Application of Albert Stain in the Diagnosis of Diphtheria

The Albert stain

The Albert stain is a specialized staining method primarily used for the identification of Corynebacterium diphtheriae. This bacterium is responsible for the disease diphtheria. The stain is useful for demonstrating metachromatic granules within the bacterial cells, which are characteristic of this pathogen.

Here’s an overview of the principle, material requirements, procedure, and results of the Albert stain:

Principle:

The principle of the Albert stain is based on the ability of certain bacteria (like Corynebacterium diphtheriae) to accumulate metachromatic granules (also known as Volutin granules) within their cytoplasm. These granules are composed of inorganic polyphosphate and exhibit a metachromatic property, meaning they change color when stained with certain dyes.

The Albert stain uses a combination of Albert's solution, which is a mixture of methylene blue and iodine. The iodine reacts with the polyphosphate in the granules, making them visible as dark blue or purple granules under a microscope.

Material Requirements:

1. Albert’s stain solution:

   • Methylene blue: 1 g

   • Iodine: 0.1 g

   • Potassium iodide: 0.1 g

   • Distilled water: 100 mL

2. Glass slide (for preparing the smear)

3. Microscope (to observe the results)

4. Bunsen burner (for heat fixing the smear)

Procedure:

1. Prepare a smear:

   • Prepare a thin smear of the sample on a clean glass slide. This sample is usually a clinical specimen suspected to contain Corynebacterium diphtheriae (such as a throat swab).

2. Heat-fix the smear:

   • Gently heat fix the smear by passing it through the flame of a Bunsen burner. Do this carefully to avoid overheating or distorting the bacteria.

3. Apply the Albert stain:

   • Add a few drops of Albert’s stain solution onto the heat-fixed smear.

   • Let it sit for about 5 minutes to allow the stain to interact with the bacterial cells and granules.

4. Rinse the slide:

   • Rinse the slide gently with distilled water to remove excess stain.

5. Dry the slide:

   • Allow the slide to air dry or blot it gently with filter paper.

6. Examine the slide:

   • Examine the stained slide under the microscope using the oil immersion lens (100x objective). Look for characteristic blue or purple metachromatic granules within the bacterial cells.

     
     
     

Results:

• Corynebacterium diphtheriae: The metachromatic granules will appear as dark blue to purple granules within the bacterial cells.

• Other bacteria: Most other bacteria will not show metachromatic granules or may not stain as distinctly with Albert's stain.

  
  
  

Interpretation:

• The presence of blue/purple granules inside the bacteria is an indicator of Corynebacterium diphtheriae.

• If no granules are present, it suggests the absence of this bacterium or the sample may not contain Corynebacterium diphtheriae.