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Laboratory Methods

	Antifungal Susceptibility Testing

	Culture Techniques and Media

	Microscopy Techniques and Stains

	Specimen Collection and Processing

School of Molecular & Biomedical Science
THE UNIVERSITY OF ADELAIDE
AUSTRALIA 5005

Contact:
Dr David Ellis

Email

Telephone:
+61 8 8161 6459
Facsimile:
+61 8 8161 7589

Potassium Hydroxide (KOH) with Parker Ink.

For the direct microscopic examination of skin scrapings, hairs, nails and other clinical specimens for fungal elements.

Method:

Dissolve the KOH in water, then add the glycerol and Parker ink. The glycerol prevents crystallisation of the reagent and prevents the specimen from drying out.

Potassium hydroxide 10 g

Glycerol 10 ml

Parker Quink Permanent Blue Ink 10 ml

Distilled water 80 ml

Making mounts for microscopy:

1. Using an inoculation needle remove a small portion of the specimen, especially from any necrotic or purulent areas, and mount in a drop of KOH on a clean microscope slide.

2. Cover with a coverslip, squash the preparation with the butt of the inoculation needle and then blot off the excess fluid. Gently heat by passing through a flame two or three times. Do not boil.

3. When the specimen has cleared, which may take from 20 minutes for skin scrapings to several hours for nail scrapings, examine microscopically for the presence of faintly blue stained fungal elements.

Note: negative specimens should be kept and re-examined the next day to avoid reporting false negative results due to delayed clearance and staining of the specimen.

Comments: the most widely used technique, but may take time for the specimen to clear and stain. Preparations last a long time and may be kept until the culture result is known.