THE DIRECT ATTACHMENT OF DYES TO TISSUES 201 



and 1 1 fibrin films took up far less acid and basic dyes respectively 

 than would be expected if all the charged groups of the protein in 

 fact reacted with dyes. The subject, however, is complicated. Dye 

 ions often aggregate (p. 238), especially at the low temperatures 

 used in the biological laboratory, and this would make stoichio- 

 metric proportions unlikely. Dyes ions with two or more charged 

 groups might not be able to use all of them to make attachment to 

 oppositely charged groups on the protein, because the latter might 

 be too far apart. 



The elTect of inorganic salts takes up a good deal of space in 

 works on textile dyeing, but is of less direct interest to the biologist. 

 In dyeing cotton and other cellulose fibres with certain direct 

 cotton dyes, it is usual to add sodium chloride or sulphate to the 

 dye-bath, as this greatly aids the uptake of dye. The dye-ions of 

 these direct cotton dyes carry the same (negative) electric charge as 

 the cellulose. In general, inorganic salts help dyeing in those 

 particular cases in which the electric charges oppose it, apparently 

 by favouring the near approach that is necessary for dyeing by 

 close-range bonds. *'^ On the contrary, they interfere with the 

 linkage between a dye-ion and an oppositely-charged group in the 

 object to be dyed, and also lessen the activity of many dyes by 

 increasing their tendency to fiocculation and thus lowering their 

 capacity to diffuse. Salts should not be used as buffers for dyes 

 without regard for these facts. 



Temperature affects dyeing in several ways.^^*' ^'^ High tem- 

 peratures increase the rate of diffusion of dye-ions and also reduce 

 any tendency they may have to aggregate into larger particles (p. 

 241), which would move more slowly and penetrate less easily. 

 High temperatures also loosen the covalent bonds that hold protein 

 chains together, and dissolve the disulphide links: thus the protein 

 becomes more easily permeable. At 100° C i hour may suffice 

 for wool to take up as much of a dye as it can hold; but if the 

 temperature be kept down to 20" C, 5 months may elapse before 

 equilibrium between wool and the same dye is reached. Tem- 

 perature does not have much effect, however, on the amount of 

 dye eventually taken up: rather more is taken up from cold solu- 

 tions, except in those cases in which a dye cannot enter at all in 

 the cold (p. 241). Most biological material is very much more 

 easily penetrated by dyes than wool is, and the temperature is only 

 raised above that of the laboratory when a dye has a special ten- 

 dency to fiocculation (e.g. azocarmine) or when a tissue-constituent 



