SEMEN STAINS 



222 



SHADOW-CASTING 



scrape surface of cloth with blunt edge 

 of a scalpel. Carry scrapings off with 

 fluid anci spread on a slide. Dry and 

 fix with heat. Cover with 4 cc. 1% aq. 

 Wollschwartz (Grubler) + 0.05 cc. 2% 

 aq. sulphuric acid, 5 min. Wash in 

 water. Counterstain 6-8 sec. with Loef- 

 fler's methylene blue diluted with 15 

 parts aq. dest. Wash in aq. dest., dry 

 and examine. Heads of spermatozoa 

 bright golden or yellowish color, all else 

 gray. Useful in legal medicine (Wil- 

 liams, W. W., J. Lab. & Clin. Med., 

 1936-37, 22, 1173-1175). See author's 

 figures. See Pollak, O. J., Arch. Path., 

 1943, 35, 140-196. 



Seminal Fluid. To study in sections 

 centrifuge fluid 5 to 1 hr. after ejacula- 

 tion for 20 min. at 3000 r.p.m. Fix 

 centrifugate in 4% formalin, 48 hrs. 

 2 changes. Take sediment into abs. 

 ale, then 9 parts abs. and 1 part xylol. 

 Gradually increase xylol to 9 parts to 

 1 part ale. Xylol paraffin 30 min. 

 Then 54 °C. melting paraffin for 3 hrs. 

 in incubator at 58°C. After 3 hrs. in 

 60°C. melting paraffin embed and sec- 

 tion 2-3 microns thick (Joel, K., J, 

 Lab. & Clin. Med., 1939, 24, 970-972). 



Sense Organs, see Eyes, Ear, Pacinian 

 Corpuscles, Meissner's Corpuscles, 

 Krause's End Bulbs, Nerve Endings. 



Sensitol Red, see Pinacyanol. 



Serum Agar, see Bacteria, Media. 



Setoglaucine O (CI, 658), a basic dye less 

 light fast than Malachite green (CI, 

 657), a constituent of some bacterio- 

 logical media (Emig, p. 47). 



Sharpening, see Microtome Knife. 



Shrinkage caused by fixation, dehydration 

 and clearing of nervous tissues has been 

 measured by King, H. D., Anat. Rec, 

 1910, 4, 213-244 and by Allen, Ezra, 

 Anat. Rec, 1916, 10, 565-589. 



Shadow-casting. — Written by Dr. W. T. 

 Dempster, Dept. of Anatomy, Univer- 

 sity of Michigan, Ann Arbor, Mich., 

 May 28, 1946 — This is a technique for 

 revealing the surface form and texture 

 of microscopical material in either light 

 or electron microscopy. It is an out- 

 growth of R. C. Williams' experience 

 with vacuum deposited metal films on 

 astronomical mirrors and of studies on 

 the physics of metallic films. Metal 

 evaporated from a hot filament in a 

 high vacuum is of atomic dimensions. 

 It was found that the particles travelled 

 in straight line paths from the filament 

 and that obstructions, however small, 

 cast a metal-free shadow. With R. W. 

 G. Wyckoff (J. Appl. Phys., 1944, 15, 

 712-716), a successful technique of 

 oblique casting of metal films was ap- 

 plied to electron microscopy for measur- 

 ing shadow lengths and calculating 

 heights of minute objects. Bacteria, 



viruses and minute chemical aggregates 

 have been studied (Williams, R. C. and 

 Wyckoff, R. W. G., Proc. Soc. Exp. 

 Biol. & Med., 1945, 58, 265-270; 59, 265- 

 270; Science, 1945, 101, 594-596; 102, 

 277-278; Nature, 156, 68). Unusual 

 contrast, surface texture and the possi- 

 bility of measuring heights are positive 

 advantages of the technique. Further 

 applications to both electron and visual 

 microscopy involved a method of study- 

 ing opaque surfaces by coUoidin replicas 

 that are shadowed (Williams, R. C, 

 and Wyckoff, R. W. G., J. Appl. Piiys., 

 1946, 17, 23-33). Applications of the 

 method to biological material viewed 

 with the light microscope and an ac- 

 count of the casting apparatus (W. T. 

 Dempster and R. C. Williams) are 

 forthcoming. 



Material is affixed to cover slips; 

 smears are thoroughly dried; paraffin is 

 dissolved from sections with solvents. 

 With no further preparation, other than 

 thorough drying, the slips are shadowed 

 with a metal deposit in a vacuum cham- 

 ber. After this, they are mounted face 

 down on slides with clarite or balsam. 

 For electron microscopy, regular screen 

 grids with a thin coUoidin film over the 

 mesh are used as substrates for suspen- 

 sions; replicas are placed directly on the 

 mesh. Metallic chromium is about the 

 best metal for general shadowing but, 

 for finest detail with the electron micro- 

 scope, gold or uranium is preferable. 

 The casting technique is similar for the 

 different metals. Shadow-casting pro- 

 duces a visually structureless deposit 

 which sticks to all surfaces save those 

 directed away from the hot filament and 

 shadow areas due to obstructions. Sur- 

 faces perpendicular to straight line 

 paths from the filament get the heaviest 

 deposit; oblique surfaces get less and 

 shadows none. Metal deposited at a 

 rather oblique angle has a distribution 

 much like light from a point source 

 shining obliquely on three-dimensional 

 objects. Highlights and shadows are 

 produced. Through the microscope, 

 shadows in the preparations transmit 

 light and appear bright; highlights are 

 dark. The eye, however, readily 

 adapts to this reversal of tone. Photo- 

 graphic negatives or prints made from 

 glass positives reverse the microscope 

 appearance; highlights then are bright, 

 shadows are dark, and variations of 

 surface texture are shown by gradations 

 of tone. 



Electron-microscope negatives show 

 the same natural appearance of light 

 and dark. The observer looks at sur- 

 faces rather than through specimens as 

 in ordinary microscopy. The appa- 

 ratus consists of a bell jar and a base 



