24 " Fixation (chap. 1) 



ones; 1-2 hours for sections on slides before staining (may be left 



overnight). 

 Wash thoroughly in running water, overnight for large gross tissues; 



15-30 minutes for slides. 

 Improves preservation and staining, particularly of mitochondria. 



De formalization 



The removal of bound formalin frequently is necessary in silver im- 

 pregnation methods, such as Ramon y Cajal, del Rio-Hortega methods, 

 and the Feulgen technique. 



Lhotka and Ferreira's (1950) Method 



1. Wash tissue blocks in distilled water: 15 minutes. 



2. Transfer through 2 changes of chloral hydrate, 20% aqueous (20 

 gm./lOO ml. water): 24 hours each. 



3. Wash in distilled water: 15 minutes. 



4. Proceed to any method. With silver stains, time of impregnation 

 needs to be lengthened: Ramon y Cajal to 2 weeks; del Rio-Hor- 

 tega to 24 hours. 



Krajian and Gradwohl's (1952) Method, on Slides 



1. Place in ammonia water (40 drops ammonia in 100.0 ml. water): 

 1 hour. 



2. Wash, running water: 1 hour. 



3. Fix in special fixative if necessary: 1 hour. 



4. Wash, running water: 1 hour. 



5. Proceed to stain. 



Removal of Mercuric Chloride Crystals 



Iodine method: see page 12. 



Gonzalez method (1959A). Gonzalez suggests the following if necessary 

 to avoid mordant action of iodine. After fixation, wash for a short 

 time, or immerse directly in cellosole (ethylene-glycol-monoethyl- 

 ether. Carbide and Carbon Chemical Co.): 24-48 hours, 3 changes. 

 Follow with toluene: 1-2 hours; infiltrate and embed. 



Decalcification 



Calcium deposits may be so heavily concentrated that they interfere 

 with future sectioning and result in torn sections and nicks on the knife 



