5 o CULTURE METHODS AND TECHNIQUE 



readily penetrable tissues may be stained in mass with a ground 

 stain, but in general it is preferable to stain sections on the slide. 

 Material of the fleshy fungi more often lends itself to mass stain- 

 ing, and this process becomes particularly desirable, moreover, 

 when carmine stains are advised. The carmine stains are most 

 important if the material has been fixed in sublimate mixtures. 



A successful method of staining fleshy fungi for histological 

 differentiation has been used by Burt as follows : Alcoholic 

 material is stained in toto twenty-four hours in Mayer's alcoholic 

 paracarmine. When the sections have been mounted on the slide 

 and dehydrated, they are stained for about five minutes in an 

 aqueous solution of fairly strong safranin, and finally washed in 

 water, previous to mounting in water and glycerin. With tissues 

 fixed in sublimate mixtures, the Ehrlich-Biondi-Heidenhain stain 

 has also been found to give effective histological differentiation. 

 This strain should be obtained ready-mixed from Grubler & Co. 

 To 100 cc. of 0.4 per cent solution of this mixture must then be 

 added 7 cc. of a \ per cent acid fuchsin solution. This stain 

 gives some nuclear differentiation, but it cannot by any means 

 be called a successful nuclear stain. Only small quantities of this 

 stain should be combined with the fuchsin at one time, since its 

 keeping qualities are not good. 



A double stain of any standard haematoxylin followed by 

 erythrosin eosin or orange G is sometimes to be recommended 

 when material is fixed in alcohol ; but, in general, many haema- 

 toxylin stains are not so valuable for work with the fungi as with 

 the higher plants. Magdala or Congo red may be followed by an 

 anilin blue or green to advantage. A stain of any solution of eosin 

 or carmine followed by a counter stain of nigrosin is often of value. 

 In cytological work more than in any other kind, it is necessary 

 to bear in mind the nature of the fixing agent in deciding upon 

 an effective stain. After sublimate fixing, one of the most success- 

 ful methods of staining on the slide for the differentiation of cyto- 

 plasmic and nuclear structures is one apparently first published by 

 Wager. It is a cumbrous and complicated process in print, but is 

 much simpler in practice. As I have used this stain, it is a slight 

 modification and simplification of Wager's process. The principal 

 solutions needed are : 



