I08 MICROSCOPIC METHODS. 



be cleaned in the mixture of sulphuric acid and potassium bi- 

 chromate described on page 89. 



I. PitfieWs Method as modified by Richard Muir. 

 Prepare the following solutions : — 



A. The Mordant. 



Tannic acid, 10 per cent watery solution, filtered . 10 c.c. 

 Corrosive sublimate, saturated watery solution . 5 c.c. 

 Alum, saturated watery solution . . . .J c.c. 

 Carbol-fuchsin (vide p. 104) ..... 5 c.c. 



Mix thoroughly. A precipitate forms, which must be allowed to deposit, 

 either by centrifugalising or simply by allowing to stand. Remove the clear 

 fluid with a pipette and transfer to a clean bottle. The mordant keeps well 

 for one or two weeks. 



B. The Stain. 



Alum, saturated watery solution . . ." . 10 c.c. 



Gentian-violet, saturated alcoholic solution ... 2 c.c. . 



The stain should not be more than two or three days old when used. It may 

 be substituted in the mordant in place of the carbol-fuchsin. 



The film having been prepared as above described, pour over it as much 

 of the mordant as the cover-glass will hold. Heat gently over a flame till 

 steam begins to rise, allow to steam for about a minute, and then wash well in 

 a stream of running water for about two minutes. Then dry carefully over the 

 flame^and when thoroughly dry pour on some of the stain. Heat as before, 

 allowing to steam for about a minute, wash well in water, dry, and mount in a 

 drop of xylol-balsam. ' 



This method has yielded the best results in our hands. 



2. Ldffler''s Method. 

 Two solutions must be made up as follows : — 



A. The Mordant. 



Tannic acid, 20 per cent aqueous solution . . 10 c.c. 

 Ferrous sulphate, cold saturated aqueous solution . 5 c.c. 

 Fuchsin, saturated alcoholic solution . . . i c.c. 



Mix well, set aside for a few days and filter always before using. This mor- 

 dant improves with age. 



B. The Stain. — Either carbol-fuchsin or aniline gentian-violet will be 

 found to be eminently satisfactory if filtered before using. 



Make a film as above described, and holding the cover-glass in a pair of 

 forceps, pour on as much of the mordant A as the cover-glass will hold. Heat 

 it carefully above a flame till steam begins to rise and then move the prepara- 

 tion gently in and out of the hot-air column over the flame for about 2 minutes. 

 Wash well in distilled water till every trace of mordant appears to be gone. 



