ZOOLOGY AND BOTANY, MICROSCOPY, ETC. 
445 
rapidly. By this procedure the bacteria are not only fixed, but deeply 
stained, and are quite ready for microscopical examination. For staining 
bacteria, in pus, faeces, and in urinary sediment, a better result is obtained 
by using a mixture of phenol-fuchsin and alcoholic solution of methylen- 
blue ; for thereby the nuclei of the cells and the bacteria are stained blue, 
while the cell-protoplasm and degenerated bacteria are stained red. 
Observing Nuclear Division in Equisetum and Chara. — Herr W. 
J. Y. Osterhout* finds Flemming’s mixture the best fixing material for 
observing the division of the nucleus in the spore-mother-cells of Equi- 
setum (see p. 416). Microtome-sections 5 /x thick were stained with 
safranin, gentian violet, and orange G, and mounted in Canada-balsam. 
The processes employed by Herr B. Debski f in observing the division 
of the nucleus in the vegetative cells and in the antheridial filaments 
of Chara (see p. 416) are described in <fetail. The best fixing material 
was found to be Flemming’s mixture ; and as a stain, iron-alum hasmato- 
xylin and Flemming’s safranin-gentian-orange gave the best results. The 
material was left for 24 hours in the fixing fluid, then for 1-2 hours in 
running water ; then placed for about 12 hours successively in 10, 15, 
20, 30, 50, 75, 90 per cent, and absolute alcohol ; and finally transferred 
through chloroform-alcohol and chloroform to chloroform-paraffin. The 
chloroform was slowly evaporated, and the material then imbedded in 
pure paraffin of 52° C. melting-point. Microtome-sections 5 u thick 
were fixed to the slide by distilled water mixed with some albumen, and 
dried. The paraffin was removed by xylol, and the xylol by alcohol, 
and the sections were then stained. 
Fixing, Imbedding, and Staining for Nuclear Division in Pollen- 
Grains.;!; — The following are the methods employed by Miss E. Sargant 
for the researches described on p. 398 : — 
A. Fixing. — Anthers fixed in absolute alcohol were usually uncut. 
Those fixed in any of the three solutions given below were either halved 
transversely or cut at both ends to ensure penetration. 
Hermanns Solution { alcoholic ). — 10 per cent, aqueous solution of 
platinic chloride, 3 ccm. ; 1 per cent, osmic acid (aqueous), 8 ccm. ; 
glacial acetic acid, 2 ccm. ; absolute alcohol, 27 ccm. The anthers were 
left in this solution for 1 J-2 hours, and then transferred to a 0 • 5 per 
cent, aqueous solution of platinic chloride for 24 hours. They were then 
placed in a 1 per cent, aqueous solution of platinic chloride for 24 hours. 
Flemming's Solution {aqueous). — 1 per cent, aqueous solution of 
chromic acid, 30 ccm. ; 1 per cent, osmic acid (aqueous), 8 ccm. ; glacial 
acetic acid, 2 ccm. The anthers were left in this solution for about 
2 hours, and then transferred to an aqueous 0 * 5 per cent, solution of 
chromic acid for 18 hours. 
Chromic acid {aqueous). — The anthers were laid in a 0 • 5 per cent, 
aqueous solution of chromic acid for 18-24 hours. 
After treatment in any of these ways, the anthers were rinsed in 
water, and transferred successively, at intervals of about 12 hours, to 
30, 50, and 70 per cent, alcohol, and finally left for several days in 
* Jahrb. f. wiss. Bot. (Pfefifer u. Strasburger), xxx. (1897) p. 159. 
f Tom. cit., pp. 229-31. 
