Scanning electron microscope autoradiography of critical point dried biological samples

A technique has been developed for the localization of isotopes in the scanning electron microscope. Autoradiographic studies have been performed using a model system and a unicellular biflagellate alga. One requirement of this technique is that all manipulations be carried out on samples that are maintained in a liquid state. Observations of a source of radiation (125I-ferritin) show that the nuclear emulsion used to detect radiation is active under these conditions. Efficiency measurement performed using 125I-ferritin indicate that 125-I-SEM autoradiography is an efficient process that exhibits a 'dose dependent' response. Two types of labeling methods were used with cells, surface labeling with 125I and internal labeling with 3H. Silver grains appeared on labeled cells after autoradiography, removal of residual gelatin and critical point drying. The location of grains was examined on a flagellated green alga (Chlamydomonas reinhardi) capable of undergoing cell fusion. Fusion experiments using labeled and unlabeled cells indicate that 1. Labeling is specific for incorporated radioactivity; 2. Cell surface structure is preserved in SEM autoradiographs and 3. The technique appears to produce reliable autoradiographs. Thus scanning electron microscope autoradiography should provide a new and useful experimental approach.