320 Microorganisms (chap. 20) 



background — yelloAv to light brown 



melanin and hematogenous pigments — may darken 



underdevelopment will result in pale background, very slender and 



pale spirochetes 

 overdevelopment will result in dense background, heavily impreg- 

 nated spirochetes, obstructed detail, sometimes precipitate 



comments: 



Faulkner and Lillie {194y) use water buffered to pH 3.6-3.8 with 

 Walpole's M/5 sodium acetate-M/5 acetic acid buffer (page 415). 



Levaditi Method for Block Staining (mallory, 1944) 

 fixation: 10% formalin. 



solutions: 

 Silver nitrate: 



silver nitrate 1.5-3.0 gm. 



distilled water 100.0 ml. 



Reducing solution: 



pyrogallic acid 4.0 gm. 



formalin 5.0 ml. 



distilled water 100.0 ml. 



procedure: 



1. Rinse blocks of tissue in tap water. 



2. Transfer to 95% ethyl alcohol: 24 hours. 



3. Place in distilled water until tissue sinks. 



4. Impregnate with silver nitrate, 37°C, in dark: 3-5 days. 



5. Wash in distilled water. 



6. Reduce at room temperature in dark: 24-72 hours. 



7. Wash in distilled water. 



8. Dehydrate, clear in cedarwood oil, and infiltrate with paraffin. 



9. Embed, section at 5 microns, moiuit on slides and dry. 

 10. Remove paraffin with xylene, 2 changes, and mount. 



results: 



spirochetes — black 

 background — brownish yello^v 



Fungi Staining 



Hotchkiss-McManus Method (mc.manus, 1948) 

 fixation: 10% formalin or any geneal fixative. 



