Phase TEM for biological imaging utilizing a Boersch electrostatic phase plate: theory and practice.

A Boersch electrostatic phase plate (BEPP) used in a transmission electron microscope (TEM) system can provide tuneable phase shifts and overcome the low contrast problem for biological imaging. Theoretically, a pure phase image with a high phase contrast can be obtained using a BEPP. However, a currently available TEM system utilizing a BEPP cannot achieve sufficiently high phase efficiency for biological imaging, owing to the practical conditions. The low phase efficiency is a result of the blocking of partial unscattered electrons by BEPP, and the contribution of absorption contrast. The fraction of blocked unscattered beam is related to BEPP dimensions and to divergence of the illumination system of the TEM. These practical issues are discussed in this paper. Phase images of biological samples (negatively stained ferritin) obtained by utilizing a BEPP are reported, and the phase contrast was found to be enhanced by a factor of approximately 1.5, based on the calculation using the Rose contrast criterion. The low gain in phase contrast is consistent with the expectation from the current TEM/BEPP system. A new generation of phase TEM utilizing BEPP and designed for biological imaging with a high phase efficiency is proposed.

The fabrication and application of Zernike electrostatic phase plate.

The Zernike electrostatic phase plate (ZEPP) consisting of three ring electrodes and two insulating rings has been fabricated using Micro Electro-Mechanical System processes. The electrostatic field produced in the phase plate was measured by electrostatic force microscopy. The electrostatic field and focal length of the phase plate is calculated using electron optics principles. The comparison of the experimental electrostatic field and the theoretical calculated is discussed. Finally, we report the application of ZEPP to enhance contrast of the SiON(x)/SiO(2) interface in an NMOS semiconductor device. The quantitative analysis of the contrast enhancement versus the applied bias is discussed in detail.