34 Microscopic Histochemistry 



ultraviolet light source) until the precipitate appears black 

 or by using photographic developers. 



Reduction of the silver salt by light has certain disadvan- 

 tages: (1) a brownish halo may sometimes form around the 

 black granules, or (2) the black precipitate may turn brown 

 when the unreacted silver is removed. For these reasons, 

 some workers prefer to use photographic developers. The 

 sHdes are first thoroughly washed in many changes of dis- 

 tilled water and then immersed in a dilute solution of hydro- 

 quinone or pyrogallol (about 0.5 per cent; concentration not 

 important) for about 2 minutes and rinsed again. After both 

 methods of reduction, the unreacted silver must be removed 

 by a short bath in a thiosulfate (hypo) solution (about 2 per 

 cent); otherwise the shdes may darken later. After photo- 

 graphic development the slide presents a very sharp contrast 

 between black and white, no intermediate shades being pres- 

 ent; however, failure to wash the slide very thoroughly be- 

 fore the use of the developer will result in the deposition of a 

 very fine, dustlike black precipitate all over the tissue. Slides 

 prepared in either way can be counterstained as desired, de- 

 hydrated, and mounted. 



The only source of error with the silver method is the pres- 

 ence of massive deposits of uric acid and of its salts, which 

 may stain very much like phosphate-carbonate. For differ- 

 entiation between urate and phosphate-carbonate see the 

 section on uric acid. 



The other heavy-metal techniques are performed in an 

 analogous way. First, the section is immersed for %-l hour 

 in the solution of a salt of the heavy metal (e.g., cobalt 

 nitrate, ferrous sulfate, etc.), then washed thoroughly and 

 treated with the reagent (ammonium sulfide, acidified po- 

 tassium f erricyanide, etc. ) . The two main disadvantages of 

 these techniques are that (1) the proteins may retain the 

 heavy metal rather stubbornly and in this way give rise to a 

 more or less intensely colored background and (2) the pene- 

 tration of heavy-metal salts into dense granules or spicules 



