(acid dye). In acid dyes, the auxochrome is generally either a hydroxyl 

 (—OH), carboxyl (— COOH), or sulfonic (— SO,H) group. The auxo- 

 chromes of basic dyes are usually an amino ( — NHo) group or one 

 of its derivatives. The chromatophores responsible for the colored 

 properties of dyes are also classified as being either acidic or basic. Ex- 

 amples of acidic chromatophores are the nitro ( — NO2) and quinoid 



(0=:<^ j>^0) groups. The basic chromatophores are the azo 



( — N=N — ) and indamin ( — N=) groups. The yellow acid dye, picric 

 acid, may be used to show the different types of groups present in a dye 

 (Figure 11-4). The color of picric acid is due to the nitro groups 

 (chromatophores), and its dyeing properties are attributed to the hy- 

 droxyl group (auxochrome) which is capable of forming salts with a 

 base. 



OH 

 0.,N 1 NO.. 



NOo 



Figure 11-4. Diagram of Molecular Structure of the Yellow-Acid Dye, 

 Picric Acid. 



Conventional Staining 

 A number of factors operate to influence the staining of cell struc- 

 tures. The most important variable is hydrogen ion concentration (pH), 

 although other factors such as tissue fixation, temperature, and ionic 

 strength of the dye are known to play a role (Singer, 1952). The effect 

 of pH on the staining of relatively pure proteins, such as fibrin, with 

 acid and basic dyes is shown in Figure 11-5. In this figure it can be 

 seen that the pH levels where the staining of fibrin with acid and basic 

 dye is maximum are widely separated (pH 3 and pH 8, respectively). 

 Also there is a range of pH where the fibrin stains with neither acid nor 

 basic dyes regardless of the time of staining. The influence of pH on 

 the staining of cell structures is a reflection of the dissociation charac- 

 teristic of the substances they contain. Proteins and nucleic acids in 

 solution are amphoteric, that is, they can act as acids or bases depending 

 on the pH of the solution. As pointed out in Chapter 2, a protein 

 molecule at its isoelectric point has a net charge of zero due to the 

 balance of positive and negative charges between the dissociated acidic 



216 / CHAPTER 11 



