172 
alcohols, to xylol, and finally mounting in Canada balsam. Or, 
and here I am about to describe the second of the two methods 
promised above, the following much more complex procedure 
may be adopted when certain finer histological details concerning 
the protoplasm and nuclei are in question. 
The slides with their adherent sections are removed from the 
alcohol to a solution of the intense red dye Safranin, and left 
in this for several hours. | When deeply stained, they are rapidly 
rinsed in water, and if necessary in alcohol to remove excess of 
the dye, and then plunged into a dilute watery solution of gentian 
violet, and again left for some minutes to an hour or more, until 
they appear deep blue-purple. | After another quick washing, to 
remove excess of adherent dye, they are put for a few minutes 
into a bath of a I per cent. solution of a dye known as Orange 
G., which exerts some peculiar action facilitating the removal of 
the excess of the violet without at the same time washing out the 
Safranin. The preparations are then finished in absolute 
alcohol, transferred to clove-oil, and finally mounted in balsam. 
It is impossible here to go into all the small points which have 
to be attended to in obtaining the proper intensity of staining, 
dehydration, &c., in this admittedly complex method,” but the 
results in a successful preparation where the more solid parts 
of the nuclei are brilliant red, other granules violet, and the 
nuclei and chlorophyll-granules of the host-plant picked out in 
equally sharp colours, are worth the trouble in certain cases. 
I now leave the subject of methods of investigating the dead 
structures, and proceed to the consideration of methods of culture 
and examination of the living fungus; methods which are, in 
my opinion, of more importance in the present state of science, 
and destined to teach us far more of the nature of these extra- 
ordinary organisms. It is obvious at the outset that our methods 
must differ according to the kind of fungus we are going to cult 
vate; not only will saprophytes behave very differently from 
parasities, but each group presents its own problems in cultiva- 
tion, and many of these problems are as yet unsolved. . 
Commencing with saprophytes, as the easier to deal with m 
practice, I distinguish between methods of isolation and methods 
of culture proper. Suppose, for instance, that we have gathered 
a species of Mucor, Sporodinia, Phycomyces, Rhizopus, or other 
easily grown Mucorine, the probability of our obtaining a pwe 
culture by direct sowing of the spores on any medium Is almost 
infinitely remote. Indeed it is sheer waste of time to attempt 
it in most cases, since the following simple method of isolating 
the fungus from its contaminating bacteria, moulds, &c., can be 
so readily employed. 
* See, for further details, Zimmerman, Hoff, Botanisches Centralblatt, 
1898, vol. 76, p. 66, 
