Analysis of Recombinant Adeno-Associated Virus (rAAV) Sample Morphology Using Negative Staining and High-Resolution Electron Microscopy

Spirillum serpens possesses a surface layer protein which exhibits a regular hexagonal packing of the morphological subunits. A morphological model of the structure of the protein has been proposed at a resolution of about 25 Å, in which the morphological unit might be described as having the appearance of a flared-out, hollow cylinder with six ÅspokesÅ at the flared end. In order to understand the detailed association of the macromolecules, it is necessary to do a high resolution structural analysis. Large, single layered arrays of the surface layer protein have been obtained for this purpose by means of extensive heating in high CaCl2, a procedure derived from that of Buckmire and Murray. Low dose, low temperature electron microscopy has been applied to the large arrays.As a first step, the samples were negatively stained with neutralized phosphotungstic acid, and the specimens were imaged at 40,000 magnification by use of a high resolution cold stage on a JE0L 100B. Low dose images were recorded with exposures of 7-9 electrons/Å2. The micrographs obtained (Fig. 1) were examined by use of optical diffraction (Fig. 2) to tell what areas were especially well ordered.

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Electron microscopy of rabbit FC crystals

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100077220 ◽

1983 ◽

Vol 41 ◽

pp. 730-731

Author(s):

Robert A. Grant ◽

Laura L. Degn ◽

Wah Chiu ◽

John Robinson

Keyword(s):

Electron Microscopy ◽

High Resolution ◽

High Resolution Electron Microscopy ◽

Molecular Replacement ◽

X Ray ◽

X Ray Crystallography ◽

Resolution Electron ◽

Replacement Technique ◽

Hydrated Crystal ◽

Molecular Structure Analysis

Proteolytic digestion of the immunoglobulin IgG with papain cleaves the molecule into an antigen binding fragment, Fab, and a compliment binding fragment, Fc. Structures of intact immunoglobulin, Fab and Fc from various sources have been solved by X-ray crystallography. Rabbit Fc can be crystallized as thin platelets suitable for high resolution electron microscopy. The structure of rabbit Fc can be expected to be similar to the known structure of human Fc, making it an ideal specimen for comparing the X-ray and electron crystallographic techniques and for the application of the molecular replacement technique to electron crystallography. Thin protein crystals embedded in ice diffract to high resolution. A low resolution image of a frozen, hydrated crystal can be expected to have a better contrast than a glucose embedded crystal due to the larger density difference between protein and ice compared to protein and glucose. For these reasons we are using an ice embedding technique to prepare the rabbit Fc crystals for molecular structure analysis by electron microscopy.

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High resolution electron microscopy of lanthanum phosphate crystals

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100108519 ◽

1978 ◽

Vol 36 (1) ◽

pp. 274-275

Author(s):

J. C. Wheatley ◽

J. M. Cowley

Keyword(s):

Electron Microscopy ◽

Catalytic Activity ◽

High Resolution ◽

Petroleum Industry ◽

High Resolution Electron Microscopy ◽

Lanthanum Phosphate ◽

Good Catalyst ◽

Resolution Electron ◽

Good Catalytic Activity ◽

Physical And Chemical

Rare-earth phosphates are of particular interest because of their catalytic properties associated with the hydrolysis of many aromatic chlorides in the petroleum industry. Lanthanum phosphates (LaPO4) which have been doped with small amounts of copper have shown increased catalytic activity (1). However the physical and chemical characteristics of the samples leading to good catalytic activity are not known.Many catalysts are amorphous and thus do not easily lend themselves to methods of investigation which would include electron microscopy. However, the LaPO4, crystals are quite suitable samples for high resolution techniques.The samples used were obtained from William L. Kehl of Gulf Research and Development Company. The electron microscopy was carried out on a JEOL JEM-100B which had been modified for high resolution microscopy (2). Standard high resolution techniques were employed. Three different sample types were observed: 669A-1-5-7 (poor catalyst), H-L-2 (good catalyst) and 27-011 (good catalyst).

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Possible applications of fraunhofer holography in high resolution electron microscopy

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100108258 ◽

1978 ◽

Vol 36 (1) ◽

pp. 222-223 ◽

Cited By ~ 1

Author(s):

N. Bonnet ◽

M. Troyon ◽

P. Gallion

Keyword(s):

Electron Microscopy ◽

High Resolution ◽

Transfer Function ◽

Radiation Damage ◽

High Resolution Electron Microscopy ◽

Far Field ◽

Field Region ◽

Crystalline Materials ◽

Resolution Electron ◽

The Ideal

Two main problems in high resolution electron microscopy are first, the existence of gaps in the transfer function, and then the difficulty to find complex amplitude of the diffracted wawe from registered intensity. The solution of this second problem is in most cases only intended by the realization of several micrographs in different conditions (defocusing distance, illuminating angle, complementary objective apertures…) which can lead to severe problems of contamination or radiation damage for certain specimens.Fraunhofer holography can in principle solve both problems stated above (1,2). The microscope objective is strongly defocused (far-field region) so that the two diffracted beams do not interfere. The ideal transfer function after reconstruction is then unity and the twin image do not overlap on the reconstructed one.We show some applications of the method and results of preliminary tests.Possible application to the study of cavitiesSmall voids (or gas-filled bubbles) created by irradiation in crystalline materials can be observed near the Scherzer focus, but it is then difficult to extract other informations than the approximated size.

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The observation of solutions in Ni3Nb

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100104765 ◽

1988 ◽

Vol 46 ◽

pp. 540-541

Author(s):

Z.M. Wang ◽

J.P. Zhang

Keyword(s):

Electron Microscopy ◽

High Resolution ◽

Phase Modulation ◽

High Resolution Electron Microscopy ◽

Antiphase Domain ◽

Boundary Area ◽

Matrix Type ◽

Resolution Electron ◽

Domain Boundaries ◽

Kontorova Model

High resolution electron microscopy reveals that antiphase domain boundaries in β-Ni3Nb have a hexagonal unit cell with lattice parameters ah=aβ and ch=bβ, where aβ and bβ are of the orthogonal β matrix. (See Figure 1.) Some of these boundaries can creep “upstairs” leaving an incoherent area, as shown in region P. When the stepped boundaries meet each other, they do not lose their own character. Our consideration in this work is to estimate the influnce of the natural misfit δ{(ab-aβ)/aβ≠0}. Defining the displacement field at the boundary as a phase modulation Φ(x), following the Frenkel-Kontorova model [2], we consider the boundary area to be made up of a two unit chain, the upper portion of which can move and the lower portion of the β matrix type, assumed to be fixed. (See the schematic pattern in Figure 2(a)).

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Atomic structure imaging of the Si/SiO2 interface with high-resolution electron microscopy

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100143559 ◽

1986 ◽

Vol 44 ◽

pp. 390-391

Author(s):

J.L. Batstone ◽

J.M. Gibson ◽

Alice.E. White ◽

K.T. Short

Keyword(s):

Electron Microscopy ◽

High Resolution ◽

Atomic Structure ◽

High Resolution Electron Microscopy ◽

Medium Voltage ◽

Voltage Electron ◽

Resolution Electron ◽

Structural Image ◽

Electron Microscopes ◽

Point To Point

High resolution electron microscopy (HREM) is a powerful tool for the determination of interface atomic structure. With the previous generation of HREM's of point-to-point resolution (rpp) >2.5Å, imaging of semiconductors in only <110> directions was possible. Useful imaging of other important zone axes became available with the advent of high voltage, high resolution microscopes with rpp <1.8Å, leading to a study of the NiSi2 interface. More recently, it was shown that images in <100>, <111> and <112> directions are easily obtainable from Si in the new medium voltage electron microscopes. We report here the examination of the important Si/Si02 interface with the use of a JEOL 4000EX HREM with rpp <1.8Å, in a <100> orientation. This represents a true structural image of this interface.

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High-resolution electron microscopy of nanostructured materials

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100137239 ◽

1995 ◽

Vol 53 ◽

pp. 172-173

Author(s):

M. José-Yacamán

Keyword(s):

Electron Microscopy ◽

High Resolution ◽

Grain Boundaries ◽

Nanostructured Materials ◽

High Resolution Electron Microscopy ◽

Hrem Image ◽

Small Particles ◽

Resolution Electron ◽

Nanophase Materials

Electron microscopy is a fundamental tool in materials characterization. In the case of nanostructured materials we are looking for features with a size in the nanometer range. Therefore often the conventional TEM techniques are not enough for characterization of nanophases. High Resolution Electron Microscopy (HREM), is a key technique in order to characterize those materials with a resolution of ~ 1.7A. High resolution studies of metallic nanostructured materials has been also reported in the literature. It is concluded that boundaries in nanophase materials are similar in structure to the regular grain boundaries. That work therefore did not confirm the early hipothesis on the field that grain boundaries in nanostructured materials have a special behavior. We will show in this paper that by a combination of HREM image processing, and image calculations, it is possible to prove that small particles and coalesced grains have a significant surface roughness, as well as large internal strain.

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