SOP 2: Centrifugal Faecal Floatation

The zinc sulfate [specific gravidity (S.G.) 1.18] centrifugal floatation procedure is suitable for the isolation and identification of a protozoan cysts and oocysts in canine and feline faeces, in particular cysts of Giardia duodenalis. Centrifugal floatation is also more sensitive for the isolation of heavier nematode eggs such as those of Trichuris vulpis and Spirocerca lupi, in which a heavier floatation solution with a SG of 1.25 is utilised (e.g. Sheather’s sugar solution). These methods are inexpensive; however, they do require the use of a centrifuge.

Reagents

  • Flotation solution (e.g., Zinc sulfate solution or Sheather’s solution)
  • Lugol’s iodine

Preparation of flotation solutions

Zinc sulfate solution (SG 1.18)

Dissolve 331 g zinc sulfate in 900 ml warmed distilled water (dH2O). Add more dH2O until the entire solution weighs 1180 g (this equates to a S.G. of 1.18). Mix solution and then check SG with hydrometer. Note: if zinc sulfate heptahydrate is used, then additional quantities will be needed (e.g., approx. 750 g).

Sheather’s solution     (SG 1.25)

To 355 ml hot water, add (while stirring) 454 g sugar. Add 6 ml formalin per 454 g sugar. Adjust to ensure SG is 1.25 using a hydrometer.

Procedure                                                                     

  1. Place ~2 g faeces into a wide-mouthed plastic disposable cup
  2. Add ~4 ml flotation solution to the jar and mix with faeces thoroughly
  3. Add a further 4 ml flotation solution to the jar and mix again
  4. Pour/filter this faecal suspension through a tea strainer into a new jar
  5. Empty the contents of the jar into a 10-15 ml test-tube supported in a rack or stand
  6. Centrifuge at 500 g for 10 min
  7. Carefully add more flotation solution until a positive meniscus forms at the top of the test tube and place a coverslip on top
  8. Stand for a further 5-10 minutes
  9. Carefully lift off the coverslip with the drop of fluid adhered to the bottom of it and place it on a microscope slide. Adding a drop of Lugol’s iodine to the slide before placing the coverslip on it can make the Giardia cysts easier to see
  10. Examine under a light microscope at low power (10x) for helminth stages and at high power (40x) for protozoal stages

Safety precautions

  • Wear lab coat and disposable gloves
  • Wash hands thoroughly when finished

Clean up procedures

  • Pour sodium nitrate into appropriate chemical waste container
  • Dispose of all slides and cover slips in a sharps container
  • Clean all equipment (tea strainer, glass test tubes) thoroughly with a 10% bleach solution
  • Wipe down work area with 70% Ethanol