METHODS OF RESEARCH: PREPARATION OF SECTIONS 115 



sections on the microtome. The tissues to be shced may be 

 frozen and cut on a special machine, and this is the quickest 

 way and the best way for preHminary studies, but it is of no 

 value for permanent sections of tissues containing bacteria, 

 since they diffuse out too readily, or they may be embedded in 

 celloidin or paraffin and cut on the ordinary microtome, which 

 is the best way for permanent mounts. I will Hmit my account 

 to paraffin, describing the way it is done in our laboratory. 

 First the tissues are fixed. We select for this purpose, clean, 

 perfectly fresh, characteristic bits which should not be large. 

 Various fixing agents are used; we often use Carnoy's fixative 

 (H glacial acetic acid, % absolute alcohol). They will fix 

 better in many cases if the air is pumped out of them as soon 

 as they are put into the fixative. I make this an invariable 

 rule. After 24 hours in this solution (12 hours is enough in 

 some cases) the pieces are shifted into 90 or 95 per cent alcohol, 

 the next day into a second alcohol, and finally from absolute 

 alcohol into mixtures of alcohol and xylol, then into pure xylol, 

 a second pure xylol, to remove all traces of the alcohol, then 

 into xylol-paraffin, then paraffin plus xylol, and finally into 

 pure melted paraffin, from which after some hours they are trans- 

 ferred to another melted paraffin in which they are embedded. 

 There is some choice in the melting point of paraffin to be used for 

 the final embedding depending on the climate. In Washington 

 we use during the summer Griibler's paraffin melting at 56° to 

 58°C., and during the winter that melting at 52°C. The em- 

 bedded pieces are trimmed and stuck to one end of a small rec- 

 tangular white pine block by means of a hot needle and a little 

 melted paraffin. The number of the specimen, corresponding 

 to an account in a record book, is written on one or more sides 

 of the pine block with a lead pencil. The wooden block is now 

 clamped into the microtome. If the material is soft, ribbon 

 sections are cut; if hard, slant-stroke sections are cut. If the 

 knife is dull, or if the tissue is gritty or contains crystals, or is 

 imperfectly embedded, the sections will be torn and worthless, 

 or nearly so. Occasionally when the tissues are full of calcium 

 oxalate crystals we have to be content with torn sections, but 

 ordinarily the tissue should be reembedded and a sharper knife 



