Resolution of electron microscope autoradiography. IV. Application to analysis of autoradiographs.

The previous publications of this series described the expected grain distributions around model radioactive structures in EM autoradiographs as a function of the specimen resolution. This family of expected distributions was called the "universal curves". In the present study, experiments on 14C-sources were compared, significant differences were found depending on the energy of the isotope. These differences were primarily in the tails of the distributions, and are therefore important in correcting for cross-scatter when analyzing electron microscope autoradiographs. Using the universal curves unique for 125I, 3H, and 14C, we designed three sets of transparent overlays, or "masks", one set for each of these isotopes. The masks can be used by an investigator in a manner similar to that suggested by Blackett and Parry to generate grain distributions in autoradiographs on the basis of any desired hypothesis regarding the levels of radioactivity in different structures. A subsequent comparison between these generated distributions and those obtained from the observed grains in these autoradiographs leads to a determination of the most likely levels of radioactivity in the tissue. A computer (described in an Appendix by Land and Salpeter) can be used to find the "best fit" levels of radioactivity in complex cases. The accuracy of the masks was checked on generated line sources for each of the three isotopes.