138 Microscopic Histochemistry 



because at sites of low enzyme concentration activity is likely 

 to drop below the threshold of sensitivity of the method. 



PREPARATION OF TISSUES FOR ENZYMATIC REACTIONS 



The tissue need not be absolutely fresh, but refrigeration 

 is advisable whenever processing cannot be prompt. In most 

 cases satisfactory results are obtained with tissues preserved 

 in the icebox for several (often up to 48) hours, although 

 the pictures may not be quite so sharp as with fresh tissue. 



If fixation is not permissible, fresh-frozen sections must be 

 used. In such cases it is imperative to keep the enzyme un- 

 dissolved; otherwise correct locahzation is impossible unless 

 the enzyme happens to be insoluble. The simplest way to 

 achieve this is to avoid contact with water or with dilute 

 saline solutions and to use strong solutions of ammonium 

 sulfate {¥2 saturated or better) up to the point in the proce- 

 dure at which the localizing precipitate has been obtained. 



It is very likely that a rapid freezing-cutting method, such 

 as that of Adamstone and Taylor^ or the simpler original 

 procedure of Schultz-Brauns,^ followed by a brief fixation in 

 acetone, would give good results with many enzymes. 



The optimal method for most enzymes is probably the 

 freezing-drying technique. Unfortunately, most workers do 

 not possess the equipment and must depend on simpler 

 methods, similar to those used in the average laboratory of 

 histology or pathology. It is to such methods that this chap- 

 ter will be devoted. 



Fixation— As a rule, the best fixative for all enzymes is 

 chilled acetone, which, of all fixatives, causes the least inac- 

 tivation. A bottle of acetone should be kept in the icebox for 

 routine use. Cytological details are not so good as one would 

 like them to be, but they are satisfactory for most purposes 

 if the slices are thin enough (not over 3 mm. in thickness). 

 It is advisable to chill (not freeze) the tissue itself before 



1. Adamstone, F. B., and Taylor, A. B.: Stain TechnoL, 23:109, 1948. 



2. Schultz-Brauns, C: Klin. Wchnschr., 10:113, 1931. 



