BOT.-VOL. II.] PEIRCE— ROOT-TUBERCLES. 297 



preparations of bacteria. The stains were made up 

 according to the directions given in Humphrey's Zimmer- 

 mann's Botanical Microtechnique (p. 186), and were used 

 as Hof (1898) directs, except that after the sections had 

 been for not more than two minutes in the gentian violet 

 solution they were rinsed with water and placed for a 

 half hour or longer in Gramm's iodine solution to differ- 

 entiate the baciUi and the infection threads from the cyto- 

 plasm. Hof says that the sections may be left from two to 

 three minutes in the anihn gentian violet. I often found 

 this quite too long, and had difficulty in washing out enough 

 of the violet without taking out the safranin also. One 

 minute is usually long enough for these tissues. Washing 

 off the Gramm's iodine with water, the slides were then 

 allowed to remain for one to two minutes in staining bottles 

 containing orange G; they were then washed with absolute 

 alcohol so long as gentian violet came off abundantly or 

 needed to be removed (as shown by microscopic examina- 

 tion), were cleared in clove oil, and mounted in xylol 

 balsam. I decidedly prefer clove oil to xylol for clearing, 

 as it aids in the differentiation for which this staining 

 method is so highly prized. 



So far as my experience goes, this method of fixing and 

 staining is perfectly certain to demonstrate the infection 

 threads and to differentiate the bacteria in the cytoplasm 

 and in the unstained matrix of the threads. I am, therefore, 

 somewhat at a loss to understand the difficulties reported 

 by some authors in staining tubercles and their contents. 

 Miss Maria Dawson (1899, p. 8) reports, for example: 

 "For some time I made use of both hand-sections and 

 microtome sections of paraffin material. The latter method 

 I afterwards abandoned, however, since I found the tuber- 

 cle tissues very difficult objects to stain upon the slide, and 

 also ordinarily thin hand-sections serve better for the exam- 

 ination of the filaments within the cells — a point to which I 

 wished to devote special attention." With the stains Miss 

 Dawson used on hand-sections, and of which she speaks 



