Calibration of stripping film autoradiography in sections of rat liver labelled with tritium

Abstract

Four different doses of 3H- l-tyrosine were injected into the peritoneal cavity of 1-week-old rats, which were killed after 2 hr. Central parts of their livers were removed, fixed, embedded in paraffin, cut into sections of 1 to 10 μ thickness, and mounted on glass slides. The sections were coated with stripping film from Ilford nuclear research plates K-5, and exposed in dry air at 4 °C for periods varying from 1 to 17 days. After processing a 15 °C in complete darkness, the fixed films were rinsed, the underlying sections stained with hematoxylin-eosin, and mounted in balsam. The developed grains were counted over known areas of the sections, as well as over inactive sections mounted on the same slides, and the total areas of the sections were measured. From these data and from the time of exposure the net number of developable grains appearing in the emulsion per unit exposure time was computed. The glass slides carrying the sections were then crushed into appropriate reaction ampoules, the tritium in them was converted into gas, and its absolute activity was determined in metal cathode internal counters. The ratio of the total number of developable grains over a section to the total number of distintegrations in that section, multiplied by 100, was taken as the autoradiographic efficiency expressed in per cent. The relative grain yield was found to decrease with time; the nonlinearity in this relationship was evident, despite the random variation in the results, from the 11th day onward. Actually, the efficiency of grain production seemed to decrease, rather linearly and largely independently of both section thickness and amount of radioactivity in it, as the time of exposure increased from 1 to 17 days. Possible reasons for this are discussed with reference to earlier results, and “coincidence saturation” is excluded, while latent image regression and “charge backscattering” remain as potential explanations. The efficiency of grain production after 7 days of exposure declined in a non-linear fashion from about 14 per cent in sections of 1 μ thickness to about 2.5 per cent in 10 μ sections. The grain yields in sections of 5 and 10 μ thickness were approximately equal. The results are compared to those of previous authors, and backscattering, not considered by them, is taken into account in a rough calculation of the sensitivity of the stripping film on the basis of available data. The effect of an electrical charge on a surface on its backscattering properties, in addition to the atomic number of the material, is suggested as one possible explanation for the difference between the sensitivity obtained for the stripping film (70 per cent) and that given for the equivalent emulsion (100 per cent).