206 DYEING 



the DNA. If, however, formaldehyde was used as fixative, the 

 protein of the chromatin will have been rendered acidic and there- 

 fore will have taken up little carmine from alkaline solution ; there 

 will consequently be little dye present in the chromatin to link up 

 with DNA on subsequent acidification. 



In routine preparations we usually want to dye the chromatin 

 in one colour with a basic dye (or basic dye-lake, p. 207) and the 

 cytoplasm in a contrasting colour with an acid dye. What does this 

 involve? Ideally, the DNA must be struck off by the fixative from 

 the protein with which it was combined, and precipitated instantly 

 without change of position in a form in w^hich it will combine 

 readily with basic dyes ; the protein of the chromatin must have a 

 low affinity for acid dyes, so that the colour given by the basic dye 

 shall not be masked ; and the cytoplasm must have little affinity for 

 basic dyes, but link strongly with acid ones. The fixative that 

 leaves tissues most nearly in this condition is chromium trioxide, 

 though chromatin fixed by this substance has not so strong an 

 affinity for basic dyes as one would wish, and retains some affinity 

 for acid ones. Acidified potassium dichromate acts similarly to 

 chromium trioxide. (See p. 132.) 



Many fixative mixtures probably owe their continued popularity 

 largely to the fact that they happen to leave the iso-electric points 

 of the tissue-constituents in convenient positions; that is to say, in 

 positions in which it is easy, without troubling to use buffers, to 

 colour the chromatin with a basic dye and the ground cytoplasm 

 with an acid dye of contrasting colour. Some shift of the iso- 

 electric point towards higher pH would be helpful towards this 

 end, w^hich is realized by such ffuids as that of Zenker. ^^* If this 

 shift did not occur, ground cytoplasm would generally be too 

 basiphil to give good colour-contrast with chromatin. 



It is a special property of mercuric chloride that it leaves tissues 

 in a state in which they are readily coloured by dyes of all sorts (but 

 particularly by basic dyes and dye-lakes). No adequate explanation 

 of this familiar fact has been provided. One must suppose that the 

 tissues are fixed in such a form that they are readily penetrated by 

 large ions, and that many acidic and basic groups (especially the 

 former) are available for linkage with dyes. 



