SPECIAL METHODS. 165 



examined in glycerine, when, if the staining has taken place 

 properly, the deep-red pads of callose stand out sharply in 

 the sieve-tubes. I have found it very useful to first over- 

 stain the sections with corallin solution and then to wash 

 them out with 4% soda solution, which quickly decolorizes, 

 all parts except the callose. This method has given es- 

 pecially good results with the fungi. Preparations stained 

 with corallin cannot be long preserved. 



290. Aniline blue has been recommended by Russow (I) 

 for staining sieve-tube callose. It may be used in a dilute 

 aqueous solution, which is allowed to act half an hour or 

 longer on the sections. Overstained sections may be washed 

 out with glycerine. They are properly stained when only 

 the callose masses appear deeply colored. The " Schlauch- 

 kopfe" of young sieve-tubes (455) are also pretty deeply 

 colored by aniline blue. To distinguish these from the 

 callose, the preparations may be subsequently stained with 

 an aqueous solution of eosin, in which they are left for a 

 few minutes. After a brief washing in glycerine the entire 

 contents of the sieve-cells, including the protoplasmic threads 

 which penetrate the sieve-plates, are colored violet or red,, 

 while the callose pads remain deep blue. These prepara- 

 tions, as well as those with aniline blue alone, can be well 

 preserved in glycerine-gelatine ; or they may be transferred 

 to Canada balsam in the usual way. 



Since it often stains the protoplasm pretty deeply, aniline 

 blue has usually given me much less instructive preparations- 

 than rosolic acid with fungi. 



[2poa. Mangin recommends (IX), for staining the callose 

 of calcified membranes, a mixture of soluble blue extra 6R 

 and vcsuvin, or of the same blue and orseillin BB. These 

 mixtures stain callose blue in a short time, the protoplasm 

 and lignified elements being brown or violet, according to- 

 the mixture used. Where incrustations are not numerous,, 

 as on many leaves, large pieces of tissue may be freed from 

 air by boiling alcohol, then placed in cold nitric acid until 

 frothing ceases, then in cold water, in boiling alcohol, and 

 finally in cold ammonia, to remove xanthoprotein and its 



