96 MICROSCOPIC METHODS 



on such, a small drop of blood is placed near one end, the edge 

 of a second clean slide is lowered through the drop on to the 

 surface of the glass on which the blood has been placed. This 

 second slide is held at an angle to the first, and the droplet of 

 blood by capillarity spreads itself in the angle between the two 

 slides. The edge of the second slide is then stroked along the 

 surface of the first slide, and the blood is spread out in a film 

 whose thickness can be regulated by the angle formed by the 

 second slide. Large-sized films can thus be obtained, and when 

 these are stained they are often examined without any cover- 

 glass being placed upon them. A drop of cedar oil is placed on 

 the preparation, and after use this can be removed by the careful 

 application of xylol. 



Films dried and fixed by the above methods are now ready to 

 be stained by the methods to be described below. 



(b) Wet Method. If it is desired to examine the fine 

 histological structure of the cells of a discharge as well as to 

 investigate the bacteria present, it is advisable to substitute 

 " wet " films for the " dried " films, the preparation of which has 

 been described. The nuclear structure, mitotic figures, etc., are 

 by this method well preserved, whereas these are considerably 

 distorted in dried films. The initial stages in the preparation 

 of wet films are the same as above, but instead of being dried 

 in air they are placed, while still wet, film downwards in 

 the fixative. The following are some of the best fixing 

 methods : 



(a) A saturated solution of perchloride of mercury in '75 per cent, 

 sodium chloride ; fix for five minutes. Then place the films for half an 

 hour, with occasional gentle shaking, in '75 per cent, sodium chloride 

 solution to wash out the corrosive sublimate ; they are thereafter washed 

 in successive strengths of methylated spirit. After this treatment the 

 films are stained and treated as if they were sections. 



(b) Formol-alcohol formalin 1 part, absolute alcohol 9. Fix films 

 for three minutes ; then wash well in methylated spirit. This is an 

 excellent and very rapid method. 



(c) Another excellent method of fixing has been devised by Gulland. 

 The fixing solution has the composition absolute alcohol 25 c.c., pure 

 ether 25 c.c., alcoholic solution of corrosive sublimate (2 grms. in 10 c.c. 

 of alcohol) about 5 drops. The films are placed in this solution for five 

 minutes or longer. They are then washed well in water, and are ready 

 for staining. A contrast stain can be applied at the same time as the 

 fixing solution, by saturating the 25 c.c. of alcohol with eosin before 

 mixing. Thereafter the bacteria, etc., may be stained with methylene- 

 blue or other stain, as described below. This method has the advantage 

 over (a) that, as a small amount of corrosive sublimate is used, less 

 washing is necessary to remove it from the preparation, and deposits are 

 less liable to occur. 



