32 STAINING METHODS. 



Mordant. Sat. aqueous sol. mercuric chloride, 2 parts 

 Tannic acid (20% aqueous sol.), 2 parts 

 Sat. aqueous sol. potash alum, 5 parts 



4. Wash in water thoroughly. 



5. Treat with 95% alcohol for one minute. (The preparation 

 should have a pale red color.) 



6. Wash well in water. 



7. Counterstain with methylene blue one-half minute. 



8. Dehydrate in alcohol. Clear in xylol and mount. (May 

 simply dry specimen with filter-paper.) 



Flagella Staining. Inoculate a tube of sterile water (gently) in 

 upper part, with just enough of an i8-to 24-hour-old agar culture, to 

 produce faint turbidity. Incubate for two hours at 37 C. From the 

 upper part of culture take a loopful and deposit it on a cover-glass. 

 Dry in thermostat for one to five hours or over night. Use perfectly 

 clean cover-glasses. To stain by. 



Muir's Modified Pitfield Method. i. Flood specimen with 

 mordant. Steam gently one minute. 



Mordant. Tannic acid (10% aqueous solution), 10 c.c. 



Sat. aq. sol. mercuric chloride, 5 c.c. 



Sat. aq. sol. alum, 5 c.c. 



Carbol fuchsin 5 c.c. 



Allow precipitate to settle or centrifuge. Keeps only one week. 



2. Wash well in water for two minutes. 



3. Dry carefully preferably in incubator. 



4. Pour on stain. Steam gently one minute. 

 Stain. Sat. aq. sol. alum, 10 c.c. 



Sat. ale. sol. gentian violet, 2 c.c. 



(May use carbol fuchsin instead of gentian violet.) 

 Stain only keeps two days. 



5. Wash well in water. Dry and mount. 



Spore Staining. The most satisfactory spore staining method is 

 really the negative staining of the spore obtained "when a bacterial 

 preparation is stained by dilute carbol fuchsin or LofHer's methylene 



