ISOLATION AND COMPOSITION OF NUCLEI AND NUCLEOLI 101 



A third recently described homogenizer for making homogenates on a 

 somewhat smaller scale than with the Waring Blendor or colloid mill is the 

 stainless steel apparatus described by Lang and collaborators.'® 



Apart from various other high-speed mixers, which undoubtedly disrupt 

 mitochondria,"'^ the devices just described constitute the principal means 

 known at the present time for preparing homogenates to be used in isolating 

 cell nuclei on a relatively large scale. It is possible that a device such as the 

 Latapie homogenizer might be useful in certain instances, but the writer has 

 had no experience with this apparatus. In the disruption of starfish egg 

 cells prior to the isolation of nucleoli, homogenization has been accom- 

 plished by forcing a cell suspension through a hypodermic needle.'^ 



The best-known small-scale homogenizer which can be used for obtaining 

 homogenates useful in isolating cell nuclei on a small scale is the ground- 

 glass apparatus now known as the Potter-El vehj em homogenizer.'^ A device 

 built on exactly the same principle was first described by Hagan in 1922,^" 

 as noted by Potter .2' The Potter-Elvehjem homogenizer was constructed in 

 a different manner and the slight modification of fusing small glass beads 

 on the bottom of the pestle was introduced, but the final product was of 

 practically the same design as the one described by Hagan. 



The Hagan-Potter-Elvehjem homogenizer has been used extensively by 

 Schneider and Hogeboom and their followers in preparing homogenates 

 from which cell nuclei are to be obtained. It has two inherent advantages, 

 one of which is the forcing of all of the homogenate between the homogeniz- 

 ing surfaces every time the ground test-tube is run up or down the rotating 

 pestle. The other is a relative gentleness of action which leaves the mito- 

 chondria in an undamaged or only very slightly damaged condition, 

 provided that the proper homogenizing medium is used. The nuclear mem- 

 branes also appear to escape damage to a considerable extent. Certain dis- 

 advantages are found, however, when this instrument is used for homogen- 

 izing tissue preparatory to the isolation of nuclei, of which the most serious 

 is the failure to break a sufficiently high percentage of the cells. Residual 

 whole cells tend to concentrate with nuclei and can be removed only by 

 special methods such as gravity sedimentation in a two-section cell' or 

 filtration through cotton flannel.' '^'^^ But if nuclei are heavily contami- 

 nated with whole cells, it is practically impossible to separate them. 



i«K. Lang and G. Siebert, Biochem. Z. 322, 360 (1952). 

 17 K. S. McCarty, J. Exptl. Med. and Surg. 7, 213 (1949). 

 '8 W. S. Vincent, Proc. Natl. Acad. Sci. U.S. 38, 139 (1952). 

 1" V. R. Potter and C. A. Elvehjem, J. Biol. Chem. 114, 495 (1936). 

 2»W. A. Hagan, /. Exptl. Med. 36, 711 (1922). 

 21 V. R. Potter, J. Biol. Chem. 163, 437 (1946). 



" G. H. Hogeboom, W. C. Schneider, and M. J. Striebich, /. Biol. Chem. 196, 111 

 (1952). 



