Quarterly Journal of Microscopical Science, Vol s3-103, 493-517, Copyright © 1962 by Company of Biologists

Experiments on the Action of Mordants 2. Aluminium-Haematein

¹ Cytological Laboratory, Department of Zoology, University Museum, Oxford

Unmordanted haematein in aqueous solution varies in colour from yellowish orange at pH 3 to bluish magenta at pH 9.

In the presence of aluminium sulphate, the colour of solutions of haematein at any particular pH depends on the number of aluminium atoms to each molecule of haematein. At pH 4.6 the colour varies from yellowish orange through dull redto rose as the number of aluminium atoms to each molecule of haematein is increased from to 8. The facts suggest that such solutions contain both uncombined haematein and an aluminium-haematein compound.

From about pH 5 upwards the aluminium-haematein compound is insoluble. Although the dominant colour is blue, a considerable proportion of red light is transmitted.

The blue colour attaches itself to tissue-constituents like a typical cationic (basic dye. It combines principally with the nucleic acids of chromatin; but if these are extracted from the tissues, it attaches itself to the acidic proteins of chromatin, and the appearance given is much the same.

In differentiation by acid, the mordant is set free from the tissue.

Chromatin that has been reddened by acid during or after dyeing becomes blue from about pH 5 upwards. Neutrality or alkalinity is not necessary for blueing.

The dull red aluminium-haematein compound present in acid solutions acts (like the blue) as a cationic dye.

When aluminium and haematein are used (as iron and haematein usually are) by the two-bath method, chromatin is dyed blue but collagen and certain other tissue-constituents yellowish. An exhaustive series of experiments shows that the yellowish colour is due to unmordanted haematein.

At room temperature haematein is about 17 times more soluble in ethylene glycol than in ethanol, and 21 times than in water. In 25% aqueous solution ethylene glycol is a suitable solvent for practical use. A solution in this solvent containing aluminium and haematein in the proportion of 8 atoms of the metal to one molecule of haematein may conveniently replace Mayer's Hämalaun and similar mixtures.