METHODS OF STAINING OF BACTERIA 49 



This is an excellent bacterial stain. 



Erythrosin^ 



(a) Erythrosin 5.0 gm. 



Alcohol (70 per cent) 100 . cc. 



(6) Erythrosin 1.0 gm. 



Carbolic acid (5 per cent) 100 . cc. 



This stain is especially recommended for root-nodule bacteria. 



1. Place a drop of the fresh culture on a glass slide, tilt the 

 slide to allow drop to spread. Dry the film in an oven at 45°C. 

 and fix in absolute alcohol. 



2. After the alcohol evaporates, flood the mount with (a) and 

 allow to stain for 10 minutes. 



3. If the stain is not deep enough, wash off this alcohol ery- 

 throsin and again stain with (6) for 10 minutes. 



1 WiNOGRADSKY, S. Ann. Inst. Pasteur, 39: 325, 1925; Gangulee, N. 

 Ann. Appl. Biol, 13: 364, 1926. 



Capsule Stain Adapted from Dorner Spore Stain 



1. Spread a loopful of culture on clean glass slide. Air dry 

 and do not fix. 



2. Stain 1 to 2 minutes with cold carbol fuchsin. Wash 

 gently in water. 



3. Cover with thin layer of nigrosin by spreading a loopful 

 of saturated aqueous solution of nigrosin. Dry quickly to 

 prevent decolorization of the cells. 



Cells will be red and capsules white against a blue-gray 

 background. 



Capsule Stain. Negative Method with Ink 



Place a loopful of liquid culture on a clean glass slide. Add 

 to it a loopful of ink (small loop carrying about one-fourth as 

 much liquid). Cover with a cover slip and examine as a wet 

 mount. The bacteria will be white against a gray background; 

 their capsules will be unstained and appear as halos around the 

 denser bacterial cells. 



By this method one can demonstrate some capsules which 

 cannot be stained by any positive staining method. 



The ink to be used is Burri's Pelikan Tusche, No. 541 (Griibler). 

 It is bacteria free when received and must be kept aseptically. 



