98 INVERTEBRATE ZOOLOGY. 



collect a bottle full of such specimens (8-10) to serve as a siapply. For inter- 

 nal anatomy they should be dissected in a fresh state, also prepared for the 

 microtome and sectioned in various planes. As with insects, their hard chit- 

 inous exo-skeleton forms a barrier to the penetration of the various preserva- 

 tives, to overcome which, various means have been adopted, as follows : — 



1. The use of hot fixing reagents. These are to be heated in a test tube 



and poured suddenly over the animal while alive. Perhaps the best 

 for this purpose is hot alcoholic corrosive sublimate. This should 

 remain 20-40 min., then washed out in 70^. 



2. The selection of specimens which have just moulted and in which the 



chitin is soft and tender. These are also best preserved with hot 

 reagents, 



3. Punch or prick a hole through the exo-skeleton in a region not after- 



wards to be used for sectioning. A living animal may be cut in two 

 with a sharp razor and the two parts instantly dropped into the 

 fixative. 



As in the case of preservation, staining also presents serious difficulties. 

 To overcome these, use an alcoholic stain of a high grade (Borax Carmine 

 may be made as high as 70 %) ; also continue the immersion for a long time 

 (several days or weeks if necessary). This will not hurt the specimen if the 

 stain contains a high percentage of alcohol. Sections may be cut from an un- 

 stained object and stained on the slide, by the method known as " slide stain- 

 ing." For this, cover the slide with an albumen fixative, rubbing it in 

 well with the finger. After this, heat in the usual way, until the paraffine is 

 melted, and then apply the usual reagents in the reverse order (4-5 minutes 

 each), turpentine — 100 '/c — 95 % — 70 </(, etc., until the specimen reaches the grade 

 of the desired stain. Apply the stain to the object and then pass it up again 

 through the same succession as far as the turpentine. Then wipe the slide 

 carefully, except the area covered by the specimen, which may be well drained 

 but must not become dry. Add a drop of Balsam and cover as usual. The 

 application of the different reagents is best performed by immersing the en- 

 tire slide successively in jars containing the desired reagents, or in the case 

 of single sections, the reagents may be pipetted on the slide, drop by drop, 

 and successively wiped away, held in the hand or rested from time to time on 

 some convenient sujjport. All the different stains may be applied in this way, 

 haematoxylin being especially recommended. 



