Studies of Aggregates of Two Muscle Proteins with Scanning Transmission and Conventional Transmission Electron Microscopy

Author(s):  
M. K. Lamvik ◽  
M. S. Isaacson ◽  
A. V. Crewe

Studies of the structures of various aggregates of proteins from vertebrate striated muscle have begun, using in particular the scanning transmission electron microscope (STEM). The high collection efficiency of this microscope has been noted elsewhere and the contrast advantage and signal selection capabilities have been demonstrated.Although the instrument can produce high contrast images of unstained and unfixed biological material, it is not clear which preparation methods might maintain the native structure of such material. So the study begins on familiar ground with conventionl negatively stained preparations, While later steps will include less conventional methods.Rabbit tropomyosin Mg-tactoids in suspension were received from Dr. Carolyn Cohen (of Children's Cancer Research Foundation, Boston). The tactoids have a primary periodicity of 395 Å. Isolated myofibrils were obtained from Joseph Etlinger (of this University); myosin was extracted from them by a method similar to that of Dow and Stracher.

Author(s):  
L. Gandolfi ◽  
J. Reiffel

Calculations have been performed on the contrast obtainable, using the Scanning Transmission Electron Microscope, in the observation of thick specimens. Recent research indicates a revival of an earlier interest in the observation of thin specimens with the view of comparing the attainable contrast using both types of specimens.Potential for biological applications of scanning transmission electron microscopy has led to a proliferation of the literature concerning specimen preparation methods and the controversy over “to stain or not to stain” in combination with the use of the dark field operating mode and the same choice of technique using bright field mode of operation has not yet been resolved.


Author(s):  
H. Rose

The scanning transmission electron microscope offers the possibility of utilizing inelastically scattered electrons. Use of these electrons in addition to the elastically scattered electrons should reduce the scanning time (dose) Which is necessary to keep the quantum noise below a certain level. Hence it should lower the radiation damage. For high resolution, Where the collection efficiency of elastically scattered electrons is small, the use of Inelastically scattered electrons should become more and more favorable because they can all be detected by means of a spectrometer. Unfortunately, the Inelastic scattering Is a non-localized interaction due to the electron-electron correlation, occurring predominantly at the circumference of the atomic electron cloud.


Author(s):  
J. F. Hainfeld ◽  
P. S. Furcinitti ◽  
J. S. Wall

Several studies of molecular structure have successfully employed computer techniques to align images of single particles. Image processing software packages (e.g., P. R. Smith's MDPP system and J. Frank's SPIDER system) have also been developed to facilitate this work. Due to the low contrast and high beam damage rate involved in the use of unstained specimens, most single particle (i.e., non-crystalline) image alignment has been of negatively stained specimens. The scanning transmission electron microscope (STEM), which has a linear transfer function at high resolution rather than the oscillating one inherent to CEMs, operationally overcomes these limitations by providing low dose, high contrast images of unstained material with high collection efficiency.


Author(s):  
S. Nomura ◽  
H. Todokoro ◽  
T. Komoda

The Scanning Transmission Electron Microscope (STEM) has made possible specimen observation with a number of advantages such as high signal collection efficiency. In addition, STEM also permits element analysis of micro-areas, when it is used in conjunction with X-ray and/or electron spectrometers. These advantages become more effective by using a high brightness electron gun.The authors have developed a field emission STEM. The schematic diagram of the instrument is shown in Fig. 1. Electrons emitted from the tungsten tip are focused on a specimen by one electro-static and two magnetic lenses. The field emission tip is surrounded by ion pumps, and the vacuum of the gun chamber is maintained at better than 5xlO-10torr.


Author(s):  
A. V. Crewe

The high resolution STEM is now a fact of life. I think that we have, in the last few years, demonstrated that this instrument is capable of the same resolving power as a CEM but is sufficiently different in its imaging characteristics to offer some real advantages.It seems possible to prove in a quite general way that only a field emission source can give adequate intensity for the highest resolution^ and at the moment this means operating at ultra high vacuum levels. Our experience, however, is that neither the source nor the vacuum are difficult to manage and indeed are simpler than many other systems and substantially trouble-free.


Author(s):  
J. S. Wall ◽  
J. P. Langmore ◽  
H. Isaacson ◽  
A. V. Crewe

The scanning transmission electron microscope (STEM) constructed by the authors employs a field emission gun and a 1.15 mm focal length magnetic lens to produce a probe on the specimen. The aperture size is chosen to allow one wavelength of spherical aberration at the edge of the objective aperture. Under these conditions the profile of the focused spot is expected to be similar to an Airy intensity distribution with the first zero at the same point but with a peak intensity 80 per cent of that which would be obtained If the lens had no aberration. This condition is attained when the half angle that the incident beam subtends at the specimen, 𝛂 = (4𝛌/Cs)¼


Author(s):  
H. Koike ◽  
S. Sakurai ◽  
K. Ueno ◽  
M. Watanabe

In recent years, there has been increasing demand for higher voltage SEMs, in the field of surface observation, especially that of magnetic domains, dislocations, and electron channeling patterns by backscattered electron microscopy. On the other hand, the resolution of the CTEM has now reached 1 ∼ 2Å, and several reports have recently been made on the observation of atom images, indicating that the ultimate goal of morphological observation has beem nearly achieved.


Author(s):  
A. Kosiara ◽  
J. W. Wiggins ◽  
M. Beer

A magnetic spectrometer to be attached to the Johns Hopkins S. T. E. M. is under construction. Its main purpose will be to investigate electron interactions with biological molecules in the energy range of 40 KeV to 100 KeV. The spectrometer is of the type described by Kerwin and by Crewe Its magnetic pole boundary is given by the equationwhere R is the electron curvature radius. In our case, R = 15 cm. The electron beam will be deflected by an angle of 90°. The distance between the electron source and the pole boundary will be 30 cm. A linear fringe field will be generated by a quadrupole field arrangement. This is accomplished by a grounded mirror plate and a 45° taper of the magnetic pole.


Author(s):  
Michael Beer ◽  
J. W. Wiggins ◽  
David Woodruff ◽  
Jon Zubin

A high resolution scanning transmission electron microscope of the type developed by A. V. Crewe is under construction in this laboratory. The basic design is completed and construction is under way with completion expected by the end of this year.The optical column of the microscope will consist of a field emission electron source, an accelerating lens, condenser lens, objective lens, diffraction lens, an energy dispersive spectrometer, and three electron detectors. For any accelerating voltage the condenser lens function to provide a parallel beam at the entrance of the objective lens. The diffraction lens is weak and its current will be controlled by the objective lens current to give an electron diffraction pattern size which is independent of small changes in the objective lens current made to achieve focus at the specimen. The objective lens demagnifies the image of the field emission source so that its Gaussian size is small compared to the aberration limit.


Author(s):  
J. R. Fields

The energy analysis of electrons scattered by a specimen in a scanning transmission electron microscope can improve contrast as well as aid in chemical identification. In so far as energy analysis is useful, one would like to be able to design a spectrometer which is tailored to his particular needs. In our own case, we require a spectrometer which will accept a parallel incident beam and which will focus the electrons in both the median and perpendicular planes. In addition, since we intend to follow the spectrometer by a detector array rather than a single energy selecting slit, we need as great a dispersion as possible. Therefore, we would like to follow our spectrometer by a magnifying lens. Consequently, the line along which electrons of varying energy are dispersed must be normal to the direction of the central ray at the spectrometer exit.


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