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1909] SAXTON—WIDDRINGTONIA CUPRESSOIDES 163 
corrosive sublimate, 52™.' Material was fixed for 24 hours and 
then washed in 50 per cent. alcohol until the alcohol was no longer 
colored yellow (a week or more as a rule), then 12-24 hours in 75 per 
cent., 85 per cent., and 94 per cent. alcohol, and at least 60 hours 
in absolute alcohol changed three or more times. Both xylol and 
cedar oil (in 25 per cent. grades with alcohol) were tried to precede 
infiltration with paraffin, but the former was entirely discarded 
_ after a few trials. Material was taken from cedar oil through 25 per 
cent., 50 per cent., and 75 per cent. solutions of soft paraffin in cedar oil, 
to pure paraffin, melting point 48° C., using tiny wire-gauze baskets, 
for the purpose, as described by Frercuson (14), and allowing 
48 hours in each solution. At least 48 hours longer was allowed in 
two fresh lots of soft paraffin (an indication of the time required here 
is given by the time taken to wash out the fixing agent), 12-24 hours 
in a mixture of soft and hard paraffin, and finally 24 hours in hard 
paraffin, melting point 55°C., in which it was imbedded. These 
very long periods in the oven were found to be absolutely necessary 
to insure proper infiltration of the para fin. 
In young stages the ovules were fixed whole, or when very young 
@ small part of the tissue of the cone was cut off bearing the ovules. 
Later the integument becomes too hard to cut, and it was necessary 
to dissect out the nucellus, a process which can be carried out 
fairly easily in practice. In some stages after fertilization the pro- 
thallus was dissected out, and where nearly mature embryos were 
Present these were fixed alone. The staminate cones were fixed 
whole, being very small, but required even longer periods in the oven 
than those mentioned above. 
The stains were (1) Delafield’s hematoxylin diluted 8-10 times, 
allowed to stain 4-12 hours, extracted with very dilute acid (aqueous), 
and followed by a thorough washing in water; and (2) Flemming’s 
triple stain. The best results were obtained with the former, 
‘specially after the fixing agent mentioned above. It appears to 
be always the case that the very best staining effects of Delafield’s 
_ hematoxylin are obtained after a fixing agent containing corrosive 
sublimate. The value of this hematoxylin as a nuclear stain has 
Not at present been realized. 
* am indebted to Mr. A. J.B range ing the use of this reagent 
do) 
