634 APPENDIX. 



should be governed by their size and the character of the tissue. When infiltration is com- 

 plete, place the specimen with a considerable amount of the thick celloidin on a block of wood 

 or of vulcanized fiber, allow to stand in the air until a firm skin forms, and then immerse 

 in 80 per cent, alcohol. This hardens the celloidin in several hours and attaches the speci- 

 men firmly to the block. The celloidin may be hardened and the specimen fixed to the 

 block more quickly by immersing in chloroform. The specimen is then ready for cutting. 

 Blocked objects may remain in the alcohol several months. A better fluid, however, is strong 

 alcohol and glycerin in equal parts; in this the celloidin does not become soft, as it does after 

 several months in plain alcohol. 



Paraffin embedding. Paraffin with a melting point of 50 to 55 C. should be used, and 

 the paraffin oven kept at a temperature of about 56° C. The hardened tissue is placed in 95 

 per cent, alcohol for several hours and then in absolute alcohol for the same time. It is 

 next transferred to oil of cedarwood for 24 hours (or xylol for 6 hours, or chloroform for 6 

 hours), then into melted paraffin in the oven for from 1 to 6 hours, depending upon the size 

 of the specimen. The paraffin should be changed twice. Make a paper box that will 

 considerably more than contain the specimen, fill the box with melted paraffin, and drop the 

 specimen into it. When the paraffin Jias become cool, so that a skin forms on the surface, put _ 

 it in cold water (ice-water if available) until the paraffin is hard. These blocked specimens 

 can be kept in the air indefinitely. For section cutting the block is attached to the metallic 

 block-holder by heating the latter and pressing the paraffin down upon it. 



VI. Section Cutting. 



In cutting celloidin sections the microtome knife is adjusted so that it passes obliquely 

 through the specimen and must be kept flooded with 80 per cent, alcohol. The sections are 

 removed from the knife with a camel's-hair brush and placed in 80 per cent, alcohol where 

 they may remain several weeks if desired. Equal parts strong alcohol and glycerin will 

 preserve the sections indefinitely. 



Celloidin sections are usually not cut thinner than 8 or 10 microns. When thinner 

 sections are desired, or when large or brittle specimens are being cut, it is often advantageous 

 to paint the surface of the block, after cutting each section, with a coat of very thin celloidin. 

 The surface of the block should be dry before applying the celloidin. 



In cutting paraffin sections, the block containing the specimen is attached to the block- 

 holder, with proper orientation, and then trimmed so that opposite surfaces are parallel. 

 The knife should be used dry and is passed straight through the specimen. The sections 

 are removed with a camel's-hair brush and laid upon a sheet of paper, where they may be 

 kept for several days at room temperature. They must be kept in some protected place, 

 for a slight draft will blow them away. Paraffin sections can usually be cut thinner than 

 celloidin; under favorable circumstances they can be cut even 2 or 3 microns in thickness. 



Serial sections undoubtedly cut to best advantage in paraffin. Sections of embryos up to 

 10 or 12 mm. should be cut 10 microns thick; sections of larger embryos, 15 or 20 microns 

 thick. The edges of successive sections adhere, and with care long "ribbons" can be ob- 

 tained. These are arranged in order on a piece of paper, and afterward mounted in order. 

 If the edges of the sections do not adhere, or if the sections curl, it usually means that the 

 paraffin is too cold, or that the room temperature is too low, or both. This trouble can some- 

 times be avoided by dipping the block in melted paraffin of a lower melting point, or by 

 keeping a gas flame near the microtome. If many sections are to be cut, however, a room 

 temperature of 70° to 73 F. will be found more satisfactory. 



Care should be taken to properly orient embryos that are to be cut serially. As a rule. 



