38 General Bacteriology. 



c. Stain with anilin-oil gentian violet 5 minutes. 



d. Pour off stain and without washing: 



e. Apply iodine solution 2 minutes (use several changes). 



/. Decolorize with 96% alcohol until drippings do not stain white filter paper. 

 g. Wash in water and counter-stain with Bismarck brown. 

 h. Mount in water and examine. 

 i. Dry and mount in balsam. 



REFERENCES. A. 162; H. 78; L. & K. 106; M. & W. 91; M. & R. 110; McF. 

 99; N. 287; P. 203. 



SPECIAL DIRECTIONS. Stain films of young cultures of B. coli and B. subtilis. Also 

 a film of an organism supplied. 



EXERCISE XXXII. TUBERCLE STAIN (Qabbett). 



EXPLANATORY. All of the differential methods of staining the tubercle bacterium 

 depend upon the fact that this germ is very resistant towards the ordinary stains and in 

 order to be stained at all must be treated with a dye containing a mordant and this 

 either allowed to remain in contact with the micro-organism several hours or be applied 

 hot. The latter method is the quicker and is usually employed, although it does not 

 give as good results. When once stained this germ withstands the effect of decoloriz- 

 ing agents to such an extent that it is possible to remove the dye from all other objects 

 on the cover-glass preparation (as in sputum) while it retains its own color. The appli- 

 cation of a second dye, of a complementary color, readily distinguishes this germ from 

 all others in the field. A few other bacteria have similar staining properties. (See 

 Part II.) Red is the usual stain and blue the counter stain. Gabbett's method is one of 

 the simplest. 



GENERAL DIRECTIONS. 



a. Spread film (sputum from tuberculous patient) . 



b. Air dry and fix. 



c. Stain with hot carbol-fuchsin 2 minutes. 



d. Wash in water. 



e. Treat with Gabbett's solution | to 1 minute. 

 /. Wash in water and examine. 



g. Dry and mount in balsam. 



REFERENCES. A. 162; M. & W. 92; McF. 214; P. 304. 



SPECIAL DIRECTIONS. Stain three samples of sputa which contain varying numbers 

 of the tubercle bacteria. 



EXERCISE XXXIII. STAINING ENDOSPORES. 



GENERAL DIRECTIONS. 

 A. Simple stain. 



a. Prepare film as usual. 



6. Fix by passing through flame 10 or 12 times instead of 3 times. (This prevents 

 the vegetative portion from taking the stain) . 



c. Stain 2-5 minutes in hot carbol-fuchsin. 



d. Mount and examine. 



