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Title page for ETD etd-03252011-125343


Type of Document Dissertation
Author Dukes, Madeline Jayne
Author's Email Address madeline.j.dukes@gmail.com
URN etd-03252011-125343
Title Scanning transmission electron microscopy of tagged proteins in whole eukaryotic cells
Degree PhD
Department Chemistry
Advisory Committee
Advisor Name Title
Darryl J.Bornhop Committee Chair
B. Andes Hess Committee Member
David W. Piston Committee Member
Niels de Jonge Committee Member
Sandra J. Rosenthal Committee Member
Keywords
  • scanning transmission electron microscopy; quantum dots;
  • molecular probes; protein labels; correlative microscopy; fluorescence microscopy; electron microsco
Date of Defense 2011-05-18
Availability unrestricted
Abstract
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.
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