METHYLEN BLUE. 205 



to four Lours before killing them. S. Mayer (Zeit. n-isn. 

 Mik., vi, 1889, p. 423) took a strength of 1 : 300 or 400 of 

 0"5 per cent, salt solution. The solutions of Rbtzius are 

 of the same strength. But the tendency of more recent 

 practice is decidedly towards the employment of weaker 

 solutions. Apathy (ibid., ix, 1892, pp. 2-5, 26 et seq.) finds 

 that it is not only superfluous, but positively disadvantageous, 

 to take solutions stronger than 1 : 1000, DoaiEL (Encycl. 

 Mik. Technili., 1st ed., p. 815) recommends i to ^ per cent., 

 or at most \ per cent. For warm-blooded animals the solution 

 should be wai-med to 36° or 37° C, and before sending in 

 the injection the blood-vessels should be well washed out 

 with similarly warmed salt solution. The injected organs 

 may be removed after 20 to 30 minutes. They should be 

 placed on a thin layer of spun glass moistened with weak 

 (I to ^ig- per cent.) methylen blue, or simply spread out on a 

 slide, and the whole placed in a Petri dish with a layer of 

 the methylen blue on the bottom. The dish is best placed 

 in a stove at 36° C, and after 15 to 30 minutes (if the pieces 

 are thin) or 1 hour to 1^ hours (if they are thick) specimens 

 may be removed for examination or preservation ; or, without 

 using the stove, specimens may be removed 10 to 20 minutes 

 after injection, placed on a slide, and moistened with weak 

 methylen blue or salt solution, and brought under the 

 microscope. Then as soon as the stain is sufficiently brought 

 out (40 to 60 minutes) they may be fixed (§ 343). 



For staining iy immersion the solutions should, if anything, 

 be still weaker. Dogiel (^Arcli. mik. Anat., xxxv, 1890, p. 

 305) places objects in a few drops of aqueous or vitreous 

 humour, to which are added two or three drops of a ye ^'^ iV 

 per cent, solution of methylen blue in physiological (0*75 per 

 cent.) salt solution, and exposes them therein to the air. In 

 thin pieces of tissues the stain begins to take effect in five or 

 ten minutes, and attains its maximum in from fifteen to 

 twenty minutes. For thicker specimens — retina, for instance 

 — several hours may be necessary. The reaction is quickened 

 by putting the preparations into a stove kept at 30° to 35° 0. 

 EouGET {Gompt. Rend., 1893, p. 802) employed a 0'05 per 

 cent, solution in 0'6 per cent, solution (for muscles of Batra- 

 chia). Allen (Quart. Journ. Micr. 8ci., 1894, pp. 461, 483) 

 takes for embryos of the lobster a solution of O'l per cent, in 



