180 Microscopic Histochemistry 



mated at 40-45,^^ 65,^^ and 75 per cent,^^ respectively. On 

 the other hand, CappeHn^^ and Cleland^^ find that embed- 

 ding causes very httle inactivation. In Coujard experiments, 

 embedding did not inactivate the shdes noticeably except 

 after formalin fixation ( ±50 per cent). No data are available 

 on the effects of celloidin-embedding. 



ParaflBn blocks, in the writer's experience, retain their ac- 

 tivity unchanged for over 10 years. Lison^* finds that they 

 may deteriorate in 2 months. 



Cut ribbons can be kept in the icebox for many months; 

 melted-on sections can be stored at room temperature for 

 years without the slightest loss in activity. Danielli, Doyle,^^ 

 and Lison, on the other hand, observed definite inactivation 

 of the enzyme in stored paraffin sections. 



1 ) The calcium phosphate method,— The first histochemi- 

 cal procedure was published by Takamatsu^^ in 1938, in a 

 journal not readily accessible to Western readers. In 19-39 the 

 procedure was republished by Takamatsu^^ and simultane- 

 ously described independently by Gomori.^^ It was based on 

 the principle that, if sections are incubated with glycero- 

 phosphate at an alkaline reaction in the presence of Ca ions, 

 the phosphate ions liberated will be precipitated in statu nas- 

 cendi ( at the site of formation ) as insoluble Ca phosphate. 

 The latter is then transformed, in a second step, into metallic 

 silver or black cobalt sulfide. 



The composition of the incubating mixture may be varied 

 within fairly wide limits without much difference in the final 

 results. The pH of the solution should be between 9 and 9.8 

 (in the lower ranges in the case of aliphatic substrates, in 

 the higher ranges in the case of aromatic ones). Below pH 



31. Doyle, W. L.: Proc. Soc. Exper. Biol. & Med., 69:43, 1948. 



32. Stafford, R. O., and Atkinson, W. B.: Science, 107:279, 1948. 



33. Cleland, K. W.: Proc. Linnean Soc. New South Wales, 75:35, 1950. 



34. Lison, L.: Bull, d'histol. appliq. a la physiol., 25:23, 1948. 



35. Takamatsu, H.: Manshu Igaku Zasshi, 31:34, 1938. 



36. Takamatsu, H.: Tr. Soc. Path. Jap., 29:492, 1939. 



37. Gomori, G.: Proc. Soc. Exper. Biol. & Med., 42:23, 1939. 



