136 MR. R. J. TILLYARD ON THE RECTAL BREATHING-APPARATUS 



Microtomy. — For this purpose two methods of procedure are available^ 

 both of which have their own peculiar advantages : — 



1. The oill-basket may be dissected out as described above, and plunged at 

 once into the fixative fluid. In the case of very complicated structures, such 

 as the Duplex System of gill-basket in well -grown larvse, some of the 

 specimens should be dealt with by everting the basket with a longitudinal 

 cut, so that the complicated foldings may be more widely separated. 



2. The larva may be taken alive (preferably, soon after an ecdysis) and 

 plunged into the fixative. By this means the fixative is drawn at once into 

 the rectum by the movements of inspiration, and fixation of the rectum 

 takes place from within. 



The second method offers the advantage of studying the complete structure 

 of the rectum in relation to the surrounding parts and body-wall. By it the 

 very delicate rectal musculature is preserved entire, and the exact positions 

 of the o-ill-e versions can be made out. But, before successful sectioning can 

 be attempted, it is necessary to soften the chitinous integument of the larva^ 

 and this is a difficult task, if one would avoid any damage to the internal 

 organs in consequence. 



The first method enables us to proceed at once with dehydration and 

 embedding, without the use of a softening process ; but it only gives us, 

 complete, the rectal structures enclosed within the circular muscle-layer. 

 Hence it is advisable to obtain plenty of material and to use both methods, 

 keeping the second only for use on larvse which have just undergone ecdysis,. 

 and whose integument, in consequence, can be softened M'ith the minimum 



of trouble. 



Probably many methods of fixing, softening, and staining could be tried 

 with success in the case before us. It seemed, however, advisable, bearing in 

 mind the limited time at my disposal and the very great variety of material 

 to be studied, to select one good method and to treat all the specimens 

 alike. I therefore made use of the following method, previously applied 

 in part by Carl and in full by Kurt-Bedau *, in studying the compound eyes 



of Insects : — 



Fixation. — The fixative used was devised by (krl, and may be called CarFs 



Fixative. It consists o£ 



Absolute alcohol 15 parts. 



Concentrated formol C „ 



Glacial acetic acid . , 2 ,, 



Distilled water 30 „ 



The object should be left in this fixative for 24 hours, and then washed in 

 70 per cent, alcohol for 24 hours with several changes. 



Softening. — If the second method of fixation, as given above, be used, the 



* "Die Facettenauge der Wasserwanzen," Kurt-Bedau, Zeit. wiss. Zool., Bd. 97 (1910). 



