go CLINICAL BACTERIOLOGY AND HEMATOLOGY 



scrupulously clean slide, allow to dry, and fix by gentle heat — 

 i.e., such that it does not get uncomfortably hot to the finger. 



Prepare a mixture of lo c.c. tap-water (or of distilled water 

 + I drop of a I in i,ooo solution of potassium carbonate), and 

 10 drops of Giemsa's stain, which must be bought ready pre- 

 pared. In mixing the two avoid violent agitation ; add the stain 

 to the water in a test-tube, stopper the latter with your thumb and 

 invert it slowly once or twice. Do not mix it until you are ready 

 to proceed with the staining. 



Take the slide in a perfectly clean pair of forceps, flood it with 

 the stain, and heat until it just begins to steam ; remove it from 

 the flame and in fifteen seconds pour off the solution, replace it 

 quickly with fresh and heat again, again removing it when steam 

 rises and allowing the action to go on for fifteen seconds. Do 

 this four times in all, allowing the action to go on for one minute 

 on the last application. Then wash in tap- water or in distilled 

 water with a drop or two of potassium carbonate solution, blot, 

 dry, and mount. 



Another method is to place the slide face downwards in a Petri 

 dish, supported on two slips of glass. The dish is then filled with 

 a mixture prepared as above, and the staining allowed to go on 

 for twelve to twenty hours in the cold, or three to four hours in 

 the incubator, the dish being covered to prevent evaporation. If 

 the slide is inserted face upwards, it will probably be covered by 

 a fine red precipitate. 



The most minute amount of acid is fatal to the process ; hence 

 all instruments must be dry and clean, and distilled water (which 

 often contains traces of acids) should be avoided. 



Examine the film with the J^ and your highest eyepiece, taking 

 great care to get a good light, white if possible. 



Another method, with which I have been fairly successful, and 

 which is strongly recommended by Herxheimer, consists in 

 fixing as before, and allowing the film to stain for twenty-four 

 hours in a I in i,ooo solution of Nile blue in distilled water. The 

 spirochaetes are stained a fine blue, and are readily recognizable. 



In making the search a good lens and good light are always, 

 and much patience frequently, necessary ; the spirochaetes may 

 be but one or two on a film, or there may be several on one field 

 of the microscope. Very occasionally they are matted together 

 in a dense mass. Having found a spirochsete, proceed to see if 

 it resembles the pallida or the refringens ; note especially its 



