328 RADIOISOTOPES IN BIOLOGY AND AGRICULTURE 



min at a collection efficiency of about 10 per cent, this would indicate that 

 a 10- to 15-day exposure period would be required. This type of measure- 

 ment may be misleading in the prediction of inadequate exposure, should 

 the radioactivity be .limited to a small area. Fitzgerald et al. (10) have 

 calculated that the following approximate tissue concentrations are 

 required for histological sections of 10 n to give an image on X-ray film 

 with a 15-day exposure: C^\ 0.05 /xc/g; Ca^^ 0.05 Mc/g; P^S 0.2 ^c/g; 

 P^-, 0.4 /xc/g; Zn^^, 2.0 Mc/g. If the percentage of uptake in a given 

 tissue can be estimated from tracer experiments, then the above values 

 wdll permit an estimation of the order of magnitude of the dose required. 



In practice, with plants and animals it is usually feasible to run some 

 preliminary experiments with graded levels of activity. This permits a 

 more accurate estimation of the dose required, as well as an indication of 

 the optimum exposure time. It is important to avoid overexposure, since 

 this causes a loss of resolution. Under standard experimental conditions 

 the optimum exposure time may be estimated from tissue counts by pre- 

 vious determination of the relationship between tissue counts and image 

 densities. However, it is often practicable to expose several sections; 

 after one or two have been developed at different times, usually being 

 underexposed, it is then possible to estimate fairly closely the best expo- 

 sure time. 



Handling of Tissues. For gross autoradiograms, thin sections offer 

 no advantage, and the problem is primarily that of presenting a smooth 

 surface to the emulsion and maintaining a close contact by pressure. For 

 studies at the cellular level it is necessary to employ thin sections of the 

 order of 5 to 10 m or less, and the conventional histological procedures are 

 heavily drawn upon for the preparation of such sections. The reader 

 unfamiliar with the basic methods of fixation, dehydration, embed- 

 ding, and sectioning should consult references (1 to 3) for background 

 information. 



In addition to the histological requirements of the fixatives and other 

 agents, it is most important that these materials do not inhibit or increase 

 photographic action and do not leach the radioisotopes from the tissue. 

 Boyd and Board (12) demonstrated the production of artifacts by chem- 

 ical action of various soft tissues on the photographic emulsion, and 

 Everett and Simmons (13) have emphasized that even the protective 

 coatings such as Formvar or silicone do not provide absolute elimination 

 of chemical reduction of the emulsion. The leaching problem is a serious 

 one, as may be noted from the papers of Williams (14), Holt and Warren 

 (15), and Blank et al. (16). It is generally agreed that mercury-contain- 

 ing fixatives such as Zenker's cannot be used. There seems to be some 

 question as to whether neutral Formalin is satisfactory (10, 17). Alcohol 

 has been used satisfactorily from the leaching standpoint but was not 



