Experiments on Staining Thin-Sections 



121 



20 mm. 



Fig. I. The preparation of formvar nets as supporting material for ullralhin tissue sections, (a) Microscope slide in 2% 

 formvar solution, (b) Microscope slide transferred to vessel in which air is saturated with vapour of solvent for the formvar 

 (ethylendichloride). The slide is left there for 20 min. (< ) Damp air is blown into this vessel, {d) Formvar net floated off 

 onto water surface. 



water the formvar net formed is floated off onto a 

 water surface and transferred to specimen grids in 

 the regular way. The upper two-thirds of the formvar 

 net can be used. 



With this method, it is possible to prepare formvar 

 nets with a large percentage of open area and with 

 a rather uniform and useful size of the holes of the 

 nets. These formvar nets can be examined in the 

 electron microscope and the useful grids selected. 

 Before being used, the nets should be stabilized by 

 coating them with a layer of metal or carbon in a 

 metal shadow casting unit. 



The gain in contrast when mounting the sections 

 in nets is so considerable that thin sections can be 

 examined without any objective aperture. This is of 

 great advantage in high resolution electron micros- 

 copy, as it means an elimination of the most frequent 

 source of astigmatism. 



References 



1. Sjostrand, F. S., Siieiue Tools 2, 25 (1955). 



2. — Exptl. Cell Research 10, 657 (1956). 



Experiments on Staining Thin-Sections for Electron Microscopy 



I. R. Gibbons and J. R. G. Bradfield 



Cavendish Laboratory, Caitihridge 



Ihe development of suitable staining techniques for 

 electron microscopy is important because staining 

 not only increases the observed contrast but also 

 provides a possible means of obtaining cytochemical 

 information about cell components. 



Experimental. — The procedure for staining is 

 simple. Cut a ribbon of sections and pick it up on a 

 filmed electron microscope grid in the usual way. 

 When the sections have dried down onto the film, 

 float the grid section-side downward on the surface 

 of the staining solution for about half an hour. 

 Remove the grid carefully with forceps, rinse by brief 

 immersion in two changes of glass-distilled water 

 and allow to dry. 



The time of staining used has generally been about 

 half an hour, but it does not appear to be critical. 

 Stereo-photographs of the fibres in locust sperm 

 tails in a rather thick section (about 700 A), which 



had been stained in osmium tetroxide for half an 

 hour, have shown the staining to be fairly uniform 

 through the thickness of the section. This suggests 

 that, even for a section as thick as this, penetration 

 is almost complete in this time — in spite of the 

 presence of the methacrylate. 



A small modification of the abo\c procedure en- 

 ables an unstained "reference area" to be obtained. 

 Before staining, the grid bearing the sections is 

 placed in the electron microscope and some areas of 

 the sections are given a brief irradiation with a fairly 

 intense electrcin beam. When the grid is removed 

 from the microscope and stained as before, those 

 areas which were pre-irradiated appear to be unaf- 

 fected by the staining procedure and hence serve as 

 reference areas in the sections. Comparison with 

 these unstained reference areas enables one to assess 

 the intensity of staining in the rest of the section. A 



