204 Silver Impregnation II (chap. 15) 



5. Rinse in distilled water, 3 changes: 1 minute total. 



6. Tone in gold chloride: 4 minutes. 



7. Rinse in distilled water, 3 changes: 1 minute total. 



8. Develop in 2% oxalic acid (2 gm./lOO ml. water). Check under 

 microscope until background is grey and fibers are sharply de- 

 fined: approxiinately 3 minutes. 



9. Wash in distilled water, 3 changes: 1 ininute each. 



10. Fix in 5% sodium thiosulfate (5 gm./lOO ml. water): 5 minutes. 



11. Wash in running water: 5-10 minutes. 



12. Rinse in distilled water. 



13. Counterstain if desired. 



14. Treat with acidified water: 5 minutes. 



15. Dehydrate, clear, and mount. 



results: 



nerve fibers — black 



background colors — depending on counterstain 



comments: 



Foley (1943) considers counterstaining essential, claiming that it 

 serves as contrast for nervous and non-nervous tissue and adds to 

 transparency of the sections. Follow step 11 with: 



12. Stain in gallocyanin (page 138): overnight. 



13. Wash thoroughly running water: 5-10 minutes. 



14. Mordant in 5% phosphotungstic acid (5 gm./lOO ml. water): 30 

 minutes. 



15. Transfer directly to dilution (20 ml./30 ml. water) of following 

 stock solution: 1 hour. 



aniline blue WS, C.I. 42780 0.1 gm. 



fast green FCF, C.I. 42053 0.5 gm. 



orange G, C.I. 16230 2.0 gm. 



distilled water 92.0 ml. 



glacial acetic acid 8.0 ml. 



16. Differentiate through 70% and 95% alcohols. 



17. Dehydrate, clear, and mount. 



Ungewitter's Modification [1943) 



procedure: 



1. Deparaffinize and hydrate sections to water. 



2. Treat with Protargol sokuion (plus copper), 37 °C: 12-24 hours. 



