Heidenhain's AZAN trichrome stain

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Trichrome stains are staining methods in which three anionic dyes are used, in conjunction with either phosphomolybdic acid (PMA), phosphotungstic acid (PTA), or a mixture of these heteropolyacids. Probably the first trichrome method was that of Frank B Mallory, an American pathologist, first published in 1900.[1] Unfortunately, none of Mallory's publications (they go from 1891[2] to 1938[3]) provide any explanation of the rationales of either his trichrome or his phosphotungstic acid-haematoxylin (PTAH) method. Nobody knows why Mallory introduced heteropolyacids into microtechnique.

Mallory's trichrome method, using acid fuchsine followed by a solution containing PTA, orange G and aniline blue, provides dark red nuclei, orange erythrocytes, and blue collagen fibres, cartilage matrix and mucus.[4] In 1915, M. Heidenhain introduced azocarmine G in place of the acid fuchsine of Mallory's method. Heidenhain also introduced visually controlled destaining to provide for different colours in cell nuclei (dark red), collagen (blue) and a variety of colours in cytoplasm.


  1. ^ Mallory FB (1900) A contribution to staining methods. Journal of Experimental Medicine 5: 15-20.
  2. ^ Mallory FB (1891) Phospho-molybdic acid-haematoxylin. Anatomischer Anzeiger 6: 375-376.
  3. ^ Mallory FB (1938) Pathological Technique. New York: Hafner.
  4. ^ Kiernan JA (2008) Histological and Histochemical Methods. Theory and Practice. 4th ed. Bloxham, UK: Scion. ISBN 978-1-904842-42-2