Degenerating Axons 209 



formalin, 10% 13.5 ml. 



citric acid, 1% aqueous 13.5 ml. 



procedure: 



1. Soak sections in 15% alcohol: 30 minutes. 



2. Wash briefly in distilled water and soak in 0.5% phosphomolyb- 

 dic acid (0.5 gm./ 100 ml. water): 15-20 minutes. 



3. No wash; transfer to 0.5% potassium permanganate (0.5 gm./lOO 

 ml. water): 4-10 minutes. Can be critical; make trial runs at 5, 7, 

 and 9 minutes. Turn sections over occasionally. 



4. Rinse in distilled water. 



5. Decolorize in equal parts of 1% hydroquinone (1 gm./lOO ml. 

 Avater) and 1% oxalic acid (1 gm./lOO ml. water): 1-2 minutes. 



6. Wash thoroughly in distilled water, 3 changes. 



7. Treat with 1% silver nitrate (1 gm./lOO ml. water): 20-30 min- 

 utes. 



8. Transfer sections individually; a brief Avash in distilled Avater 

 and then into freshly prepared ammoniacal silver: 1 minute. 



9. Transfer to reducing solution, allowing sections to float on sur- 

 face of solution. Brown color Avithin 1 minute. If too light, add 

 a little sodium hydroxide to ammoniacal silver solution; if too 

 dark add some ammonia. 



10. Wash briefly, fix in 1% sodium thiosulfate (1 gm./lOO ml. water; 

 2 miniues. 



11. Wash in distilled water, 3 changes. 



12. Mount, Albrecht's {1934) method: 



(ft) Transfer into equal parts of 1.5% gelatine (1.5 gm./lOO ml. 

 Avater) and 80% alcohol: 5 minutes or longer. 



[b) Tease section onto slide, AvithdraAV from fluid Avith slide tilted 

 alloAving rapid drainage of excess fluid. W^ipe aroiuid section, 

 but do not alloAv it to completely dry. 



(c) Blot gently Avith smooth filter paper. If section sticks to paper, 

 first moisten paper Avith 95% alcohol. 



(cl) Immerse rapidly in 95% alcohol to congeal gelatine. 



13. Dehydrate, clear, and mount. 



results: 



disintegrating axons — black 



normal axons — various shades of broAvn 



comments: 



See also Nauta and Gygax (1951); Nauta and Ryan (1952); Wall 

 (1950); White (1960); and additional references included therein. 



