Starry Starry Night? No, Warthin-Starry Stain!

Continuing in our series, the Warthin-Starry (WS) stain is another type of special stain often in the histology lab to detect certain types of organisms.

Why Use Warthin-Starry?

Some Gram-negative organisms do not stain well by the Gram stain technique. These include spirochaetes (such as Helicobacter, Leptospira, Borrelia, and Treponema species) and small bacilli (including Campylobacter, Bartonella, and Legionella species).

Silver stars

The WS stain is considered the best staining technique to detect these types of organisms. It was first developed in 1920 by American pathologists Aldred Scott Warthin and Allen Chronister Starry, and is commonly referred to as a “silver stain” because it is a silver nitrate-based staining method.

General Principles of the Stain

The procedure involves 2 basic steps:

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1. Slides of tissue sections are immersed in an acidified aqueous solution of silver nitrate (optimum pH 3.5-4.0). Two reactions occur during this time. Firstly, large numbers of silver ions form bonds with protein molecules throughout the tissue. Secondly, smaller numbers of silver ions are reduced to silver atoms at specific sites in the tissue- within or on the surfaces of organisms. These metallic silver deposits are tiny particles known as “nuclei” (but these are not the same as cell nuclei).
2. Slides are immersed in a reducing solution that typically contains hydroquinone, gelatin, and a lower concentration of silver nitrate. This solution acts as a type of “developer”: the gelatin sequesters silver ions, slowing the rate at which they are reduced to metal by the hydroquinone. This slow reduction of the silver ions is catalysed by the nuclei that were formed in the first step. Eventually, these tiny nuclei enlarge as more and more silver deposits on the organisms, until enough is present to make the organisms visible as black objects under the microscope.

Consequently, the end product of the reaction is such that the organisms stain dark brown to black. The lower concentrations of silver deposits stain golden brown and provide a contrasting background staining of the architecture of the tissue.

Who Uses Warthin-Starry Stain?

This stain is used widely for both diagnostic and research purposes. For example, researchers investigating diseases such as leptospirosis, cat scratch disease, and Helicobacter-associated gastric cancer may routinely examine WS-stained tissue sections to detect the presence of organisms in tissues of interest.

In diagnostic labs, pathologists often use this stain in conjunction with a Gram stain to help answer questions that may arise after findings in routine haematoxylin and eosin-stained sections suggest bacterial involvement.

How frequently do you use a Warthin-Starry stain in your lab?

Nicola Parry

Dr Nicola Parry graduated from veterinary school at the University of Liverpool and spent several years working in mixed general practice in the UK before moving to the USA to pursue Anatomical Pathology at the American College of Veterinary Pathologists.

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