76 THE CELL. 



methods already described may be used; viz., the treatment with pyro- 

 ligneous acid of objects previously fixed in osmic acid mixtures. 



According to Hermann (93, II), sections from such preparations 

 can be double-stained as well as those that have not been treated with 

 pyroligneous acid. They are accordingly stained with safranin in the 

 usual manner, and afterward treated from three to five minutes with 

 the following solution of gentian violet : 5 c.c. of a saturated alco- 

 holic solution of the stain is dissolved in 100 c.c. of anilin water. 

 The 'latter is composed of 4- c.c. of anilin oil in 100 c.c. of distilled 

 water. This is shaken in a test-tube and then filtered through a wet 

 filter. The sections are then placed in a solution of iodin and iodid of 

 potassium (iodin i gm., iodid of potassium 2 gm., water 300 c.c.) 

 until they have become entirely black, after which they are immersed in 

 alcohol until they receive a violet tinge with a slight dash of brown. By 

 this means the chromatin network, the resting nuclei, and the chromosomes 

 in both of the spirem stages appear bluish-violet, while the true nucleoli 

 are pink. The chromosomes of the aster and diaster are colored red. 



Flemming (91, III) recommends the following method: Fixation by 

 his mixture ; the specimens or thin sections are then placed in safranin 

 from two to six days, washed for a short time in distilled water, and then 

 immersed in absolute alcohol weakly acidulated with hydrochloric acid 

 (i : 1000), until no more color is given off. They are then washed again 

 with distilled water and placed in a concentrated solution of anilin-water- 

 gentian-violet from one to three hours. After a third rinsing in distilled 

 water, they come into a concentrated aqueous solution of orange G, until 

 they begin to assume a violet color. Then wash with absolute alcohol, 

 clear in clove or bergamot oil, and mount in Canada balsam. 



A comparatively simple method showing the different structures 

 of the cell and its nucleus with great clearness consists in staining with 

 Heidenhain's hematoxylin. 



Solger (89, I and 91) has discovered that both chromosomes 

 and polar rays are shown in an exquisite manner in the pigment cells of 

 the skin (corium) of the frontal and ethmoidal regions of the common 

 pike (vid. Fig. 37). The preliminary treatment is optional, Flemming's 

 solution or corrosive sublimate being the best. These cells illustrate the 

 stability of the radiate structures of protoplasm, the polar rays showing 

 as parallel rows of pigment granules. 



The various structures of resting and dividing nuclei and cells 

 are of such a complicated nature that they can be observed only with 

 great difficulty in ordinary objects, because of the crowding of so many 

 elements into a comparatively small space. For example, salamandra 

 maculosa, which has become a classic histologic object through the 

 researches of Flemming, possesses somatic cells whose nuclei have no less 

 than twenty-four chromosomes. (It may here be remarked that, curiously 

 enough, salamandra atra has only half this number. ) Consequently, van 

 Beneden's discovery (83), that the somatic cells of ascaris megalocephala 

 have only four primary chromosomes, is a fact of considerable import- 

 ance. Boveri (87, II and 88) has even found an ascaris showing only 

 two chromosomes. As these animals also show distinct achromatic fig- 

 ures in the protoplasm of their ova and sperm cells, they are certainly 

 worthy of being regarded as typic specimens for laboratory __ purposes. 

 The processes of cell-proliferation are almost diagrammatic in their dis- 

 tinctness. 



