Pluteus larva 



SECTIONS FROM DOURLE-EMBEDDED MATERIAL 



155 



The specimens should now be looked 

 over in order to select those in which the 

 "arms" are relatively straight, and these 

 should then be transfei'retl to a tube of 

 absolute alcohol and changed at intervals 

 until they are completely dehydrated. The 

 absolute alcohol is then replaced withamix- 

 ture of equal parts of absolute alcohol and 

 anhydrous ether, which is changed once 

 or twice. About three hours between 

 changes will be enough to dehydrate ol:)- 

 jects of this size. 



The embedding solution is prepared by 

 taking 16% stock solution of celloidin in 

 alcohol-ether and adjusting it with addi- 

 tions of alcohol-ether and methyl sali- 

 cylate to the composition selected. The 

 solution of Heinz 1923, for example 

 (Chapter 17, 21.1 Heinz 1923), with 

 which the writer has had excellent results, 

 is obtained by diluting 12 milliUters of the 

 16% stock solution of celloidin to a total 

 volume of 50 with a mixture of ecjual parts 

 absolute alcohol and anhydrous ether, and 

 then mixing with this 50 milliliters of 

 methyl salicylate. The larvae are trans- 

 ferred directly from the alcohol-ether so- 

 lution to this mixture and may remain for 

 as long as required, but at least for 48 

 hours. 



The author prefers, at this stage, to add 

 a drop or two of a saturated solution of 

 ethyl eosin in absolute alcohol to the 

 medium, in order that the block cast from 

 it may be sufhciently colored to be visible 

 when embedded in wax. After the larvae 

 have remained in this mixture long enough 

 to become impregnated, the tube contain- 

 ing them is tipped into a watch glass. Each 

 individual larva is then taken uj) in a 

 pipet with a consideralile amount of its 

 embedding medium, and the pipet is held 

 vertically until the larva has sunk to the 

 bottom. The largest possible drop con- 

 taining the larva is then extruded from the 

 pipet into a beaker of anhydrous chloro- 

 form and finally shaken off the tip, so that 

 a spherical globule of the embedding me- 

 dium containing the larva falls into the 

 chloroform, where it coagulates instantly. 

 This is repeated with each of the succes- 

 sive larvae until the batch is finislied. If 

 orientation were not of importance, it 

 would now be possible to proceed to paraf- 



fin embedding. In a case like this, how- 

 ever, orientation is of primary imjwrtance, 

 and the author prefers to transfer, after 

 about an houi' in chloroform, each of the 

 little globules to a tube of cedar oil where 

 they remain until they are clear. Each glo- 

 bule is then trinnned, under cedar oil, until 

 the larva lies in a rectangular block of 

 celloidin with the long axis of the larva 

 exactly parallel to the long sides of the 

 rectangular block. 



It is now necessary to prepare the hairs 

 which will be used to jirovide the guide 

 lines used in reconstruction. Take a num- 

 ber of human hairs and dip them into any 

 sticky substance before rolling them back- 

 ward and forward in a watch glass of 

 lamjiblack. Remove each hair, shake off 

 as much lampblack as possible, and lay the 

 hair aside until required. When it is re- 

 quired for use, a bow of wood, or plastic, is 

 strung with the hair, which therefore re- 

 mains straight and under tension. 



Now return the celloidin blocks to chlo- 

 roform where they remain until most of 

 the cedar oil has been removed. Then 

 change to fresh chloroform and add 

 enough chips of the embedding medium to 

 cover the blocks. Now place the tube in the 

 oven until the wax has dissolved in the 

 chloroform. Pour off the chloroform-wax 

 mixture and replace it with fresh wax, in 

 which the objects msiy continue to impreg- 

 nate for another hour before being cast 

 into blocks. 



Objects as small as this may be con- 

 veniently cast in a watch glass. Place a 

 pipet full of wax in a watch glass, hold 

 this touching the surface of a fingerbowl 

 of water, and, ag soon as the underside has 

 haixlened, transfer a celloidin block to it 

 and orient the block roughly. Now lay one 

 of the stretched hairs alongside the celloi- 

 din block, not quite touching, but abso- 

 lutely parallel. Return the watch glass to 

 the surface of the water so that the block 

 will harden. Then detach the block from 

 the watch glass and tiim it, first cutting 

 away the ends of the hair from the bow, 

 and then jerking out the hair to leave a 

 straight line of lampblack behind it. The 

 block is then mounted and cut into serial 

 sections as in Chapter 12. 



The only difficult}^ which is Ukely to be 



