222 DYEING 



if 5%, to 2-3. Now 5% chrome alum and 5% iron alum by them- 

 selves (p. 212) are considerably more acid than 5% potassium 

 alum acidified in this way; yet their low pH does not prevent 

 dyeing. 



Another problem is presented by the peculiar resistance of the 

 tissue/iron/haematein complex to acidity. Quite strong acid must 

 be used to break this bond. This fact is very useful in microtech- 

 nique, for we can use weak acids in subsequent procedures without 

 removing the lake from chromatin ; but the reason for the difference 

 from other lakes is not obvious. We have not yet a full explanation 

 of the action of acids in loosening the tissue/mordant and mordant/ 

 dye bonds that are formed in the ordinary processes of micro- 

 technique. It is probable that we should have had a better insight 

 into it if there had been any process corresponding to differentia- 

 tion in the dyeing of textile fibres. In industry, however, there 

 would be no purpose in trying to colour one part of one cell (in 

 wool, for instance) with a dye, while leaving another part colour- 

 less; and that is the whole purpose of differentiation in ordinary 

 microtechnique. 



Benda ^^ long ago distinguished oxydirenden from einfach 

 losenden differentiators of mordant dyes. Einfach is not a happily 

 chosen word, yet the distinction is on the whole a useful one. 

 Potassium permanganate has been used as an oxidizing differentia- 

 tor, and ordinary bleaching agents are also available for the pur- 

 pose. The dye is presumably oxidized to a colourless substance. 

 The effect is one of differentiation, because certain objects contain 

 so much dyed matter that they still hold plenty of it when the 

 background has lost all visible trace. It has also been suggested ^^^ 

 that oxidizing differentiators may act on the metal of the mordant. 

 Chromium trioxide has been used as a differentiator of iron 

 haematein.^* Here one cannot be certain of the relative roles played 

 by oxidation and acidity. The same applies to picric acid, which 

 is a moderate oxidizer and a weak acid. It differentiates iron 

 haematein slowly. ^^' 



Another oxidizing differentiator is potassium ferricyanide. This 

 is used after tissues have been mordanted with potassium di- 

 chromate and then dyed with haematein. The technique was intro- 

 duced by Weigert ^^^ for the colouring of myelin, and subsequently 

 perfected as a histochemical test for phospholipids. ^^^'^^ The 

 mordant is anionic chromium, which acts very differently from the 

 cationic complexes that we have been considering in this chapter. 



