CHROMATIN, ANIMAL CHROMOSOMES, NUCLEOLI 267 



Painter {Anat. Rec, xxvii, 1924, p. 77). and many others, 

 have used a slight modification of Allen's modihcation of Bouin's 

 lluid (10 c.e. of aeetie acid is used instead of 5 c.e. in the formula 

 given in § 115). Tissue is cut into small pieces and teased directly 

 in the preservative with needles. Fixation lasts for 1^ to 3 hours. 

 The drop method is used for changing fluid, and Allen's anilin- 

 wintergreen oil method of clearing is followed (see § 629 for 

 details). 



The Japanese cytologists have been using, with success, either 

 the original Champy's fornmla, or some modification of it. (See 

 especially Minouchi, Jap. Jour. ZooL, viii, 1928, p. 219, also 

 § 632 below.) After sectioning and bleaching, sections are treated 

 for twenty-four hours with Chura's solution {Zeit. rviss. Mikr.. 

 xiii, 1925), which consists of equal parts of glacial acetic acid and 

 a saturated aqueous solution of picric acid, in order to dissolve 

 out the cytoplasmic inclusions and to render the chromosomes 

 more deeply staining. 



If the tubules of the testis are teased apart, it is important 

 that these small structures be protected from injury during 

 dehydration and imbedding. In the higher grades of alcohol, 

 especially after osmic acid fixation, the tubules become extremely 

 brittle and the very best fixed material may be lost. It is advisable 

 to put the testicular fragments into a little cage of some sort 

 during dehydration, clearing and imbedding. The common 

 Gooch crucible is good for this purpose, or a small square of porous 

 cloth may be folded into a bag to hold the material. 



629. Precautions in Dehydrating and Clearing. It is well 

 known that too rapid dehydration and clearing will cause shrinkage 

 and often distortion. Material to be used for chromosome study 

 should be dehydrated gradually, either by the drop method, or 

 by some one of the devices which have been developed to ensure 

 the gradual exchange and diffusion of fluids. For clearing, 

 the least shrinkage seems to occur with some of the 

 vegetable oils, such as bergamot, wintergreen, cedar wood or 

 origanum, and if any of these do not mix readily with paraffin, 

 the clearer can be washed out with benzol or xylol. The use of 

 xylol, as a clearing agent directly from alcohol, is not recom- 

 mended for delicate tissues as it causes some shrinkage. 



Ezra Allex {Anat. Rec, x, 1915, p. 565), following Suchannek 

 (§ 150) uses anilin oil as a substitute for alcohols by a method 

 developed to follow his modification of Bouin's lluid, but applic- 

 able generally to other types of fixation except those involving the 

 use of osmic acid. 



Following Allen's lead, Painter employs the following pro- 

 cedure : After fixation with Bouin-Allen, small pieces of the 

 testis are placed on a square of cloth of porous weave, the four 

 corners are brought together, forming a sort of bag. A pin is 



