Abstract
In this dissertation, we present a new route to achieve the same goals by hollow cone dark field (HCDF) imaging using thermal diffuse scattered (TDS) electrons giving about a 4 times contrast increase as compared to bright field imaging It can be divided into three topics: (1) negative staining, (2) Cryo-electron microscopy, and (3) in situ liquid cell TEM. In the first part, we demonstrated the optimum condition of HCDF and contrast enhancement by negative stained GroEL. After that, we demonstrated the 3D reconstruction of a negative stained GroEL particle can yield about 13.5Å resolution but using a strongly reduced number of images. In the second part, we established the process flow of HCDF technique for Cryo-EM field and demonstrated the concept by liposome. Later, we demonstrated the first result utilized HCDF technique in various biological sample including Keyhole Limpet Hemocyanin (KLH) protein, Adenovirus and Nervous Necrosis Virus (NNV) and the 3D reconstruction model of NNV. In the last part, we demonstrated the HCDF in in situ liquid TEM field and observed the Magnetospirillum magnetotacticum, (MS-1). We proved the HCDF technique provided contrast enhancement and more detail than tradition bright field mode.