*Absolute alcohol.*--Used as a hardening agent where the tissues are to be examined for micro-organisms, and for specimens to be stained by Nissl’s method (p. 101). A cheaper and equally effective hardening medium is made by dehydrating methylated spirit by adding one ounce of fused carbonate of potassium to each pint of methylated spirit, and decanting. Small pieces must be used. The depths of the block should not exceed 3/8 inch. The fluid should be changed on the third day. Hardening will be completed in about ten days or even earlier. *Osmic acid.*--For rapidity of action, and for rapid fixing of all the tissue elements in their natural position osmic acid is one of the best hardening reagents we possess. Its disadvantages as a hardening agent are:-- 1. Its expense. 2. Its irritating and corrosive vapour. 3. The fact that only small pieces of tissue can be hardened in it, since the external surface is very rapidly hardened and thus the fluid is prevented from penetrating into the centre of the lump. It is most frequently used as a hardening agent for very delicate structures, such as the retina, or embryos, or for fresh sections of brain (p. 95). The acid itself may be procured in sealed tubes, each containing one gramme. These should be broken in a bottle under sufficient distilled water to make a one per cent. solution. The bottle containing it should be covered with brown paper to exclude the light. For hardening purposes small pieces of the tissue, not much larger than a pea, should be placed in the acid, the one per cent. solution being diluted with five to ten volumes of distilled water. The tissues may be left in this for from three to five days. They must then be *thoroughly* washed in distilled water and may afterwards be preserved in methylated spirit. Both the hardening and the subsequent washing must be carried on *in the dark*. Osmic acid is also a most valuable staining reagent (see p. 81). *Carbolic acid* (5 per cent.).--May be used to harden almost any tissue, but is particularly useful for hardening nervous tissues such as brain or spinal cord which are afterwards to be preserved as museum specimens. It does not discharge the colour of a specimen so rapidly as spirit. Three or four times the bulk of fluid should be used. It requires changing at the end of twenty-four hours, and again at the end of the first week. Saturated aqueous solution of *corrosive sublimate* is one of the most convenient hardening reagents for small pieces of delicate tissue, _e.g._, embryos. It hardens them in a few days. When they are sufficiently hardened the mercurial salt should be removed by washing first in methylated spirit for a few hours and then in running water.