universe84a

Introduction of Albert’s Stain 

Albert’s stain is a metachromatic dye and is useful to stain Corynebacterium diphtheriae.

Principe of Albert’s stain 

It stains the volutin or metachromatic granules of Corynebacterium species. Volutin granules appear as bluish-black around bodies at the end of bacilli and the bacilli appear green. A typical Chinese letter arrangement of the different bacilli with the volutin granules is the indication of Corynebacterium diphtheriae being positive in the throat swab smear.

Preparation of Albert’s stain 

It is of two types.

• Albert’s stain A
• and Albert’s stain B

Composition of Albert’s Stain A

• Toluidine blue: 1.5 gm
• Malachite green: 2.0 gm
• Glacial acetic acid: 10 ml
• 95% ethanol: 20 ml
• Distilled water (D/W): up to 1000 ml

Preparation

• Dissolve both the dyes in alcohol.
• Add water and acetic acid.
• Make up the final volume with distilled water up to 1000 ml.
• Mix thoroughly.
• Filter and store in a brown color bottle with a label.

Albert’s  Stain B (Albert’s iodine) Composition

• Iodine: 6.0 gm
• Potassium iodide (KI): 9.0 gm
• Distilled water: 900 ml

Preparation

1. First, dissolve the potassium iodide in little quantity of water (50 ml).
2. Then dissolve the iodine in the above solution.
3. Prepare final volume up to  900 ml.

Test procedure

1. Take the throat swab from the affected site checking for the presence of the membrane that is normally present in diphtheria.
2. Prepare the smear.
3. Leave for air drying and then heat fix.
4. Cover the slide with Albert’s stain A and allow it to act for 3-5 minutes.
5. Drain the solution, do not wash.
6. Flood the smear with Albert’s iodine i.e.  Albert’s Stain B for 1 minute.
7. Rinse with water, blot dry and examine under oil immersion objective.

Result and Interpretation

The granules stain bluish black but the protoplasm stains green. Other organisms appear light green. In brief-

• Granules: bluish-black
• protoplasm: green
• Other organisms: light green

Further Readings

• Topley and Wilson’s microbiology and microbial infection Topley and Wilson’s microbiology and microbial infection – Bacteriology-2-10th Edn.
• Manual of Clinical Microbiology   -Patrick R. Murray -8th Edn.
• Bailey and Scott’s  Diagnostic Microbiology -13th Edn.
•  Mackie & Mc Cartney  Practical Medical Microbiology- 14th  Edn.
• Diagnostic Microbiology -Connie R. Mahon & George Manuselis
• Koneman Color Atlas and Textbook of Diagnostic Microbiology-6th  Edn.
• Jawetz Melnick and Adelberg’s Medical Microbiology- 25th Edn.
• Lippincott’s –Illustrated- review-Microbiology-3rd Edn.
•  Mandell’s Infectious Disease-7th Edn.
• Bergey’s Manual of Systemic Bacteriology – 2nd  Edn.
• Medical Microbiology-The Practice of Medical Microbiology Vol-2-12th Edn. –Robert Cruickshank
• District Laboratory Practice in  Tropical Countries  –  Part-2-   Monica Cheesebrough-   2nd Edn Update