278 OYTOLOGICA.L METHODS. 



the last two in saline media. It is sometimes advisable to 

 rub them up with serum, as recommended by v. LA VALETTE 

 ST. GEORGE. These methods are most important for the study 

 of the Nebenkern. 



The student will remember that no known reagent will 

 stain any part of the nucleus whilst alive. The " Nebenkern " 

 stains sometimes, but feebly. Most frequently the colour is 

 only taken up by certain granules of the cytoplasm, which 

 may or may not be identical with the " granules " or " bio- 

 blasts " of ALTMANN. 



These matters have already been discussed in the paragraph 

 headed " Staining ' intra vitam, 3 " 93. 



601. Study of Fresh and Lightly fixed Cells. It has been 

 rightly pointed out by Flemming that so-called " indifferent " 

 liquids must not be believed to be without action on nuclei. 

 Iodised serum, salt-solution, serum, aqueous humour, lymph, 

 better deserve the name of weak hardening agents. Between 

 these, and such energetic hardening agents as Flemming' s 

 mixture, come such light fixing agents as picric acid, or very 

 dilute acetic acid. These it is whose employment is indicated 

 for the study of fresh isolated cells. 



A typical example of this kind of work is as follows : Tease 

 out a piece of living tissue in a drop of acidulated solution of 

 methyl green (0*75 per cent, of acetic acid). This is a 

 delicate fixing agent, killing cells instantly without change of 

 form. Complete the fixation by exposing the preparation for 

 a quarter of an hour to vapour of osmium ; and add a drop of 

 solution of Ripart and Petit, and a cover. 



Or you may fix the preparation, after teasing, with vapour 

 of osmium for half a minute to two minutes, then add a drop 

 of methyl green, and after five minutes wash out with 1 per 

 cent, acetic acid, and add solution of Ripart and Petit, and 

 cover. 



Or you may kill and fix the cells by teasing in solution of 

 Ripart and Petit (to which you may add a trace of osmium 

 if you like), and afterwards stain with methyl green. 



Other fixing agents, such as picric acid, or weak sublimate 

 solution, may of course be used, and in some cases doubtless 

 should be preferred. Other stains, too, such as Bismarck 

 brown, or Del afi eld's haBinatoxylin, may be used as occasion 



