5µ paraffin sections of neutral buffered formalin fixed tissue are suitable.
Other fixatives are likely to be satisfactory.
Most trichrome stains benefit from picric acid or mercuric chloride fixation.
Formalin fixed tissues may benefit from secondary fixation of sections in Bouin's fluid.
Bring sections to water via xylene and ethanol.
Oxidise in chromic acid for 40-60 minutes.
Rinse with tap, then distilled water.
Place into Schiff;s reagent for 15 minutes.
Place into sulphurous acid rinses, 3 changes of 2 minutes each.
Wash with running tap water.
Counterstain with hemalum for 1 minute, and blue
Dehydrate with ethanols.
Clear with xylene and mount with a resinous medium.
Glycogen, mucin – red
Fungi – red
Nuclei – blue
Modern practice is to leave out the sulphite rinses and wash with large amounts of tap water.
A progressive hemalum should be used as counterstain because regressive hemalums sometimes stain
Mucins are not usually as dark as with a PAS.
Applying chromic acid for too long weakens staining due to continued oxidation of the aldehydes
McManus, J. F. A. and Mowry, R. W., (1960) Staining Methods Histologic and Histochemical
Harper & Row, New York, NY, USA.