134 MICROBIOLOGY 



glass. Gently touch the surface growth on the agar culture 

 with the end of the platinum needle and immerse it in the 

 water on the cover-glass without spreading the drop. The 

 impregnated needle should carry bacteria enough for 3 or 4 

 preparations. Then place the tray of cover-glasses in the 

 incubator to dry. The bacteria become disseminated through- 

 out the water by means of their power of locomotion. When 

 dry they are ready for the staining treatment. 



The bacteria are fixed to the cover-glass by holding them, 

 film upward, between the thumb and forefinger, over a gas 

 flame for about a minute. They are then treated with 

 Loeffler's mordant. 



Place the fixed cover-glass preparation in a large test 

 tube, cover it with mordant, and carefully heat over a gas 

 flame or in a water bath until steam is given off. Allow the 

 mordant to act for from 3 to 5 minutes. Then remove the 

 cover- glass with a bent wire loop and fine forceps and thor- 

 oughly rinse it in water. Then place it in a similar tube and 

 cover with carbol fuchsin for staining. Heat this as the mor- 

 dant was heated and allow the stain to act for from 5 to 10 

 minutes. Remove the cover-glass as before and thoroughly 

 rinse in water. If the stain is too deep, decolorize by rinsing 

 the preparation for a few seconds in alcohol and again in 

 water. It is then ready for the microscopic examination in 

 water, or it may be allowed to dry and then be mounted in 

 balsam. If the first preparation fails, add 2 drops of a 10% 

 solution of sulphuric acid to the mordant. 



The flagella should appear as fine, hairlike appendages 

 radiating from the bacteria. 



Van Ermengem's 2 method. The films are prepared as 

 described above. Three solutions are necessary. 



SOLUTION A (FIXING BATH) 



Osmic acid, 2% solution 1 part 



Tannin, 10 to 25% solution 2 parts 



2 Van Ermengem. Ref. Centralb. f. Bakteriologie, Bd. XV (1894) 

 No.' 24, p. 969. 



