48 MICROSCOPIC TECHNIQUES 



nitrate method which Pap (1929) employed for the staining of 

 reticulum visualized glycogen more intensely and consistently than 

 either the Best carmine or Bauer-Feulgen procedures. The admitted 

 drawback to this method is the fact that since reticulum fibers of 

 connective tissue are also stained it cannot be applied to this tissue. 

 However, the authors feel that in other cases the ammoniacal silver 

 nitrate method has advantages over those previously employed. 

 Gomori (1946) subsequently modified this procedure and developed 

 a more selective method which demonstrates glycogen and mucin, 

 but eliminates possible interference by desoxyribonucleic acid, uric 

 acid, and granules of enterochromaffin cells, all of which can reduce 

 silver solutions under certain conditions. However, melanin is stained, 

 and except for this the method enables the same localizations of the 

 reducing substances as the Bauer-Feulgen procedure. Should in- 

 soluble calcium salts be present they too will stain black. A prelim- 

 inary 10 min. treatment of the sections with citrate buffer of pH 

 3-4 will remove calcium deposits. 



The differentiation of glycogen from other substances that give 

 positive reactions may be made in some instances by employing 

 saliva to digest away the glycogen selectively. See page 46 foi* the 

 staining of mucoproteins. 



Bauer-Feulgen Stain for Glycogen (after Bensley) 



SPECIAL REAGENTS 



Alcohol-Formalin Fixative. 9 vol. absolute alcohol plus 1 vol. 

 neutral formalin. The alcohol may be first saturated with picric 

 acid. 



Feulgen Reagent* Heat to dissolve 1 g. basic fuchsin in 100 ml. 

 distilled water. Filter while warm, cool, add 20 ml. 1 A^" hydro- 

 chloric acid and 1 g. sodium bisulfite. Let stand 24 hr. The soln. 

 should be straw colored. 



1% or 4% Chromic Acid. 



Bisulfite Rinsing Solution. 1 vol., 1 M sodium bisulfite plus 19 

 vol. tap water. 



PROCEDURE 



1. Fix very small pieces (2-3 mm.) of fresh tissue in the alcohol- 



*See pages 65 and 67 for other methods of preparing this reagent. 



