92 STUDY OF BACTERIA 



Flood the cover-slip with the mordant and gently steam for one 

 minute, then wash and dry thoroughly, pour the stain on and 

 steam for one minute more. Wash, dry and mount. 



This method yields very good results. 



Pitfield's Method. 



This is the simplest stain and the easiest to use, but does not 

 give the good results that the previous one does. But one solution 

 is needed, this is made in two parts and mixed. 



A. Tannic acid i gram. 



Water 10 c.c. 



B. Saturated watery solution alum (old) 10 c.c. 



Saturated alcoholic solution gentian violet i c.c. 



Mix. 



A heavy precipitate is formed by this process which is useful in the stain- 

 ing. The stain is almost a saturated solution of alum and tannic acid, and 

 when it becomes supersaturated by evaporation and heat, staining takes place. 

 After this the process is very simple. The cover-slip is carefully flooded with the 

 stain and warmed for a minute over the flame of a bunsen burner, turned very 

 low, until steam arises. Not too much stain should be run over the cover-slip. 

 After steaming occurs, the stain should remain for a minute, then the preparation 

 is washed, dried, and mounted. It will be found that the best stained flagella are 

 on those bacteria nearest to the edges where the evaporation has been most 

 intense. If the preparation is not equally stained, Weigert's aniline gentian 

 violet can be run on for a minute to deepen the color. 



Loffler's Method. 



This is the original flagella stain and is a very good one. 

 It is made as follows: 



A. Mordant 



20 percent watery solution tannic acid 10 c.c. 



Sat. solution ferrous sulphate 5 c.c. 



Fuchsin sat. alcoholic solution i c.c. 



Mix 



B. Stain 

 Carbol-fuchsin. 



Proceed as in the previous methods. 



