METHYLEN BLUE. 85 



with picrate of ammonia. More recently (Zeit. f. wiss. Mik., 

 viii, 1, 1891, p. 15) lie has recommended an increased duration 

 of the picrate of ammonia bath up to eighteen or twenty-four 

 hours, and mounting, without washing out, in chemically pure 

 glycerin, free from acid. There is a defect in this process, 

 namely, that picrate of ammonia has a very injurious action 

 of a macerating nature on some tissues. This may, however, 

 be avoided by adding to the fixing-bath 1 to 2 per cent, of a 

 1 per cent, osmic acid solution. (If it be desired to harden 

 the tissues so that sections may be cut, the proportion of 

 osmium solution should be increased fourfold.) 



S. MAYER (Zeit. f. wiss. Mile., vi, 4, 1889, p. 422) preferred 

 a mixture of equal parts of glycerin and saturated picrate of 

 ammonia solution, which served to fix the colour and mount 

 the preparations in. This was also in principle the method 

 followed by RETZIUS (Intern. Monatsschr. Anat. u. Phys., Bd. 

 vii, H. 8, 1890). DOGIEL, after careful study, quite refuses to 

 admit that this is in any way an improvement. 



Other workers have employed saturated solution of iodine 

 in iodide of potassium (so ARNSTEIN), or picro-carmine (so 

 FEIST, Arch. f. Anat. u, Entwickel., 1890, p. 116; cf. Zeit. f. 

 wiss. Mik., vii, 2, 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 LAVDOWSKY, but this, too, after 

 careful study, is rejected by DOGIEL. 



APATHY (Zeit.f. wiss. Mik., ix, 1, 1892, p. 30) has found, as 

 stated above, that free ammonia is a capital factor in the 

 differentiation of the stain. He brings preparations (after 

 washing in salt solution if the staining have been performed 

 with a strong methylen-blue solution, or without washing if 

 it have been done with a very dilute solution) 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 preferable, 

 into a 1 to 2 per cent, freshly prepared solution of neutral 

 carbonate 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. 



