Factors which Affect Efficiency of Autoradiography with Tritiated Thymidine

Abstract

The overall efficiency of autoradiography with tritium-labeled thymidine has been found to be influenced by the following conditions: (1) exposure in an atmosphere of CO₂ and the use of the stripping-film technique, both of which increase the autoradiographic efficiency when compared to exposure in air or to dip-coating technique; (2) latent image fading, which increases with increasing exposure. Up to 2 wk of exposure, however, this disadvantage is counterbalanced by the fading of the mechanical background produced during stripping or dip-coating; (3) the thickness of the inert coating interposed between the labeled locus and the sensitized emulsion. A layer of inert coating can be obtained that will arrest all beta particles from tritium, while having no effect on more energetic emitters like C¹⁴; (4) the amount of tritiated thymidine given, with relatively large amounts producing an increase in the mean grain count per labeled cell but not in the percentage of cells identifiable as labeled; and (5) the type of fixative and the staining procedure used. Feulgen stain reduces the mean grain count in cells labeled with radioactive thymidine, while fixation with acetic alcohol (1:3) reduces the grain count in cells labeled with precursors of ribonucleic acid.