200 Silver Impregnation II (chap. 15) 



6. Wash in several changes of distilled water: 2-3 hours. 



7. Transfer to 50% alcohol, change twice over a period of 24 hours. 



8. Dehydrate, clear, and embed. 



9. Section, mount on slides, deparaffinize, clear, and cover. 



results: 



nerve fibers and endings — brown to black 



comment: 



Gold toning does not improve impregnation. 



Pericellular End-Bulbs (or Boutons) 



Chloral Hydrate-Silver Method (davenport, 1933) B 



fixation: Perfusion (page 22) with 10% chloral hydrate is a desirable 

 procedure, followed by fixation in 10% chloral hydrate: 1-2 hours. 

 (After perfusion cut whole organ into tissue blocks, 5-8 mm. thick.) 



procedure: 



1. Treat tissue blocks with 20-40% pyridine (aqueous): 2 days. 



2. Wash in distilled water: 1-2 hours, no more. Change water every 

 half hour. 



3. Impregnate with 1.5% aqueous silver nitrate (1.5 gm./lOO ml. 

 water): 3-7 days. (Time is not critical within these limits.) 



4. Wash in distilled water: 20-30 minutes, change water once. 



5. Reduce in 2% aqueous pyrogallol (2 gm./lOO ml. water): 5 hours. 



6. Dehydrate, clear, and embed. 



7. Section (15-20 microns) and mount on slides. 



8. Deparaffinize and gold tone if desired, or clear and cover. 



results: 



nerve fibers and endings — black 

 cells — pale yellowish brown 



comments: 



Davenport suggests that the chloral hydrate is perfused into the aorta 

 and can be pumped through the system for about 1-3 minutes before 

 the heart stops. 



The washing after the silver impregnation is critical. If not suffi- 

 ciently washed the periphery will be overstained. One-half hour 

 leaves the superficial as well as the central zones well stained. Five 

 hours of washing produces pale staining and leaves the end bulbs 

 unstained. Cat tisstie gave excellent results. 



