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Overview of Gram Stain of Hans Christian Gram

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Hans Christian Gram is a Danish Bacteriologist who acquired his biological knowledge at the University of Copenhagen, where he mastered the study of plants and the use of the microscope. Following graduating from the medical school in the year 1883, Hans Christian Gram travelled throughout Europe before settling in Berlin, where he carried out most of his research studies. One of the most renowned findings of Hans Christian Gram is the ‘gram stain’ procedure which has been used ever since with a few modifications to recognize two major types of bacteria, the ‘gram positive’ and the ‘gram negative’ forms.

Origin of gram stain

The findings which lead to the formulation of ‘gram staining’ is the result of studying human lung tissues obtained from patients died of pneumonia by Gram as a pathologist. During his observations, Gram noted that certain bacterial cells took up certain stains and retained while the other bacteria failed to achieve the same. However, the procedure adapted by Gram is somewhat different to the ‘gram stain’ process practiced by modern day scientists. Even then, the initial few steps are relatively the same.

Steps in the initial gram stain procedure

The procedure adapted by Gram included drying a fluid smear on a glass slide by holding it over a burner flame. He then poured Gentian violet solution over the dried smear. Following rinsing the same in water, Gram added Lugol’s solution (potassium triiodide solution) in hope of fixing the dye. It is after this step that Gram poured ethanol over the slide to wash away the excess dye. When examined under the microscope after bleaching with Ethanol, Gram observed that some bacteria retained the purple color, while certain bacteria became bleached or decolorized. The bacteria, which retained the purple color, were named as ‘gram positive’ and the ones which became decolorized were given the name ‘gram negative’. It should also be stated that the initial gram stains were performed on streptococcus penumoniae and Klebsiella pneumonia.

Modifications and the mechanism of gram stain

However, it was the German pathologist Carl Weigert who developed a counter staining using safranin, which is now regarded as the last step of the ‘gram staining’ technique.

Although not really understood at the time of its discovery, the mechanism behind the ‘gram stain’ is now known to the scientists. Thus, it has been recognized that gram-positive bacteria are able to retain the stain as its mesh-like cell wall which is 50 to 90% made out of peptidoglycan stains purple while the gram negative bacterial cell wall which is only 10% peptidoglycan stains pink. At the same time, having a lipid laden outer membrane which is separated by a periplasmic space also contribute towards the difference between gram positive and gram negative staining.

Thus, although the main concepts of ‘gram stain’ have not been changed over the years, a few additions to its main procedure have promoted the effectiveness of the ‘gram stain’ as a preferred diagnostic method in certain disease processes.

More about this author: Dr Pandula Siribaddana

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