218 Microscopic Histochemistry 



bate sections for 3-18 hours. Remove excess Fe ions bound 



by tissue proteins with an oxalate buffer of pH 4. Convert 



ferric hydroxyquinoline into Prussian blue by treating the 



sections with an acidified solution of potassium ferrocyanide. 



Counterstain with a red nuclear stain. Sites of activity are 



blue. 



Campbell^^^ reports the successful use of these methods. 



Carbonic Anhydrase 



Kurata^^^ has described a histochemical method for the 

 localization of carbonic anhydrase, based on the following 

 principle: 



If a solution of a salt of a heavy metal (such as manga- 

 nese or cobalt) is added to a solution of sodium bicarbon- 

 ate, a precipitate of metal carbonate ( and bicarbonate ) will 

 form. At the same time some carbon dioxide will be liber- 

 ated, owing to the acid reaction of salts of heavy metals. 

 This free carbon dioxide will lower the pH of the solution 

 slightly. On standing, the pH will gradually shift back in 

 the direction of the equilibrium value ( ±8), and additional 

 metal carbonate will precipitate. In the presence of carbonic 

 anhydrase this reaction will be greatly accelerated. 



Kurata fixes thin slices of tissue in cold acetone for 1 hour, 

 washes them briefly in distilled water, and incubates them 

 in a freshly prepared and filtered medium at 37° C. for 45 

 minutes. The medium is a mixture of 100 ml. of an 8 per 

 cent solution of Na bicarbonate and of 10 ml. of a 10 per 

 cent solution of either MnCl2 or C0CI2. The tissues are 

 washed with a bicarbonate buffer of pH ± 7.2 and embed- 

 ded in paraffin. The sections are treated with ammonium 

 sulfide to reveal C0CO3 as black CoS or with neutral peri- 

 odate to convert MnCOs into dark-brown Mn02. Typical 

 localizations (parietal cells of the gastric mucosa; red cells) 

 are claimed. 



132. Campbell, J. G.: Brit. J. Exper. Path., 30:548, 1949. 



133. Kurata, K.: Tr. Soc. Path. Jap., 38:108, 1949. 



