64 CALIFORNIA ACADEMY OF SCIENCES. [Proc. 3D Ser. 



The following fixing fluids were tried: Flemming's 

 strong mixture; one per cent, chromic acid; two per cent, 

 iron trichloride; Wilson's corrosive sublimate-acetic; Bove- 

 ri's picro-acetic; Lindsay's potassium bichromate-platinum 

 chloride-osmic-acetic; one per cent, palladium chloride and 

 one-half per cent, iridium chloride. 



Flemminfj's strong mixture was used undiluted and also 

 with the addition of one, two and three parts of water. The 

 quantities of chromic and acetic acids in the original mixture 

 were also varied. Dilute solutions were found to shrink the 

 cells much more than the strong ones. Var3ang the amount 

 of chromic acid did not improve the action of the fixing 

 fluid. An increase in the amount of acetic acid, however, 

 gave the best results obtained. Flemming's strong mixture 

 with an excess of acetic acid was therefore almost exclu- 

 sively used. Fair" results were also obtained with palladium 

 chloride and iridium chloride to which a small amount of 

 glacial acetic acid had been added. 



After remaining in the fixing fluid for twenty-four hours, 

 the anthers were washed in running water for six hours. 

 They were then placed in a deh3^drator ^ for twenty-four 

 hours, with 95 per cent, alcohol below and distilled water 

 above. 



Some alcohol was then removed from the material and 

 mixed with an equal volume of 95 per cent, alcohol. The 

 material was transferred to this stronger mixture for two 

 hours. By repeating this process three or four times the 

 material was brought into 95 per cent, alcohol without 

 shrinkage. It was left in 95 per cent, alcohol for twent}^- 

 four hours. It was then placed for six hours in each of the 

 following, successively: absolute alcohol; absolute alco- 

 hol and bergamot oil (equal parts) ; bergamot oil ; bergamot 

 oil and paraffin, 47° (equal parts); paraffin 47°; paraffSn 

 47*^ and paraffin 54^ (equal parts); paraffin 54°. 



Sections 3 to 4 /i in thickness were cut with the Minot 

 wheel microtome. Of the stains tried, Flemming's triple 

 stain (safranin, gentian violet and orange "G ") gave the 

 best results. 



1 For a description of the dehydrator see Lawson, 1898, and Williams, 1899. 



