METHYLENT BLUE. 207 



343. Fixation of the Stain. — I'lie staia obtained by any of 

 these methods may be fixed, and more or less permanent 

 preparations be made by one or other of the following 

 methods : 



Aenstein {Anat. Anz., 1887, p. 551) puts the tissue for 

 half an hour into saturated aqueous solution of picrate of 

 ammonia. 



S. Mayee [Zeit. wiss. Mik., vi, 1889, p. 422) preferred a 

 mixture of equal parts of glycerine and saturated picrate of 

 ammonia solution, which served to fix the colour and mount the 

 preparations in. This was also in principle the method of 

 Retzius [Intern. Monatschr. Anat. Phyx., vii, 1890, p. 328). 



DoGiEL [Encyd. mik. Techn., ii, p. 105) puts for 2 to 24 

 hours into saturated aqueous picrate of ammonia, and then 

 into equal parts of glycerin and the picrate solution. (Thin 

 membranes, and the like, may be fixed with 1 or 2 per 

 cent, of 2 per cent, osmic acid solution added to the picrate 

 solution and stained with picro-carmine before putting into 

 the glycerin mixture.) 



Other workers have employed saturated solution of 

 iodine in iodide of potassium (so Aenstein) or picro-carmine 

 (so Feist, Arch. Anat. Entwickel., 1890, p. 116 ; cf. Zeit. 

 wiss. Mik., vii, 1890, p. 231), the latter having the advan- 

 tage of preserving the true blue of the stain if it be 

 not allowed to act too long, and the preparation be mounted 

 in pure glycerin. 



Picric acid has been used by Lavdowskt, but this after 

 careful study is rejected by Dogiel. 



Apathy [op. cit., § 342) brings preparations either into a 

 concentrated aqueous solution of picrate of ammonia free 

 from picric acid, and containing five drops of concentrated 

 ammonia for every 100 c.c. ; or, which is generally prefer- 

 able, into a 1 to 2 per cent, freshly prepared solution 

 of neutral carhonate of ammonia saturated with picrate. 

 They remain in either of these solutions, preferably in the 

 dark, for at least an hour. They are then brought into 

 a small quantity of saturated solution of picrate of ammonia 

 in 50 per cent, glycerin, where they remain until thoroughly 

 saturated. They are then removed into a saturated solution 

 of the picrate in a mixture of 2 parts 50 per cent, glycerin, 

 1 part cold saturated sugar solution, and 1 part similarly 



