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Applicability of thin film phase plates in biological electron microscopy

View Article: PubMed Central - PubMed

ABSTRACT

Presented is an evaluation of phase contrast techniques in transmission electron microscopy. The traditional defocus phase contrast is compared to two recently developed phase plate techniques. One is the Zernike phase contrast transmission electron microscope, the other is the Hilbert differential contrast thransmission electron microscope. The imaging characteristics of each technique are discussed. Phase plate techniques provide improved contrast for ice-embedded biological samples which are a challenge for the conventional defocus phase contrast. The flat spectral response of the Zernike and Hilbert modes extends towards the low frequencies which are severely suppressed in the conventional defocus mode. Target applications for each of the phase contrast techniques are discussed based on the specifics of image formation and spectral transfer.

No MeSH data available.


Images of polystyrene latex particles demonstrating the effect of the central diffraction beam to HDC-TEM phase plate edge distance on the image and Fourier space. (a) central beam positioned close to the phase plate edge. (c) central beam moved some distance from the edge. (b) and (d) modulus of the Fourier transforms of (a) and (c) respectively.
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f5-2_35: Images of polystyrene latex particles demonstrating the effect of the central diffraction beam to HDC-TEM phase plate edge distance on the image and Fourier space. (a) central beam positioned close to the phase plate edge. (c) central beam moved some distance from the edge. (b) and (d) modulus of the Fourier transforms of (a) and (c) respectively.

Mentions: The low cutoff frequency in the case of HDC-TEM is not uniform in all directions. It is lowest in direction perpendicular to the edge. In parallel to the edge direction there is a stripe in the Fourier space exhibiting conventional TEM phase contrast transfer. The width of this stripe is determined by the central beam to edge distance. Fig. 5 demonstrates the effect of this distance on the appearance of the image and the Fourier space. When the beam is close to the phase plate edge (Figs. 5a, b) the image exhibits a strong topographic contrast. This is due to the realization of phase contrast for low and very low spatial frequencies. Larger details are characteristic of biological samples on the medium and large scale such as organelles, bacteria, cells, etc. Moving the beam further from the edge results in vast reduction of the overall contrast in the image (Figs. 5c, d). Only a slight hint of topographic contrast remains. In practice working as close as possible to the edge is most desirable. This produces images with the highest contrast and most low frequency information. The pCTF envelope of HDC-TEM is identical to that of ZPC-TEM so the optimal focus condition is also similar. Working close to focus or with slight underfocus is most desirable.


Applicability of thin film phase plates in biological electron microscopy
Images of polystyrene latex particles demonstrating the effect of the central diffraction beam to HDC-TEM phase plate edge distance on the image and Fourier space. (a) central beam positioned close to the phase plate edge. (c) central beam moved some distance from the edge. (b) and (d) modulus of the Fourier transforms of (a) and (c) respectively.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC5036645&req=5

f5-2_35: Images of polystyrene latex particles demonstrating the effect of the central diffraction beam to HDC-TEM phase plate edge distance on the image and Fourier space. (a) central beam positioned close to the phase plate edge. (c) central beam moved some distance from the edge. (b) and (d) modulus of the Fourier transforms of (a) and (c) respectively.
Mentions: The low cutoff frequency in the case of HDC-TEM is not uniform in all directions. It is lowest in direction perpendicular to the edge. In parallel to the edge direction there is a stripe in the Fourier space exhibiting conventional TEM phase contrast transfer. The width of this stripe is determined by the central beam to edge distance. Fig. 5 demonstrates the effect of this distance on the appearance of the image and the Fourier space. When the beam is close to the phase plate edge (Figs. 5a, b) the image exhibits a strong topographic contrast. This is due to the realization of phase contrast for low and very low spatial frequencies. Larger details are characteristic of biological samples on the medium and large scale such as organelles, bacteria, cells, etc. Moving the beam further from the edge results in vast reduction of the overall contrast in the image (Figs. 5c, d). Only a slight hint of topographic contrast remains. In practice working as close as possible to the edge is most desirable. This produces images with the highest contrast and most low frequency information. The pCTF envelope of HDC-TEM is identical to that of ZPC-TEM so the optimal focus condition is also similar. Working close to focus or with slight underfocus is most desirable.

View Article: PubMed Central - PubMed

ABSTRACT

Presented is an evaluation of phase contrast techniques in transmission electron microscopy. The traditional defocus phase contrast is compared to two recently developed phase plate techniques. One is the Zernike phase contrast transmission electron microscope, the other is the Hilbert differential contrast thransmission electron microscope. The imaging characteristics of each technique are discussed. Phase plate techniques provide improved contrast for ice-embedded biological samples which are a challenge for the conventional defocus phase contrast. The flat spectral response of the Zernike and Hilbert modes extends towards the low frequencies which are severely suppressed in the conventional defocus mode. Target applications for each of the phase contrast techniques are discussed based on the specifics of image formation and spectral transfer.

No MeSH data available.