Enzymes 163 



with peroxide and a suitable substrate is not, in itself, a 

 cogent proof for the presence of true peroxidase. Even when 

 it is known that a morphological structure does contain per- 

 oxidase (e.g., leucocytes), it is always questionable whether 

 the reaction obtained under the conditions of the histochemi- 

 cal experiment is due to enzymatic action or to a nonprotein 

 catalyst ( "pseudo-peroxidase" ) . 



For the histochemical demonstration of peroxidase, benzi- 

 dine,"^^ naphthol,^^ and various leuco-dyes are utilized^-' ^^ in 

 the presence of hydrogen peroxide. Benzidine is oxidized to 

 a blue ( quinhy drone ) or brown ( quinoneimine ) dye; naph- 

 thol to a purple-black one, the chemical nature of which is 

 not clear; leuco-dyes are recolorized to their original shades. 

 It is important to run controls without peroxide because 

 positive reactions may be obtained even in its absence (see 

 previous section, "Phenol Oxidase" ) . 



The two main histological localizations of peroxidase are 

 hemoglobin and myeloid granules. However, the reactions 

 given by hemoglobin differ from those obtained in myeloid 

 granules in several important points: 



1. Zinc leuco-dyes are readily recolorized by hemoglobin 

 but not by myeloid granules. 



2. In the benzidine reaction the optimal concentration of 

 H2O2 is about 0.01 M in the case of myeloid granules and 

 much higher, about 0.1 M, in the case of hemoglobin. 



3. The activity of myeloid granules is rapidly destroyed 

 by heating to 75°-80° C. or by extraction with a warm chloro- 

 form-methyl alcohol mixture. The activity of hemoglobin 

 is entirely resistant to these influences. 



In model experiments, stale (peroxidized) linseed oil im- 

 bibed in filterpaper strips gives rather intense reactions with 

 benzidine but none with leuco-dyes or with naphthol. An- 



70. Adler, O., and Adler, R.: Ztsclir. f. physiol. Chem., 41:59, 1904; 

 Lepehne, G.: Beitr. z. path. Anat. u. z. allg. Path., 65:163, 1919; Good- 

 pasture, E. W.: J. Lab. & Clin. Med., 4:442, 1919. 



71. Loele, W.: Foha haemat., 14:26, 1912. 



72. Lison, L.: Compt. rend. Soc. de bioL, 106:1266, 1931. 



73. Jacoby, F.: J. Physiol., 103:25P, 1944-45. 



