BINUCLEATE CELLS IX TISSUE CULTURES. 85 



be interpreted as due to the fixative they should be abundant. Again, the actual 

 observation of such nuclei in living cells is proof absolute that they are not artifacts. 



The only type of nuclear fission which I have observed in tissue cultures is that 

 which occurs, apparently, by constriction. 



An estimate of the frequency of occurrence of such transitional amitotic nuclear 

 forms as those shown in figures 2, 3, 6, etc., was made by making careful counts, the 

 aforementioned series of 20 heart cultures being used. Out of a total of 41,725 

 cells in this series, 50 cells were found to contain constricted nuclei of such a character 

 as would warrant their being considered as amitotic. This is a proportion of one 

 amitotic nucleus to 835 ordinary nuclei, or 0.1198 per cent. In the same series 

 there were 375 binucleate cells, which are regarded as end products of nuclear 

 amitosis. The proportion of transitional forms to end products is thus 50 : 365, 

 1 : 7.5 or 13.33 per cent. So high a percentage of transitional forms seems to indi- 

 cate that the nuclei remain a long time in this condition, and the observations upon 

 living cells bear this out. The final stages of direct nuclear fission, as shown in 

 figures 6 and 8, are, as has been noted, rarely found. 



In this connection it is of interest to compare the incidence of amitotic with 

 that of mitotic nuclei. In the same series there were found to be 170 cells undoubt- 

 edly in mitosis. The ratio of mitotic cells to total cells is thus 170:41,725, or 

 1 : 245, or 0.4 per cent. There were probably many more mitoses than this, for some 

 are undoubtedly rubbed off in preparation, since their rounded and thickened form 

 exposes them to friction in washing, etc.; also 62 doubtful mitotic forms were not 

 included. 



It is an easy matter to calculate the relative proi)ortion of amitotic to mitotic 

 forms. As has been stated, the ratio of amitotic nuclei to total cells is 1 : 835, while 

 that of mitotic nuclei to total cells is 1 : 245. It is evident that the mitotic forms 

 are 3.4 times as numerous as the amitotic, even when we leave out the doubtful 

 forms and the cells in mitosis which have been rubbed off. Again, when we con- 

 sider that the amitotic process is a slow one, as has been shown, and that mitosis 

 is relatively rapid (1 j to 2| hours according to Lewis and Lewis, 1915, p. 371), it will 

 be realized that the amitotic method of nuclear division is unimportant, so far as 

 nuclear multiplication is concerned, as compared with mitosis. 



Thus, examination of living and fixed preparations makes reasonable the view 

 that direct division of the nucleus occurs where this structure is elongated, and 

 sometimes bent upon itself, by a karyoplasmic streaming, away from the nuclear 

 equator, and a gradually deepening constriction which encircles the nucleus more or 

 less symmetrically and cuts it into two parts, the constricted area becoming a 

 narrow tube and finally a thread, which ultimately disappears. 



The behavior of the nuclear membrane during amitosis in the cells of tissue 

 cultures seems to be essentially the same as that of the same structure in the cells 

 of the trematode described by Cary (1909) during intranuclear mitosis. There is, 

 however, no intranuclear sjiindle in the cells which I have examined. 



The final separation of the consti'icted nucleus takes only a short time, as has 

 been noted, but a nucleus may remain for a long time apparently about to divide 

 without actuallj' doing so. 



