26 LABORATORY METHODS OF UNITED STATES ARMY 



CARBOL-THIONIN. 



Saturated solution of thionin in 50 per 



cent, alcohol lo.oc.c. 



Two per cent, phenol loo.oc.c. 



Stain for two minutes. 



GRAM'S METHOD AND MODIFICATIONS. Preparations are 

 made on cover-slips or slides in the usual way. 



It is always necessary to control Gram stains with organ- 

 isms of known type. 



The preparation is covered with an anil in gentian-violet 

 solution, which is best made up freshly before use. 



The staining fluid is made up, according to Gram's original 

 directions, as follows: 



Five c.c. of anilin oil are shaken up thoroughly with 125 

 c.c. of distilled water. This solution is then filtered through 

 a moist filter paper. 



To 1 08 c.c. of this anilin water add 12 c.c. of a saturated 

 alcoholic solution of gentian violet. The stain acts best 

 when twelve to twenty-four hours old, but may be used at 

 once. It lasts, if well stoppered, for three to five days. A 

 more convenient and simple method of making up the stain 

 is as follows: 



To 10 c.c. of distilled water in a test-tube add anilin oil 

 until on shaking the emulsion is opaque roughly, i to 10. 

 Filter this through a wet paper until the filtrate is clear. 

 To this add saturated alcoholic solution of gentian violet 

 until the mixture is no longer transparent and a metallic 

 film on the surface indicates saturation. One part of alco- 

 holic saturated gentian violet to nine parts of the anilin water 

 will give this result. This mixture may be used immediately 

 and lasts two to five days if kept in a stoppered bottle. 



Cover the preparation with this ; leave on for five minutes. 

 Pour off excess stain and cover with Gram's iodin solution 

 for two or three minutes. 



