154 



THE ART OF MAKING MICROCOPE SLIDES 



Pluteus larva 



is in the form of reconstructions. The pres- 

 ent example, since reconstruction has not 

 been previously described, must be pre- 

 fixed by a discussion of this process. 



Reconstruction involves reproduction, 

 either as a side view or as a solid model, of 

 greatly enlarged areas of a section. This 

 book is not the right place to discuss the 

 process in detail, since it is no part of the 

 making of microscope slides. A general un- 

 derstanding of the processes is, however, 

 necessary in order to explain the steps 

 which are taken in preparing the sections. 

 To make a wax reconstruction, camera 

 lucida drawings of the parts of the re- 

 quired sections are transferred to sheets 

 of wax which have been cast of the same 

 thickness as the section would be if it 

 were magnified to the size of the drawings. 

 The relevant portions of the wax are now 

 cut out and piled on top of each other 

 until a solid model, representing an en- 

 largement of the section, has been built. 

 Graphic reconstruction, on the contrarj^, 

 is done on sheets of graph paj^er. Lines 

 of the thickness which correspond to the 

 magnification at which one is studying the 

 section are drawn, these lines represent a 

 side view of the section which is being re- 

 constructed. It must be obvious that in 

 both cases it is necessarj^ to have one fixed, 

 straight line running from front to back 

 of the object in order to relate either the 

 wax blocks or the lines drawn to some 

 stable point. Were this not done, any 

 curve, when reconstructed, would appear 

 as a straight line. In reconstructing a ver- 

 tebrate embryo the problem is simplified 

 because either the center of the notochord, 

 or the dorsal aorta, may be used as a point 

 of reference. Invertebrate larvae, how- 

 ever, suffer from the disadvantage that 

 they rarely have any structure which runs 

 in a straight line through them and some 

 straight lines must be synthesized. This is 

 best done by embedding alongside the ob- 

 ject a hair which has been thickl}' covered 

 with lampblack. After the block has been 

 cast the hair is withdrawn with a sharp 

 jerk (it might really just as well be left in 

 place) leaving a line of lamp black run- 

 ning from front to back. This line will ap- 

 pear as a dot in each successive section and 

 each structure seen in the section may be 



orientated with regard to the lampblack 

 line. 



The first problem in dealing with echino- 

 derm larvae is that of fixation. In the 

 author's experience nothing is to be com- 

 pared, for this purpose, with the "strong 

 fluid" of Flemming (Chapter 18, F 1600.0010 

 Flemming 1884). Larvae from plankton 

 samples or from breeding tanks, are accu- 

 mulated in a small fingerbowl of clean sea 

 water, each one then taken up in a pipet 

 with the smallest possible quantity of sea 

 water, and squirted rapidly into a large 

 volume of fixative. They will be perfectly 

 fixed in about 10 minutes and must im- 

 mediately be removed to distilled water, in 

 several changes of which they are washed. 

 It is necessary to remove the fixative as 

 rapidlj' as possible since the osmic acid is 

 liable to deposit osmium hydroxides as a 

 blackened layer over the tissue. Since 

 these objects are to be embedded in cel- 

 loidin and paraffin, it is necessary for them 

 to be stained before embedding, or end- 

 less difficulties will result. The writer 

 prefers, for echinoderm larvae, Mayer's 

 "paracarmine," the formula for which will 

 be found in Chapter 20 as DS 11.22 Mayer 

 1892a. Fixed specimens should be passed 

 from 70% alcohol to 50% alcohol, which 

 is then replaced with the stain in which 

 the larvae are left from 24 to 48 hours. 



It is easy to lose small larvae in chang- 

 ing the staining solutions. It is recom- 

 mended, therefore, that the tube in which 

 the staining is done be tipped out into a 

 large fingerbowl of the differentiating solu- 

 tion, since the resultant dilution is fight 

 enough in color to enable one to see the 

 small larvae floating about. These larvae 

 are then picked out one at a time and 

 placed in a clean tube of the differentiating 

 solution for about half a day. They are 

 then removed to distilled water for two 

 changes of about one hour each, and from 

 this passed through 30% alcohol, 50% al- 

 cohol, and back to 70% alcohol, in which 

 they are well washed, and in which they 

 may be preserved until required for sec- 

 tioning. It may be added that the cal- 

 careous sjiines contained in later pluteus 

 larvae are dissolved by the acetic acid 

 in the fixative and no decalcification is 

 necessary. 



