A scanning microscope with 5 Å resolution

Crewe, A. V.; Wall, Joseph S.

doi:10.1016/0022-2836(70)90052-5

Cited by 214 publications

(69 citation statements)

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“…In contrast the scanning transmission electron microscope (STEM) was already employed to make nano-scale measurements on biological samples in the 1970s [3] [4] and retains its importance today (for reviews see [5][6]). Key was the development of the field emission gun and its incorporation in the STEM [7]. This electron source delivers a highly coherent beam of electrons that can be focused to a spot less than 0.5 nm in diameter.…”

Section: Introductionmentioning

confidence: 99%

Biological Scanning Transmission Electron Microscopy: Imaging and Single Molecule Mass Determination

Müller¹,

Engel²

2006

Chimia

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scanning transmission electron microscopes can both measure the mass of single protein complexes and take amazingly clear images thereof, offering a wide range of applications in structural biology. The principle of mass measurement is presented and discussed and the scope of scanning transmission electron microscopy is illustrated by selected examples. Keywords:Mass determination · scanning transmission electron microscopy · sTEM unstained proteins to be clearly visualised under low dose conditions. Each pixel of the digital image is a quantitative scattering experiment from which the mass of the irradiated volume can be calculated [9]. Thus, the mass of a single protein complex can be determined by integrating over all pixel values that lie within its boundary. The method is applicable to macromolecules of widely varying mass and form (reviewed in [10]). When combined with SDS-gel electrophoresis and, where available, sequence information, STEM mass measurements serve to define protein stoichiometry. Further, as electron energy loss spectroscopy [11][12] or energy dispersive X-ray spectroscopy [13] can be performed while imaging, the STEM also offers the possibility of determining the distribution of specific elements in protein complexes. Indeed the feasibility of mapping single atoms bound to protein molecules has recently been demonstrated [14].The high contrast delivered by the dark-field detector system [15] can also be exploited in negative or positive stain microscopy [16]. Here the STEM yields clear single-shot images that are free from phase contrast fringes. These images frequently provide information only otherwise visible after averaging several hundred projections recorded with a transmission electron microscope (TEM). They thus make it possible to document the presence of distinct protein conformations, information that would be lost by averaging techniques. Indeed, STEM images of both negatively stained [17] and unstained [18] protein complexes have been used to reconstruct their 3D structure.The few dedicated STEMs employed in biology today are invaluable. In particular, the coupling of image and mass gives them the power to answer questions not resolvable by ultracentrifugation or mass spectrometry. Here we discuss the use of the STEM both as an imaging tool and to measure mass, and examine the uncertainties to be expected in STEM mass data sets. The Importance and Versatility of STEM Mass MeasurementsWorldwide, only four laboratories routinely use dedicated STEMs to measure the mass of biological samples. Their facilities are open for external collaborations and the many papers in the literature containing STEM mass measurements document the importance of this rare technique to the scientific community. Although the precision of STEM cannot match that of the mass spectrometer, its capability to measure an enormously wide mass range, its independence from shape assumptions and the presence of an image make the STEM an invaluable tool.The methodology of mass spectrometry has develop...

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Section: Introductionmentioning

confidence: 99%

Biological Scanning Transmission Electron Microscopy: Imaging and Single Molecule Mass Determination

Müller¹,

Engel²

2006

Chimia

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“…The study of field emitters as potential electron sources for electron-beam instruments started around 1954 (Dyke & Dolan, 1956). The first practical utilization of such sources appeared late in 1968 as a part of a scanning electron microscope (Crewe et al, 1968(Crewe et al, ,1969Crewe & Wall, 1970). Many experimental FE studies have been carried out on tungsten, lanthanum hexaboride and carbon fiber emitters.…”

Section: Introductionmentioning

confidence: 99%

The Effects of Dielectric Coatings on Electron Emission from Tungsten

Al-Qudah

Alnawasreh

Madanat

et al. 2017

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Field electron emission measurements were performed on dielectric-coated tungsten emitters, with apex radii in the nanometer and micrometer range, which were prepared by electrochemical etching in NaOH solution. Measurements were performed in a field electron microscopy (FEM) with a base pressure <10 -6 Pascal (10 -8 mbar). Four different types of dielectric were used, namely: (1) Clark Electromedical Instruments epoxylite resin, (2) Epidian 6 produced by Ciech Sarzyna S. A., (3) a Radionox solution of colloidal graphite; and (4) Molyslip 2001 E compound (MoS 2 and MoS). Currentvoltage measurements and FEM images were used to investigate the characteristics of these composite emitters, and to assess how the different types of dielectric coating affect the suitability of the composite emitter as a potential electron source.

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“…Our aim is to emulate and complement the achievements of scanning transmission electron microscopy (2), ultimately at comparable resolution, by taking advantage of the interaction properties of fast ions with matter (3). The latter include processes such as charge exchange and molecular ion dissociation which should result in contrast mechanisms unavailable to the electron microscope.…”

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confidence: 99%

“…[2] From [2] we obtain an expression for the radius of the Gaussian image of the source in terms of probe current and specific brightness: rG = 1118 = (Io/3V)i/'(1/irac) [3] This contribution to the final spot size must be combined with those arising from diffraction and from spherical and chromatic aberration. As often done (9), we add these contributions in quadrature and seek the optimum value a0p, of as that reduces the overall probe radius r to its minimum value ropt.…”

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Scanning transmission ion microscope with a field ion source.

Escovitz¹,

Fox²,

Levi‐Setti³

1975

Proc. Natl. Acad. Sci. U.S.A.

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Experiments with a low-resolution scanning transmission ion microscope, using hydrogen ions from a field ionization source, indicate that it will be feasible by this approach to aim at high-resolution ion microscopy. Micrographs of unstained biological specimens have been obtained by critical range absorption of a 55 keV hydrogen ion beam at a resolution of 2000 A.As a first step toward the development of a high-resolution scanning transmission ion microscope, we have constructed and operated a 65 kV prototype ion-gun scanning transmission ion microscope which accelerates and focuses hydrogen ions from a field ionization (1) source. Our aim is to emulate and complement the achievements of scanning transmission electron microscopy (2), ultimately at comparable resolution, by taking advantage of the interaction properties of fast ions with matter (3). The latter include processes such as charge exchange and molecular ion dissociation which should result in contrast mechanisms unavailable to the electron microscope.We present here experimental data that are relevant to the feasibility of a high-resolution scanning transmission ion microscope and show the first low-resolution images of biological specimens obtained with the prototype instrument.Previous attempts at using ions, and in particular protons, to make images of small objects with conventional electron microscope optics have been reviewed by Grivet (4) and more recently by Levi-Setti (3). The aim was to obtain higher resolution than with electrons. This was believed to be possible because of the ion's shorter De Broglie wavelength. The potential of proton charge exchange as a possible new source of contrast was recognized by Chanson and Magnan (5).Several factors, however, conspired to abort this development of the proton microscope as a high-resolution and viable instrument. In our view, the principal limitations originated in the approach of conventional microscopy itself, where the large rate of proton energy loss (10-30 eV/A in traversing organic materials at 100 keV) causes irreparable chromatic aberration. For this reason, we have chosen the approach of scanning transmission microscopy, where no optics is affected by the beam-specimen interaction so that the latter can be used to advantage in producing contrast.The considerations leading to the optimum parameters and the resulting performance in the scanning transmission electron microscope have been recently reviewed by A. V. Crewe (6) and E. Zeitler (7). We have extended the analysis of these authors to the case of a proton probe (8). The basic problem consists of optimizing the design parameters to obtain the maximum probe current in the smallest beam spot. How closely the theoretical microscope resolution can be approached in practice depends essentially on the specific brightness of the ion (or electron) source, since the probe current must exceed that required to obtain an intelligible picture in a practicable scan time. We consider a microscope consisting of a source of radius 6 and an ...

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A scanning microscope with 5 Å resolution

Cited by 214 publications

References 8 publications

Biological Scanning Transmission Electron Microscopy: Imaging and Single Molecule Mass Determination

Biological Scanning Transmission Electron Microscopy: Imaging and Single Molecule Mass Determination

The Effects of Dielectric Coatings on Electron Emission from Tungsten

Scanning transmission ion microscope with a field ion source.

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