APPENDIX. 363 



acid, and to 100 c.c. of this add 2 c.c. of concentrated 

 sulphuric acid; filter, and to the filtrate add 300 c.c. 

 of distilled water (Kleinenberg). 



Tissues should be left in this for a comparatively short 

 time, varying from three hours to a few days; the 

 hardening may be completed with alcohol. If the 

 tissue contains earthy salts to be extracted, nitric 

 acid should be substituted for sulphuric acid. 



A simple concentrated aqueous solution is also fre- 

 quently usef uL 



Osmic acid, a 1 p.o. solution of this is most useful. 

 To avoid reduction of the osmic acid care should be 

 taken to obtain water free from organic matter. 

 The acid is obtained in tubes containing one grm. 

 each, the tube should be well washed and then broken 

 in a bottle containing 100 c.c. of distilled water and 

 shaken occasionally; it takes some little time to 

 dissolve completely. The bottle should have been 

 previously well washed with sodium hydrate, sul- 

 phuric acid and water. If a solution is required to 

 be made at once, the tube with the osmic acid may be 

 ground up in a mortar with a little water, but some 

 care is required to do this without sufiering from the 

 hurtful vapour. If the solution begins to turn brown, 

 the bottle should be covered with black paper and 

 kept in the dark; this however should not be necessary. 

 Osmic acid does not penetrate well, so that a small 

 piece only of the tissue should be taken ; the tissue 

 may be left in the osmic acid any time from half-an- 

 hour to a day, in the latter case a -5 p.c. solution should 

 be used. Many tissues are hardened sufficiently by 

 a stay of 12 to 24 hours in osmic acid' to allow of 



