosing full compliance and peace of mind.

Our ready-to-use, validated stain kits are designed to support the modern BMS by saving valuable time and delivering consistent, reliable results within proven protocols. Each kit offers complete batch and raw material traceability, ensuring a robust audit trail whenever required. In addition, we provide full technical support to help you maximise the performance and reliability of your staining processes.

Conclusion

From Dmitri Romanowsky’s original discovery in 1891 to the numerous refinements made by pioneers such as Giemsa, Wright, Leishman, and others, Romanowsky-type stains have become a cornerstone of diagnostics. Their unique ability to differentially stain blood cells, parasites, and bone marrow elements with vivid clarity continues to make them essential in both clinical and research settings.

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