INTRODUCTION
The Romanowsky stains are used to stain smears in order to provide better cellular definition and also identify blood parasites such as haemobartinella.
They use 2 dyes:

• HAEMATOXYLIN - which stains basic tissue blue.
• EOSIN - which stains acidic tissue red.

There are 3 main types:

• DIFF QUIK - very quick.
• GIEMSA - ideal for staining haemobartinella.
• LEISHMAN'S - excellent cellular definition.

EQUIPMENT

• Lab coat and protective gloves.
• Sink.
• Drainer (for Giemsa and Leishman's).
• Slide rack.
• Paper towel to protect work surfaces from the stains.
• Pipettes (for Giemsa and Leishman's).
• Buffered distilled water.
• Alcohol/methanol (for Giemsa).
• Stains.
• Pencil for labelling smears.
• Watch or timer

METHOD

1. Put on lab coat and protective gloves; ensure that long hair is tied back and any jewellery removed.
2. Label the smears in pencil (if this has not already been done); information should include the date, owner's surname and animal identification.
3. Ensure that you have a flat, clean, dry surface upon which to work.
4. Handle the slides correctly; ie at the edges only.

Diff Quik.

• Place the slide in the fixer (pale blue solution) for 5 seconds.
• Place the slide in the red solution for 30 seconds.
• Place the slide in the dark blue solution for 10 seconds.
• Rinse gently with distilled water.
• Air dry and place on a slide rack.

Giemsa

• Fix slide in alcohol/methanol for 3 seconds.
• Place the slide the correct way up on a drainer over the sink.
• Use a pipette to flood the slide with Giemsa stain.
• Leave for 30 minutes (if this task is given to you in the examination - you will be told to pretend that the time is up).
• Rinse gently with distilled water.
• Air dry and place on a slide rack.

Leishman's.

• Place the slide the correct way up on a drainer over the sink.
• Use a pipette to flood the slide with Leishman's stain.
• Allow to stand for 15 minutes until a metallic scum appears on the solution surface (again, if this is an examination task - you will have to pretend that the 15 minutes is up).
• Rinse gently with buffered distilled water.
• Air dry and place on a slide rack.

EXAMINATION

Blood smears are examined under oil immersion (high power).

NB. Please refer to Module 4 - Blood Cells for further information.