200 MICROSCOPIC AND CULTURAL STUDY OF BACTERIA 
(6) Wash with water. 
(c) Flood with 5 per cent chromic acid sohition for two minutes. 
(f/) Wash thoroughly in running water. 
(e) Flood with carbol-fuchsin and steam for five minutes. 
(/) Wash thoroughly in water. 
(f/) Decolorize with 1 per cent sulphuric acid until the excess of stain 
is removed. 
(h) Wash thoroughly in water. 
(i) Flood with Loffler's alkaline methylene blue and allow to stain 
one minute. 
(j) W^ash in water, dry and mount. 
Spores stain red, vegetative cells blue. 
Muir Spore Stain. — (a) Prepare a very thin film, which is dried in 
the air and then steamed gently after flooding with filtered carbol- 
fuchsin for about thirty seconds. 
(b) Wash rapidly with 90 per cent alcohol. Then wash well with 
water. 
(c) Flood the preparation with the following mordant for thirty to 
sixty seconds: 
Saturated solution of mercuric chloride 2 cc. 
20 per cent tannic acid solution 2 cc. 
Saturated solution of potassium alum 5 cc. 
(f/) Wash thoroughly with water. 
(e) Wash with 90 per cent alcohol for one minute, or until the color 
is pinkish to pale red. 
(/) Wash thoroughly with water. 
ig) Counterstain with methylene blue for one minute. 
{h) Dry with filter paper. 
Bacterial bodies are colored deep red, the spores light blue. 
2. Capsule Stains.— WelchSuttaW Method.^— (a) Fixed films of 
bacteria are flooded with glacial acetic acid for a few seconds. 
(6) The acid is poured oft* and the preparation is washed two or 
three times with anilin oil gentian violet, then flooded with the stain, 
which is allowed to act for three to five minutes. 
(c) Wash with 2 or 3 per cent aqueous solution of sodium chloride. 
(d) Mount in salt solution and examine. 
Capsules stain faint purple, bacterial body deep purple. 
Hiss Method.^— (a) Prepare the smear on perfectly clean cover-glass, 
do not use any water; dry in the air, and do not fix by heat. 
(6) Cover smear with aqueous gentian violet, and heat gently until 
steam is just visible. 
(c) Wash off immediately with 20 per cent copper sulphate solution. 
{d) Blot and dry. 
1 Johns Hopkins Hosp. Bull., 1892, 3, 81. 
2 Jour. Med. Res., 1901-1905, 6, 317. 
