Heidenhain's AZAN trichrome stain

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. Unfortunately, none of Mallory's publications (they go from 1891 to 1938 ) 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. 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.