1 6 MARGARET C. FERGUSON 



prepared by Dr. G. Griibler ; Guignard's combination of methyl 

 green, acid fuchsin, and orange G; Flemming's safranin- 

 gentian-violet-orange combination : and Heidenhain's iron- 

 haematoxylin. The last two proved the most satisfactory and 

 were almost exclusively used. The iron-haematoxylin was fol- 

 lowed by orange G, or, if it was desirable to stain cell-walls, 

 by Bismarck brown. Iron-haematoxylin followed by Flem- 

 ming's triple stain, or by gentian-violet and orange G, brought 

 out the so-called kinoplasmic structures with great definiteness. 

 The best differentiation was obtained with the iron-haematoxy- 

 lin by allowing the haematoxylin to act from 12 to 18 hours, 

 decolorizing in iron-alum, and then washing in running tap- 

 water from 2 to 6 hours. Flemming's triple stain was often 

 used without the safranin with excellent results. Both anilin 

 and aqueous solutions of gentian-violet were used. As a rule, 

 a one-half percent, solution was employed, the slides remain- 

 ing in it from 5 to 20 minutes. The achromatic figures in the 

 divisions of the pollen-mother-cell, especially in Pinus Strobus, 

 were, however, brought out with great difficulty with this stain. 

 The best results were obtained for these stages by allowing the 

 slides to stand from 24 to 48 hours in stender dishes of distilled 

 water to which not more than 10 drops of a one percent, solution 

 of gentian-violet had been added. Pinus sections take the 

 orange with such avidity, that a fraction of a minute was in all 

 cases a sufficiently long time to allow this stain to act. After 

 washing out the superfluous gentian-violet and' dehydrating in 

 absolute alcohol, differentiation was effected by dashing with 

 clove oil. Bergamot oil was used for fixing and clearing, and 

 I have found it expedient to pass the slides from bergamot oil 

 to jars of xylol. They can remain in the xylol for hours, if 

 desirable, without injury, and the xylol is so readily miscible 

 with the balsam that the preparations become clear and more 

 satisfactory for studying in a much shorter time than when car- 

 ried directly to the balsam from the bergamot oil. 



