Compact 4pi beam scanning microscope for imaging cells with a sevenfold increase in axial resolution

We have become accustomed to differentiating between the scanning microscope and the conventional transmission microscope according to the resolving power which the two instruments offer. The conventional microscope is capable of a point resolution of a few angstroms and line resolutions of periodic objects of about 1Å. On the other hand, the scanning microscope, in its normal form, is not ordinarily capable of a point resolution better than 100Å. Upon examining reasons for the 100Å limitation, it becomes clear that this is based more on tradition than reason, and in particular, it is a condition imposed upon the microscope by adherence to thermal sources of electrons.

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A New Image Processing Technique for STEM

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100052171 ◽

1974 ◽

Vol 32 ◽

pp. 432-433

Author(s):

Yasushi Kokubo ◽

Hirotami Koike ◽

Teruo Someya

Keyword(s):

Electron Microscopy ◽

Image Processing ◽

Scanning Electron Microscopy ◽

Elastic Scattering ◽

Field Emission ◽

Processing Technique ◽

Image Processing Technique ◽

Energy Analyzer ◽

Scanning Microscope ◽

Scanning Electron

One of the advantages of scanning electron microscopy is the capability for processing the image contrast, i.e., the image processing technique. Crewe et al were the first to apply this technique to a field emission scanning microscope and show images of individual atoms. They obtained a contrast which depended exclusively on the atomic numbers of specimen elements (Zcontrast), by displaying the images treated with the intensity ratio of elastically scattered to inelastically scattered electrons. The elastic scattering electrons were extracted by a solid detector and inelastic scattering electrons by an energy analyzer. We noted, however, that there is a possibility of the same contrast being obtained only by using an annular-type solid detector consisting of multiple concentric detector elements.

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A High Resolution Scanning Microscope

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100101529 ◽

1968 ◽

Vol 26 ◽

pp. 356-357

Author(s):

A. V. Crewe ◽

J. Wall ◽

L. M. Welter

Keyword(s):

High Resolution ◽

Field Emission ◽

Spherical Aberration ◽

Focal Length ◽

Spot Size ◽

Emission Source ◽

Field Of View ◽

Scanning Microscope ◽

Magnetic Lens ◽

High Quality

A scanning microscope using a field emission source has been described elsewhere. This microscope has now been improved by replacing the single magnetic lens with a high quality lens of the type described by Ruska. This lens has a focal length of 1 mm and a spherical aberration coefficient of 0.5 mm. The final spot size, and therefore the microscope resolution, is limited by the aberration of this lens to about 6 Å.The lens has been constructed very carefully, maintaining a tolerance of + 1 μ on all critical surfaces. The gun is prealigned on the lens to form a compact unit. The only mechanical adjustments are those which control the specimen and the tip positions. The microscope can be used in two modes. With the lens off and the gun focused on the specimen, the resolution is 250 Å over an undistorted field of view of 2 mm. With the lens on,the resolution is 20 Å or better over a field of view of 40 microns. The magnification can be accurately varied by attenuating the raster current.

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The Potentials of Scanning Microscopy

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100101505 ◽

1968 ◽

Vol 26 ◽

pp. 352-353

Author(s):

A. V. Crewe

Keyword(s):

Salient Feature ◽

Secondary Emission ◽

Resolving Power ◽

X Rays ◽

Scanning Microscope ◽

Forming Process ◽

Additional Tool ◽

Detection Systems ◽

Conventional Microscope ◽

Present Information

If the resolving power of a scanning electron microscope can be improved until it is comparable to that of a conventional microscope, it would serve as a valuable additional tool in many investigations.The salient feature of scanning microscopes is that the image-forming process takes place before the electrons strike the specimen. This means that several different detection systems can be employed in order to present information about the specimen. In our own particular work we have concentrated on the use of energy loss information in the beam which is transmitted through the specimen, but there are also numerous other possibilities (such as secondary emission, generation of X-rays, and cathode luminescence).Another difference between the pictures one would obtain from the scanning microscope and those obtained from a conventional microscope is that the diffraction phenomena are totally different. The only diffraction phenomena which would be seen in the scanning microscope are those which exist in the beam itself, and not those produced by the specimen.

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3-D imaging of cells using a confocal laser scanning microscope and digital image processing

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100102560 ◽

1988 ◽

Vol 46 ◽

pp. 96-97

Author(s):

Thomas M. Jovin ◽

Michel Robert-Nicoud ◽

Donna J. Arndt-Jovin ◽

Thorsten Schormann

Keyword(s):

Laser Scanning ◽

Confocal Laser Scanning Microscope ◽

Laser Scanning Microscopy ◽

Confocal Laser Scanning ◽

Confocal Laser ◽

Scanning Microscope ◽

Laser Scanning Microscope ◽

Biochemical Processes ◽

Adjacent Structures

Light microscopic techniques for visualizing biomolecules and biochemical processes in situ have become indispensable in studies concerning the structural organization of supramolecular assemblies in cells and of processes during the cell cycle, transformation, differentiation, and development. Confocal laser scanning microscopy offers a number of advantages for the in situ localization and quantitation of fluorescence labeled targets and probes: (i) rejection of interfering signals emanating from out-of-focus and adjacent structures, allowing the “optical sectioning” of the specimen and 3-D reconstruction without time consuming deconvolution; (ii) increased spatial resolution; (iii) electronic control of contrast and magnification; (iv) simultanous imaging of the specimen by optical phenomena based on incident, scattered, emitted, and transmitted light; and (v) simultanous use of different fluorescent probes and types of detectors.We currently use a confocal laser scanning microscope CLSM (Zeiss, Oberkochen) equipped with 3-laser excitation (u.v - visible) and confocal optics in the fluorescence mode, as well as a computer-controlled X-Y-Z scanning stage with 0.1 μ resolution.

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