WASHING OUT. 57 



General Directions for the Indirect or " Flemming " staining 



Method* 



96, Staining, Sections only can be stained by this method. 

 The solutions employed are made with alcohol, water, or 



anilin, according to the solubility of the colour. There seems 

 to be no special object in making them with alcohol if water 

 will suffice, the great object being to get as strong a solution 

 as possible. The sections must be very thoroughly stained in 

 the solution. As a general rule they cannot be left too long 

 in the staining fluid. With the powerful solutions obtained 

 with anilin a few minutes or half an hour will frequently 

 suffice, but to be on the safe side it is frequently well to leave 

 the sections twelve to twenty-four hours in the fluid. Up to 

 a certain point the more the tissues are stained the better do 

 they resist the washing-out process, which is an advantage. 

 For researches on nuclei the solutions made with anilin had 

 better be employed only with preparations well fixed in 

 chromo-aceto-osmic acid, as the basic anilin oil may easily 

 attack chromatin if not specially well fixed. 



97. Washing out. Washing out is generally done with 

 alcohol, sometimes pure, sometimes acidulated (with HC1). 

 The stained sections, if loose (celloidin sections), are brought 

 into a watch-glassful of alcohol ; if mounted in series on a slide 

 they are brought into a tube of alcohol (washing out can be 

 done by simply pouring alcohol on to the slide, but it is 

 better to use a tube or other bath) . It is in either case well 

 to just rinse the sections in water before bringing them into 

 alcohol. 



The sections in the watch-glass are seen to give up their 

 colour to the alcohol in clouds, which are at first very rapidly 

 formed, afterwards more slowly. The sections on the slide 

 are seen, if the slide be gently lifted above the surface of the 

 alcohol, to be giving off their colour in the shape of rivers 

 running down the glass. In a short time the formation of 

 the clouds or of the rivers is seen to be on the point of ceasing ; 

 the sections have become pale and somewhat transparent, and 



* Historically the principle of this method is due to HERMANN and 

 BOETTCHER ; but it is universally known by the name of Flemming, to 

 whom is due the credit of having greatly improved the method in its prac- 

 tical details. 



