As the seventh methyl group is very loosely attached, there is 

 always some methyl violet present, either because it is not all com- 

 pletely converted into the higher homolog or because it has broken 

 down again. It has been stated that to obtain free methyl green 

 the commercial dye should be shaken in a separatory funnel with 

 amyl alcohol or chloroform, which dissolves the methyl violet. As 

 a matter of fact, however, pure methyl green may not be always 

 desired by the biologist, as the dye owes part of the metachromatic 

 properties for which it is prized to the presence of small amounts of 

 the violet compound. 



Methyl green is at present one of the most valuable nuclear 

 stains known to the histologist, and is widely used as a chromatin 

 stain by the cytologist. On the other hand it has been used by 

 Galeotti as a cytoplasm stain following acid fuchsin and picric 

 acid. In the Ehrlich-Biondi technic it is used to stain nuclei in 

 contrast to acid fuchsin; while Bensley employs it to stain chroma- 

 tin in contrast to acid fuchsin which stains the mitochondria. It is 

 an ingredient of the Ehrlich triacid mixture (with orange G and 

 acid fuchsin) for staining blood smears. Botanists find it a valu- 

 able stain, combined with acid fuchsin, for lignified xylem. One of 

 its most valuable uses today is in the Pappenheim stain, in which 

 it is combined with pyronin and used for staining the gonococcus 

 and mast cells as well as by Unna in studying chromolysis. It is 

 also a useful chromatin stain for protozoa, and is employed in weak 

 acetic acid solution for staining fresh material beneath the cover- 

 glass. 



When the foreign supply of dyes was first shut off, this stain 

 proved one of the most difficult to obtain in satisfactory quality, 

 largely due to the looseness with which the seventh methyl group 

 is attached and the resulting instability of the compound. At first 

 certain green dyes of an entirely different nature were furnished, 

 but as soon as an investigation of the dye was begun manufac- 

 turers proved perfectly able to produce methyl green; the difficulty 

 came in obtaining the right degree of purity. Samples were finally 

 furnished so pure that they lacked completelj^ the necessary meta- 

 chromatic staining quality; and it proved necessary to add a certain 

 small percentage of the violet dye to obtain the proper results. This 

 problem seems to have been solved at present and satisfactory 

 methyl green is available. The chief problem now is to standardize 

 it. With other stains this can ordinarilv be done on the batch 

 basis, approving some batch large enough to meet the demand for 

 a period of years. With methyl green this cannot safely be done, 

 on account of its instability. Hence large batches are impractical; 

 and the stain ought to be sold with the caution that the dye does 

 not keep indefinitely without change. That this is not generally 

 realized is shown by the fact that when a certain company recently 

 announced for sale a supply of German stains imported before the 

 war and kept on their shelves since then, one laboratory ordered a 



70 



