18 General Bacteriology. 



SPECIAL DIRECTIONS. 



a. Incubate all cultures of the non- pathogenic bacteria at 28 C., except the gelatin. 

 Keep these in the cool chamber. After growth has taken place, the cultures can be taken 

 from the incubator and kept at the room temperature. 



b. Study and make diagrams of an incubator, a Reichert thermo-regulator, a Roux 

 thermo-regulator and Koch's safety burner. 



EXERCISE XV. CLEANING SLIDES AND COVER-GLASSES. 



GENERAL DIRECTIONS. Slides can be sufficiently cleaned by washing in water or 

 alcohol and drying with a towel. The cover-glasses for bacteriological work, however, 

 must not only be freed from visible dirt but must be rendered free from fat. One of the 

 best methods is the following: New cover-glasses are cleaned by washing in water and 

 drying from alcohol between driers (two blocks 20xlOx2J- mm. covered with several layers 

 of cotton cloth or chamois skin) , and then heating them on a piece of sheet iron or in hot 

 air sterilizer for one hour at about 200 C. They are best kept in a clean Petri dish and 

 handled with forceps. (Novy). Old slides and covers having balsam on them should first 

 be dropped one by one into a cleaning solution (potassium bichromate 60, sulphuric acid 

 60, water 1000) , and boiled for one-half hour and then treated as above. 



SPECIAL DIRECTIONS. Clean oz. of cover-glasses and place them in a clean Petri 

 dish. 



EXERCISE XVI. PREPARATION OF STAINING SOLUTIONS. 



GENERAL DIRECTIONS. The dyes most useful for staining bacteria are the basic 

 anilin dyes which come in powdered or crystalline form. (Gruebler's dyes are standard.) 

 Those in most common use are Fuchsin, Methylen blue, Gentian violet and Bismark 

 brown. They keep well in powdered form, with perhaps the exception of Methylen 

 blue, but because of greater convenience and equally good keeping qualities, saturated 

 alcoholic solutions are kept in stock. These are made by adding the dry dye to 95% 

 alcohol to saturation and filtering. This form can not be used for staining bacteria. The 

 following solutions are required to begin work with: 



1. Aqueous solution of Gentian violet. 



Saturated alcoholic solution of Gentian violet, - 2.5 cc. 



Distilled water, 47.5 cc. 



2. Saturated aqueous solution of Bismark brown. 



3. Ziehl's carbol-fuchsin. 



Saturated alcoholic solution of Puchsin. 5 cc. 



Solution of carbolic acid (5%)- - 45 cc. 



4. Loeffler's Methylen blue. 



Saturated alcoholic solution of Methylen blue, - 15 cc. 



Potassium hydrate ( 1 : 10 , 000 ) , 50 cc . 



5. Ehrlich's Anilin Oil Gentian violet. 



Saturated alcoholic solution of Gentian violet, 6 cc. 



Absolute alcohol, - 5 cc. 



Anilin water, - - 50 cc. 



Anilin water is prepared by adding 2-3 cc. of anilin oil drop by drop to 50 cc. of 

 water, thoroughly shaking and then filtering through moistened filter paper until per- 

 fectly clear. 



