194 Methods of Observing Micro-organisms 



cover in distilled water. It is the precipitate that clings to the flagella 

 and renders them distinctly visible. If no precipitate occurs, the 

 flagella will not be seen. 



L. Smith * offers the following modification of Newman's 

 method f as being a simple and excellent method of staining 

 flagella: 



The material and cover-glasses are prepared with care as 

 for the foregoing methods, after which one proceeds as follows : 



1. Transfer a loopful of the bacillary emulsion to the clean slide 



or cover-glass and allow it to dry in the air. 



2. Expose to a mild degree of heat, holding the glass in the fingers 



this is rather drying than actual heating. 



3. Allow the stain to drop from a filter upon the film and remain 



in contact five to ten minutes. 

 The formula for the stain is 



I. Tannic acid i gram 



Potassium alum i 



Distilled water 40 c.c. 



Dissolve by shaking or allow to stand overnight in the in- 

 cubator. 



II. Dissolve "night blue" J 0.5 gram 



95 per cent, or absolute alcohol 20.0 c.c. 



Mix I and II thoroughly and remove the heavy precipitate 

 by filtration. If not used at once, drop from a filter upon 

 the film. The stain does not keep more than a few days. 



4. Wash carefully but thoroughly in water. 



5. Apply a saturated aqueous solution of gentian violet for about 



two minutes to stain the bodies of the bacteria. 



6. Wash thoroughly in water, dry with smooth blotting-paper, 



and mount in balsam. 



To secure a perfectly clean background for photomicrography, it is 

 best to stain on a slide. The stain is then poured into a Petri dish, the 

 slide inverted, the end of the slide used to push aside the film on the 

 surface of the stain, and the film then immersed downward, one end of 

 the slide supported, during staining on a match-stick or bit of glass rod. 

 In this way the adherence of the precipitate to the slide can be avoided. 



THE OBSERVATION OF LIVING PROTOZOA. 



When protozoa are to be examined in transparent fluids, 

 such as pond-water or culture fluids in which they have been 

 artificially nourished, use can be made of a " live-box " or 

 of the " hanging drop." Ordinarily, however, the organisms 

 to be examined are contained in blood, in pus, in sputum, 

 in feces, or in some other more or less opaque fluid, of which 

 an extremely thin layer must be prepared in order that the 



* "Jour. Med. Research," vi, 1901, p. 341. 



t" Bacteria," John Murray, London, 2d edition. 



t James Strong & Son, Glasgow and Manchester. 



