154 



SCIENCE 



[N. S. Vol. XXXVI. No. 918 



the point of a dissecting needle. The pollen 

 mother-cells float out uninjured. Most other 

 genera are easily treated in the same way. A 

 few genera present difficulties; e. g., the Mal- 

 vaceaa, where the anthers are so charged with 

 mucilage that the mother-cells can not be 

 handled successfully without considerable 

 trouble. 



Before passing to the technic, which is ex- 

 ceedingly simple, I wish to say that the 

 method can readily be adapted to study of the 

 cells of more compact tissues by the simple 

 process of teasing out with needles a few cells 

 and separating them from the rest of the tis- 

 sue. Although the microtome method is of 

 extreme value to every worker, it has the tend- 

 ency of tyrannizing over all other methods, 

 some of which are unquestionably better for 

 special purposes. 



The killing of the pollen mother-cells can be 

 done either while they are still in the anthers 

 or after their separation. I have found weak 

 Flemming's solution excellent, but Bouin's 

 solution, on the whole, the better for this pur- 

 pose. It has long been a favorite of the zoolo- 

 gists, but rather neglected by botanists. The 

 formula is : 



Picric acid, sat. aq. sol 75 



Acetic acid, glacial 5 



Formalin, commercial 20 



Fix 4 to 8 hours; wash with 50 per cent, alcohol 

 until no color remains, in which the material may 

 then be kept indefinitely. 



After killing and washing, the pollen 

 mother-cells are stained in toto by any satis- 

 factory method. I find that both Heiden- 

 hain's iron-hematoxylin and Hermann's modi- 

 fication of riemming-'s triple stain are espe- 

 cially good. The latter works vastly better 

 than the regular Plemming stain. The for- 

 mula is: 



Safranin, water soluble 1 



Alcohol 10 



Anilin water 90 



Stain 4 to 8 hours; wash in 50 per cent, alcohol 

 and, if necessary, in acidulated 50 per cent, alcohol. 

 Pass into 



Gentian violet 1 



Alcohol 10 



Anilin water 90 



Stain 2 to 6 minutes; wash in water. Pass into 

 Orange G., aq. sol. 



I find the concentrated aqueous solution too 

 intense, therefore dilute it with 9 volumes of water. 

 Stain 1 to 3 minutes. Wash quickly in 50 per 

 cent, alcohol and finish with absolute alcohol. 



The material is cleared in cedar oil or, where 

 dampness is a drawback to this method, in oil 

 of cloves. There need be no shrinkage what- 

 ever in the finished preparation and there can 

 be practically no disturbance in the arrange- 

 ment of the cell contents. 



Pollen mother-cells thus prepared present to 

 the investigator the original packages with 

 unbroken walls, from which no histological 

 particle has escaped. The karyokinetic figures 

 are complete. The chromosomes are all in 

 situ, not sliced up into incomplete ribbons 

 that need to be matched in successive sections, 

 but each one complete and undisturbed. The 

 whole machinery of mitosis, as well as all the 

 adjacent cell contents, make an unbroken unit. 

 The picture under the eye is that of a lot of 

 spheres, transparent, translucent, revealing the 

 contents, with as little chance of artifacts as 

 is possible by human methods. 



By mounting such cells in a somewhat 

 limpid medium, such as heavy glycerine or 

 thin Canada balsam and placing a triangular 

 blotting-paper at one side of the cover-glass, 

 the cells may be rolled over under the ob- 

 server's eye, presenting to view all sides of the 

 karyokinetic spindle and enabling one to count 

 the chromosomes, to notice their position, and 

 to study the entire mechanism with an ease 

 and an absolute certainty that no series of 

 sections can possibly equal. 



Three points in this method deserve em- 

 phasis : 



1. The comparative ease with which pollen 

 mother-cells can he secured in an unhroTcen 

 and perfect state, and prepared for observation 

 as to their internal structure. 



2. The positiveness of interpretation that 

 can be secured by this method in contrast to 



