Enzymes 161 



hours. Temperature should be between 20° and 37° C. It is 

 advisable to change the incubating solution once or twice to 

 avoid the deposition of a melanin precipitate (by spontane- 

 ous oxidation of the substrate). At pH 7.7 the reaction is 

 much faster (about 1 hour), but the danger of precipitates 

 is also increased. Rinse sections, counterstain as desired, de- 

 hydrate, and mount. Sites of dopa oxidase activity appear 

 dark brown-gray or brown-black. For greater contrast, mel- 

 anin formed during the reaction can be blackened by silver 

 (see under "Pigments"). 



D) Amine oxidase.— 



This enzyme, originally described by Hare,^^ oxidizes ali- 

 phatic and aromatic amines, such as isoamylamine, tyramine, 

 histamine, tryptamine, etc., first to the corresponding alde- 

 hydes and then to acids. Ammonia and hydrogen peroxide 

 are formed in the course of the reaction. The corresponding 

 amino acids ( tyrosin, histidine, etc. ) are not attacked. 



Oster and Schlossmann^^ described a method for the histo- 

 chemical localization of amine oxidase, based on the demon- 

 stration of the aldehyde formed. The detailed technique is 

 as follows: 



Fresh-frozen sections are used. Since tissues contain plas- 

 malogen, which, on standing, may break down to plasmal, 

 an aldehydic compound, it is necessary first to transform 

 plasmal into a nonreacting form to avoid its being mistaken 

 for newly formed aldehyde. This is accomplished by treating 

 the sections with a solution of bisulfite; the bisulfite addition 

 compound of plasmal does not react with the usual aldehyde 

 reagents. After thorough washing, the sections are incubated 

 with the substrate (tyramine) and treated with Schiff's rea- 

 gent. The aldehyde formed is demonstrated by the localized 

 development of an indigo-blue shade. 



The theoretical foundation of this method is not sound. 



63. Hare, M. L. C: Biochem. J., 22:968, 1928. 



64. Oster, K. A., and Schlossman, N. C: J. Cell. & Comp. Physiol., 

 20:373, 1942. 



