EXAMPLE 7 



Staining a Bacterial Film with Crystal Violet 

 by the Technique of Lillie 



This technique is so simple that it would be scarcely worth the trouble 

 to describe it were it not necessary for the benefit of those who have never 

 previously handled bacterial material and who may wish to attempt this 

 for the first time. 



The only tools and reagents necessary are a clean glass slide, a wire loop 

 of the type used normally in bacteriology, a drop bottle containing the 

 crystal violet stain, and a wash bottle containing distilled water. 



The freshly flamed wire loop is touched as lightly as possible either to 

 the surface of the medium in a test tube, or to the surface of the colony 

 in an agar petri-dish culture, and then touched lightly to the center of the 

 clean slide to transfer the bacteria. The only mistakes likely to be made 

 by the beginner are in having too great a quantity of material and hav- 

 ing it on a greasy slide. The result of this combination is seen in Fig. 142. 

 It must be remembered that the specimen is to be examined under an oil- 

 immersion lens, so that the smallest possible smear, derived just by touch- 

 ing the slide with the moist platinum loop, will have an ample area for 

 the purpose required. 



If the microorganisms have been taken from a liquid culture that has 

 not yet reached a very thick stage of growth, the spot on the slide now 

 may be permitted to dry in air; but if the bacteria have been taken 

 from a colony on the surface of a dish, it is necessary to dilute the drop 

 on the slide. For this purpose, a small quantity of water in the same 

 platinum loop is touched to the same spot, rubbed backward or forward 

 once or twice, and then the slide is permitted to dry. 



As soon as the slide has dried, which should only be a moment or two 

 if a sufficiently small quantity of suspension of the specimen has been 

 used, it is taken and "heat fixed" in the flame of a bunsen burner or a 

 spirit lamp. This is done by passing the slide rather rapidly twice through 



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