Scanning transmission electron microscopy of tagged proteins in whole eukaryotic cells
Dukes, Madeline Jayne
The purpose of the dissertation is to develop whole cell electron microscopy (EM) techniques capable of imaging tagged proteins. The utility of lanthanide chelate molecules as membrane penetrating bimodal fluorescent and EM imaging agents is presented. In order to obtain EM images of cellular components in physiologically relevant environments, I introduce a correlative approach to image quantum dots on whole eukaryotic cells in liquid with fluorescence microscopy and with scanning transmission electron microscopy (STEM). A key feature of this method is that it requires no additional sample processing for EM, beyond chemical fixation. In addition to liquid cell samples, I demonstrate utilizing aberration-corrected STEM to study the three dimensional distribution of gold nanoparticles in whole cells by means of a focal series, without tilting the specimen. A quantitative method to measure the structural stability of dry whole cells was developed and samples were optimized for stability. The axial positions of gold-tagged epidermal growth factor receptors were determined.