RADIOAUTOGRAPHY 



299 



RADIOAUTOGRAPHY 



Large specimens of soft tissues may 

 be frozen, ground in the same way as 

 above, or sectioned with a chilled blade 

 or fine electric saw while being kept 

 in the frozen state, and thus applied 

 to the photographic emulsion. 



1. Contact method: This technique, 

 the oldest and crudest method, has been 

 used extensively especially in the in- 

 vestigations of the sites of localization 

 of isotopes in hard tissues such as bones 

 and teeth, and in frozen sections of soft 

 tissues. (See reviews: Gross, J. and 

 Leblond, C. P., Canadian Med. Assoc. 

 J., 1947, 57, 102; Gross, J., Bogoroch, R., 

 Nadler, N. J. and Leblond, C. P., Am. 

 J. Roentg. and Radium Therap., 1951, 

 65, 420-458.) The method consists es- 

 sentially of placing a microscopic slide 

 bearing a radioactive tissue in close con- 

 tact with a photographic plate; and 

 of keeping these in close contact by 

 pressure. 



1. A slide containing the histological 

 section is dipped twice into an ether- 

 alcohol solution of 1% celloidin fol- 

 lowing staining and passage through 

 95% and absolute alcohol. The sec- 

 tion is then dried overnight to in- 

 sure hardening of the celloidin. 



2. In the darkroom, the histological 

 slide, which should be free from dust 

 particles or granules which may 

 make for uneven contact, is placed 

 gently against a photographic plate 

 or film (x-ray, Eastman Kodak 

 Medium Lantern Slide plate, etc.). 

 Various methods may be used to 

 insure an intimate uniform contact 

 between slide and plate or film. 

 They may be held in a roentgen-ray 

 type pressure cassette or in a print- 

 ing frame. Naturally, these holders 

 should be light-tight. 



3. After exposure, the photographic 

 plate or film is processed according to 

 the routine photographic procedures, 

 i.e., developed in D-72 for 2 min., 

 rinsed in water, fixed in acid fixer 

 for 10 min., washed for 30 min. and 

 dried. To protect the radioauto- 

 graphic image in the emulsion from 

 scratches or abrasions, it is useful 

 to cover the image with a drop of 

 Canada balsam and a histological 

 coverslip. The histological section 

 and the radioautographic image may 

 then be simultaneously examined 

 under a dissection or ordinary micro- 

 scope. 



This method has the advantage of 

 simplicity but the localization at best 

 is rather crude due to low resolution. 



2. Mounting method: The one most 

 usually followed is that of floating sec- 

 tions onto a photographic emulsion. 



Recent attempts at doing this under 



dry conditions will be mentioned later. 



In the Wet Mounting method (Evans, 



T. C, Proc. Soc. Exp. Biol, and Med., 



1947, 64, 313-315; Endicott, K. M., and 

 Yagoda, H., Thid.j p. 170-172; Boyd, 

 G. A., and Williams, A. I., Ibid., 



1948, 69, 225-232) the tissue section is 

 floated directly onto a photographic 

 film or plate. This insures a more 

 intimate contact between tissue and 

 emulsion and results in a better resolu- 

 tion, i.e., permits a finer localization. 

 This method, however, cannot be used 

 if the radioactive substance in the 

 tissue is water soluble. 



1. Unstained strips of paraffin sections 

 of radioactive tissues are floated in 

 a 40°C. water bath to remove any 

 wrinkles from the tissue. The water 

 is cooled to 18°C. by the addition of 

 ice cubes at 18°C. and all subsequent 

 steps are carried out in the darkroom 

 using a Wratten No. 1 safelight. 



2. A photographic plate or film (East- 

 man Kodak Medium Lantern Slide 

 Plate, NTA, NTB 1, 2, or 3 plates, 

 etc.) is slipped under the tissue sec- 

 tion, the corner of the tissue being 

 held against the photographic plate 

 with a needle, and the plate with ad- 

 herent tissue is lifted out of the 

 water. After the excess water has 

 drained off the plate and the emul- 

 sion has dried completely, the tissue 

 adheres intimately and permanently 

 to the photographic emulsion. 



3. After proper exposure at refrigerated 

 temperatures (4°C. or lower), the 

 section is deparaffinated in two 

 changes of xylol and hydrated 

 through graded alcohols. 



4. The preparations may be developed 

 in D-72 developer for 2 minutes or in 

 D-19 for 10 minutes, rinsed in water, 

 fixed in acid fixer F5 or in 30% thio- 

 sulphate, and washed in running 

 water for 30 minutes. 



5. Unstained sections may be dehy- 

 drated, cleared and mounted im- 

 mediately or they may be stained 

 with either dilute hematoxylin over- 

 night, or metanil yellow (Gross, J. 

 and Leblond, C. P., Canadian Med. 

 Assoc. J., 1947, 57, 102-122; Simmel, 

 E. B., Fitzgerald, P. J. and Godwin, 

 J. T., Stain Techn., 1951, 26, 25-28). 

 For staining prior to development, 

 Doniach and Pelc have used hot 

 carbol fuchsin-neutral red which re- 

 sists the destaining action of the 

 developing reagents. 



Because the mounting method allows 

 for a very intimate contact between 

 tissue section and emulsion, a very 

 very fine localization is possible. 



