Inorganic Substances 41 



for about 30 minutes. Wash the shde under the tap and sub- 

 ject it to the Prussian blue method. 



Pohcard^^ finds that the most sensitive method to demon- 

 strate iron is microincineration. 



Copper.— According to Okamoto and co- workers, ^^ certain 

 invertebrate tissues contain some kind of cupric analogue of 

 hemosiderin. The form in which copper occurs in the tissues 

 of higher species is not known, but it is almost invariably 

 masked and not demonstrable directly, except when cupric 

 salts have been administered parenterally. 



Okamoto and Utamura^^ suggest the use of two reagents 

 for the demonstration of copper: rubeanic acid and p-di- 

 methylamino-benzylidenerhodanine. The former was found 

 in model experiments to be highly specific and sensitive, 

 although, contrary to the findings of Okamoto,*^ positive re- 

 sults were never obtained in human tissues. Perhaps an un- 

 masking pretreatment with strong hydrogen peroxide would 

 help to reveal copper even in mammalian organs. 



Method 



Fixation in absolute alcohol or in formalin. 



Incubate sections at 37° C. for 12-24 hours in a mixture 

 consisting of 50 ml. of 10 per cent Na acetate and 1-3 ml. 

 of a 0.1 per cent solution of rubeanic acid ( dithiooxamide ) 

 in alcohol. Copper greenish black. 



Theoretically, copper should be demonstrated also by acid- 

 ified potassium ferrocyanide, in the form of reddish-brown 

 cupriferrocyanide. However, the color would be overlaid 

 beyond recognition by the shade of Prussian blue, since the 

 amount of iron present in the tissues far exceeds that of 

 copper. 



37. Policard, A.: Bull, d'histol. appHq. a la physiol., 11:216, 1934. 



38. Okamoto, K., Utamura, M., and Mikami, G.: Acta Scholae Med. Univ. 

 Kioto, 22:335, 1938-39. 



39. Okamoto, K., and Utamura, M.: Acta Scholae Med. Univ. Kioto, 

 20:573, 1937-38. 



40. Okamoto, K., Utamura, M., and Mikami, G.: Acta Scholae Med. Univ, 

 Kioto, 22:348, 1938-39. 



