142 STAINED PREPARATIONS 



cover-glass near the edge with the point of a needle, and the side on which the film 

 has been spread will be easily distinguished by the little scratches which will remain. 

 But the possibility of losing the film side can be avoided by marking the upper 

 limb of the forceps with a glass pencil, or by the use of a pair of Cornet's forceps 

 on one limb of which a small knob-like depression is impressed in the metal ; if 

 the forceps be always used with this knob upwards it will act as a guide. 



(6) A small quantity only of culture should be used for making the film. The 

 shape of the organisms can best be made out when there are only a few in the field 

 of the microscope. 



(c) The Canada balsam should be dissolved with xylol and the solution should be 

 of such a consistency that it does not tail when a drop is taken out with the glass 

 rod. Balsam should be kept in a bottle stoppered with a glass bell- stopper and 

 having a rim arranged so that the excess of balsam taken up on the glass rod can 

 be drained off. 



(d) Alcohol-ether, alcohol and volatile reagents generally are best kept in 

 drop-bottles stoppered with ground-glass stoppers (several patterns can be obtained 

 at the shops) : shallow thick glass bottles of 60 to 100 c.c. capacity are perhaps 

 the most convenient. 



B. The method which has just been described is especially useful for delicate 

 work and for making films which are to be preserved. But for the provi- 

 sional examination of cultures and for routine work it is quicker and more 

 economical to work with slides. 



1. Take a slide between the fingers or hold it in a pair of Debrand's forceps, 

 and place a little drop of the culture on it. 



2. Spread, dry and fix as in the former case (A). 



3. Stain the film and wash it in the manner described, and then dry the 

 slide. Put a drop of cedar- wood oil straight on to tlie film without using 

 a cover-glass and examine with the oil-immersion lens. 



If after examination it is desired to preserve the preparation the cedar- wood oil 

 can be washed off with a few drops of xylol, and when this has evaporated the 

 slide is put away dry. Or after washing off the cedar oil the film may be mounted 

 with a drop of balsam and cover-glass. 



[C. Another method which gives excellent results and is commonly adopted 

 by us seems to deserve description here. 



[1. The film is spread, dried and fixed on a slide as in B. 



[2. Wash the film for a moment or two in a 10 per cent, aqueous solution 

 of acetic acid. Wash thoroughly in water. Blot and dry. 



[3. Place a drop of the stain on the centre of the film and lower a cover- 

 glass on to the stain, avoiding the introduction of air bubbles. Blot the 

 upper surface of the cover-glass with blotting paper. 



[4. Put a drop of oil on the dried surface of the cover-glass and examine 

 with a y^th oil immersion. 



[If the preparation is to be preserved, float off the cover-glass from the 

 slide by putting a drop or two of water at the edge of the cover-glass. Wash 

 the slide in water, blot and dry it. The slide may then be kept indefinitely. 



[Both in this method and in the preceding, the film may be decolourized 

 in a 10 per cent, aqueous solution of acetic acid or in alcohol, and then restained 

 with another dye ; so that using the same film one may first determine the 

 morphology of the organism by examination in a simple stain and then 

 ascertain its reaction to Gram's stain (vide infra).] 



3. Grain's method of staining. 



Gram devised a method of staining which serves to divide bacteria into two 

 large groups. 



Some bacteria, when stained with a basic pararosanilin dye in aniline or 



