Gloves should be used and the procedure carried out in a fume cupboard

1 With unfixed specimens add a spoonful (about 1g) of faeces to about 7ml 10% formalin in a tube. Mix well with two applicator sticks. Wait about 30min for inactivation and fixing of specimen.

2 Filter a suitable amount of faeces suspension through one layer of dampened surgical gauze into a beaker. With very thin or mucoid specimens, see below.

3 Transfer the filtrate to a centrifuge tube. Adjust the volume to 7ml with formalin.

4 Add 3ml ethyl acetate and stopper the tube. Shake the tube for 15s on a vortex mixer or 30s by hand.

5 Centrifuge the stoppered tube at 1 000g for 60 s or for 5 min if investigation for Cryptosporidium is requested. There will now be four layers, the sediment, a formalin/debris layer, a fatty plug and an ethyl acetate layer.

6 Unstopper the tube with care because of positive pressure inside. Carefully loosen the fatty plug with an applicator stick. Pour off the plug and fluid layers leaving the sediment in the tube. A little fluid will run back to the sediment.

7 Dry the inside of the tube with a cotton swab. If the sediment appears too dry add a drop or two of 0.9% saline and mix well.

8 Leave the tube in the fume cupboard for a few minutes to allow any remaining ethyl acetate to evaporate.

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