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Serial-section electron microscopy is the method of choice for studying cellular structure and network connectivity in the brain. We have built a pipeline of parallel imaging using transmission electron automated microscopes (piTEAM) that scales this technology and enables the acquisition of petascale datasets containing local cortical microcircuits. The distributed platform is composed of multiple transmission electron microscopes that image, in parallel, different sections from the same block of tissue, all under control of a custom acquisition software (pyTEM) that implements 24/7 continuous autonomous imaging. The suitability of this architecture for large scale electron microscopy imaging was demonstrated by acquiring a volume of more than 1 mm3 of mouse neocortex spanning four different visual areas. Over 26,500 ultrathin tissue sections were imaged, yielding a dataset of more than 2 petabytes. Our current burst imaging rate is 500 Mpixel/s (image capture only) per microscope and net imaging rate is 100 Mpixel/s (including stage movement, image capture, quality control, and post processing). This brings the combined burst acquisition rate of the pipeline to 3 Gpixel/s and the net rate to 600 Mpixel/s with six microscopes running acquisition in parallel, which allowed imaging a cubic millimeter of mouse visual cortex at synaptic resolution in less than 6 months.

Despite the use of electrons with wavelengths of just a few picometers, spatial resolution in a transmission electron microscope (TEM) has been limited by spherical aberration to typically around 0.15 nanometer. Individual atomic columns in a crystalline lattice can therefore only be imaged for a few low-order orientations, limiting the range of defects that can be imaged at atomic resolution. The recent development of spherical aberration correctors for transmission electron microscopy allows this limit to be overcome. We present direct images from an aberration-corrected scanning TEM that resolve a lattice in which the atomic columns are sepa-rated by less than 0.1 nanometer.

Electron-optical aberration correction has recently progressed from a promising concept to a powerful research tool. 100–120 kV scanning transmission electron microscopes (STEMs) equipped with spherical aberration (Cs) correctors now achieve sub-Å resolution in high-angle annular dark field (HAADF) imaging, and a 300 kV Cs-corrected STEM has reached 0.6 Å HAADF resolution. Moreover, the current available in an atom-sized probe has grown by about 10x, allowing electron energy loss spectroscopy (EELS) to detect single atoms. We summarize the factors that have made this possible, and outline likely future progress.

Aberration correction of the scanning transmission electron microscope (STEM) has made it possible to reach probe sizes close to 1 A at 60 keV, an operating energy that avoids direct knock-on damage in materials consisting of light atoms such as B, C, N and O. Although greatly reduced, some radiation damage is still present at this energy, and this limits the maximum usable electron dose. Elemental analysis by electron energy loss spectroscopy (EELS) is then usefully supplemented by annular dark field (ADF) imaging, for which the signal is larger. Because of its strong Z dependence, ADF allows the chemical identification of individual atoms, both heavy and light, and it can also record the atomic motion of individual heavy atoms in considerable detail. We illustrate these points by ADF images and EELS of nanotubes containing nanopods filled with single atoms of Er, and by ADF images of graphene with impurity atoms. (C) 2010 Elsevier B.V. All rights reserved.

An all-magnetic monochromator/spectrometer system for sub-30 meV energy-resolution electron energy-loss spectroscopy in the scanning transmission electron microscope is described. It will link the energy being selected by the monochromator to the energy being analysed by the spectrometer, without resorting to decelerating the electron beam. This will allow it to attain spectral energy stability comparable to systems using monochromators and spectrometers that are raised to near the high voltage of the instrument. It will also be able to correct the chromatic aberration of the probe-forming column. It should be able to provide variable energy resolution down to approximately 10 meV and spatial resolution less than 1 Å.

Aberration-corrected scanning transmission electron microscopes (STEMs) can now produce electron probes as small as 1 A at 60 keV. This level of performance allows individual light atoms to be imaged in various novel materials including graphene, monolayer boron nitride, and carbon nanotubes. Operation at 60 keV avoids direct knock-on damage in such materials, but some radiation damage often remains, and limits the maximum usable electron dose. Elemental identification by electron energy loss spectroscopy (EELS) is then usefully supplemented by annular dark-field (ADF) imaging, for which the signal is much larger and the spatial resolution significantly better. Because of its strong dependence on the atomic number Z, ADF can be used to identify the chemical type of individual atoms, both heavy and light. We review the instrumental requirements for atomic resolution imaging at 60 keV and lower energies, and we illustrate the kinds of studies that have now become possible by ADF images of graphene, monolayer BN, and single-wall carbon nanotubes, and by ADF images and EEL spectra of carbon nanotubes containing nanopods filled with single atoms of Er. We then discuss likely future developments.

Progress in aberration correction of electron microscopes has been rapid in the last decade. CEOS and Nion, the two companies chiefly responsible for the advent of practical aberration correctors, were started 12 and 11 years ago, respectively, at a time when aberration correction still seemed an impractical dream to many electron microscopists. The first sub-A resolution, directly interpretable aberration-corrected images were published in 2002 [1]. Today, there are more than 100 aberration-corrected electron microscopes in the world, and several more are installed each month. The resolution attained has reached 50 pm (0.5 A) in both STEM and TEM, and the electron current in an atom-sized electron probe can now be such that atomic-resolution STEM/EELS elemental maps can be acquired in less than one minute [2].

Improved resolution made possible by aberration correction has greatly increased the demands on the performance of all parts of high-end electron microscopes. In order to meet these demands, we have designed and built an entirely new scanning transmission electron microscope (STEM). The microscope includes a flexible illumination system that allows the properties of its probe to be changed on-the-fly, a third-generation aberration corrector which corrects all geometric aberrations up to fifth order, an ultra-responsive yet stable five-axis sample stage, and a flexible configuration of optimized detectors. The microscope features many innovations, such as a modular column assembled from building blocks that can be stacked in almost any order, in situ storage and cleaning facilities for up to five samples, computer-controlled loading of samples into the column, and self-diagnosing electronics. The microscope construction is described, and examples of its capabilities are shown.

Through the correction of spherical aberration in the scanning transmission electron microscope (STEM), the resolving of a 78 pm atomic column spacing has been demonstrated along with information transfer to 61 pm. The achievement of this resolution required careful control of microscope instabilities, parasitic aberrations and the compensation of uncorrected, higher order aberrations. Many of these issues are improved in a next generation STEM fitted with a new design of aberration corrector, and an initial result demonstrating aberration correction to a convergence semi-angle of 40 mrad is shown. The improved spatial resolution and beam convergence allowed for by such correction has implications for the way in which experiments are performed and how STEM data should be interpreted.

A dedicated STEM developed for operation at primary energies from 200 keV to 40 keV and lower is described. It has a new cold field emission gun (CFEG) that gives a normalized brightness of 3 × 10 8 A/(m 2 sr V), and excellent short-term and long-term stability. It includes two gun lenses (one electrostatic and one electromagnetic), a fast electrostatic beam blanker, three condenser lenses, a corrector of third- and fifth-order geometric aberrations, an objective lens with low aberration coefficients, a flexible set of projector lenses, an ultra-stable sample stage, and provision for storing up to five samples under high vacuum and loading them into the microscope’s objective lens under remote control. The microscope is enclosed in a magnetically and acoustically shielding enclosure, which allows it to operate at a high performance level even in non-optimal environments. It has reached 53 pm resolution at 200 keV and 123 pm at 40 keV, and an EELS energy resolution of 0.26 eV.

We compare different trajectories for C5-free C3 aberration correctors for scanning transmission electron microscopes, and find an asymmetric solution that minimizes the CC contributions and power supply stability requirements compared to a symmetric one. We further study the field aberrations of a C5-free quadrupole-octupole C3 aberration corrector and find that they are similar to those of sextupole C3-correctors in their suitability for wide-beam imaging applications. © 2008 Elsevier B.V. All rights reserved PACS: 41.81.Gy