1903] KELLOGG: — STUDIES FOR STUDEXTS II9 



in which the specimens may be preserved indetinitely. Or, transfer the specimens 

 from the boiUng water to a warm concentrated sokition of corrosive sublimate in 

 35 C{, alcohol for three hours (puncturing the body wall just before removing to this 

 solution) ; then wash in 75% alcohol ; then transfer to 759^ alcohol to which a few 

 drops of tincture of iodine has been added, to extract the corrosive sublimate, then 

 wash in clear 75% alcohol and transfer to 85*^ alcohol iox keeping. Do not use 

 metal instruments in handling material fixed in corrosive sublimate. Put specimens 

 in at least ten times their own bulk of the various solutions used. Keep in corked 

 shell vials or large homeo vials. 



For more detailed account of killing and fixing methods for insects, with refer- 

 ence to other fixing agents and special cases, see Comstock and Kellogg's Elements 

 of insect anatomy, chap, VIII (p. 121-139), 1901 ; for exhaustive account of 

 many killing and fixing agents (for miscellaneous animals) see Lee's Microtomists' 

 vade-mecum. Also see these two references for more detailed consideration of 

 the subjects of the following paragraphs. 



Hardening, dehydrating, and clearing. — When ready to carry material further, 

 select from the stock of properly killed and fixed specimens (preserved in 85% 

 alcohol) the particular specimens desired to study and transfer to 95% alcohol for 

 from 12 to 24 hours ; then to absolute alcohol for from 12 to 24 hours; then to a 

 half-and-half mixture of absolute alcohol and cedar-wood oil (or xylol). Pour the 

 oil slowly into the vial containing the specimens in absolute alcohol ; the oil and 

 alcohol will remain distinct at first, the specimens keeping in the alcohol ; as the 

 two liquids gradually mix the specimens will become gradually (and hence safely) 

 infiltrated with the new mixture. Leave in this mixture for from 12 to 24 hours. 

 Transfer to pure cedar-wood oil (or xylol); leave from 12 to 24 hours. The speci- 

 mens are now ready to be infiltrated with and imbedded in paraifine preparatory 

 to cutting by the microtome. 



Infiltrating and imbedding. — Remove specimens from pure cedar-wood oil, in 

 which they may remain without injury indefinitely if for any reason the work must 

 be interrupted, into cedar-wood oil into which about half the same bulk of parafiine 

 shavings have been dropped and allowed to dissolve. This mixture should be kept 

 in a watch glass or small dish at a temperature of about 45° C. To do this keep 

 the dish in the parafiine oven or at the back end of an imbedding triangle. (Paraf- 

 fine oven or imbedding triangle may be obtained of dealers in microscopic supplies.) 

 Remove specimens from mixture of parafiine and cedar-wood oil after from three 

 to six hours, depending on size and thickness of body wall of specimens, to melted 

 pure parafiine of 54° C. melting point. This paraffine must be kept melted in paraf- 

 fine oven or on imbedding triangle. The temperature should not be allowed to go 

 up much higher than the melting point of the paraffine and never to fall below it 



