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Scanning Electron Microscopy Under Gaseous Environment
Chapter
3
Scanning Electron Microscopy Under Gaseous Environment
Zahava Barkay Tel-Aviv University, Tel-Aviv, Israel
CHAPTER OUTLINE
3.1 3.2 3.3 3.4 3.5
Introduction 85 Basics of Electron Optics and Vacuum Conditions 87 ElectroneSpecimen Interaction, Generated Signals and Detectors 88 Resolution at the Various Vacuum Modes 90 Imaging at Various Vacuum Modes and Corresponding Applications 91 3.5.1 Imaging at High-Vacuum Mode 91 3.5.2 Imaging at Low-Vacuum Mode 92 3.5.3 Imaging at Wet Mode Using Reflected Secondary Electron Signal 93 3.5.4 Imaging at Wet Mode Using Transmitted Electron Signal
3.6 Benefits and Limitations of Imaging in Gaseous/Liquid Environment 3.7 Summary 100 Acknowledgments 100 References 101
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3.1 INTRODUCTION Variable-pressure scanning electron microscopy (VP-SEM) and environmental scanning electron microscopy (ESEM) differ from conventional scanning electron microscopy (SEM) because the sample can be viewed not just under high vacuum but also in a gaseous high-pressure environment. ESEM operates at an extended pressure relative to VP-SEM, but the principles are similar. VP-SEM and ESEM under low-vacuum conditions are suitable for imaging poor-conducting samples without grounding or preconductive coating. Microscopy Methods in Nanomaterials Characterization. http://dx.doi.org/10.1016/B978-0-323-46141-2.00003-1 Copyright © 2017 Elsevier Inc. All rights reserved.
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Low-vacuum imaging in materials sciences refers to large disciplines in which conductive coating is not desirable or sample postprocessing is required. It includes the fields of geology, optical devices, papers, plastics, ceramics, fabrics, composite materials, soft condensed matter (polymers, lithography resist patterns/structures), and various porous or outgassing materials. Research in biology at low vacuum includes imaging of uncoated dehydrated samples such as tissues, cells, inner-cell structures, drug encapsulation in cells, and immunolabeling. The applications include the fields of zoology, botanic, biotechnology, and medical devices such as nonconducting stents and implants. Low-vacuum imaging at nanoscale refers to bulk material surfaces as well as to nanomaterials such as nanoparticles (NPs) and carbon nanotubes (CNTs), which are frequently deposited on silicon dioxide or other nonconducting substrates. We will further focus on NPs and CNTs, which have major applications in biology, fuel catalytic reactions, electronic devices, and energy research fields. The ESEM, which operates at three vacuum modes, enables the investigation of (1) conductive samples at high vacuum, (2) nonconductive or highvacuum incompatible materials at low vacuum, and (3) wet samples. The research in ESEM is compatible for both biological and materials sciences by combining two main advantages: n
n
Nanometer resolution in multivacuum range: high-vacuum, low-vacuum, and wet modes. Real “wet” mode in the sense of 100% humidity in the specimen chamber.
Wet mode, often-named “environmental mode”, corresponds to biology and pathology research on hydrated cells and tissues as well as emulsions, cosmetics, oils, paints, gels, and food industries. In situ dynamic experiments in ESEM refer to wetting experiments, hydration and dehydration, corrosion, oxidation and reduction experiments, materials growth (tubes, wires, films, and coatings), gas-mediated reactions, and beaminduced chemical reactions. Several books have been published on SEM [1e3], VP-SEM, and ESEM [4], providing comprehensive overview on various instrumental and analytic aspects. In addition, a large variety of book chapters, review articles, journal articles, and internet media have been devoted to SEM research and development in the past 70 years. This chapter thus cannot replace that large amount of literature. It rather intends to highlight the relevance of SEM under gaseous environment for various applications in both biology and materials sciences. It further emphasizes the benefits and limitations of the various vacuum and imaging modes for nanoscale applications.
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The chapter is organized in sections, which refer to (1) basics of electron optics and vacuum conditions (Section 3.2); (2) electronespecimen interaction, generated signals, and detectors (Section 3.3); (3) resolution at the various vacuum modes (Section 3.4); (4) imaging at various vacuum modes and corresponding applications (Section 3.5); and (5) benefits and limitations of imaging in gaseous/liquid environments (Section 3.6) followed by summary (Section 3.7). Section 3.5 consists of subsections, which include imaging at high-vacuum, low-vacuum, and wet modes using the reflected secondary electron (SE) signal and the transmitted electron signal. All demonstrations in this chapter have been carried out using FEI Company (FEI) Quanta 200 FEG ESEM. However, the concepts are typical to imaging in gaseous environments using other systems and manufacturers of VP-SEM or low-vacuum SEM.
3.2 BASICS OF ELECTRON OPTICS AND VACUUM CONDITIONS All SEM systems consist [1e4] of an electron column with electron gun and magnetic lenses for e-beam generation, acceleration, and focusing. In most configurations, the electron column is at a vertical position, whereas the electron gun is at the top and the sample chamber is adjacent below the electron column. At the bottom of the column, a set of scan coils deflect the beam to generate a scanning electron beam (e-beam) on the sample. Electrone specimen interaction and signal generation take place at the sample chamber and the signals are collected by detectors above the sample or by inlens detectors, which provide the analysis of bulk samples. The SEM configuration differs from the transmission electron microscope (TEM) configuration, which requires thin samples that are usually mounted on a TEM grid. Unlike the high-vacuum SEM, which holds high vacuum through the column and sample chamber, the VP-SEM or ESEM use low vacuum at the sample chamber, whereas the e-beam column is maintained at high vacuum. The VP-SEM or ESEM configurations use differential pumping with a series of pressure-limiting apertures to maintain the pressure gradient through the system. The exact vacuum scheme depends on the SEM manufacturer. VP-SEM or ESEM are designed to operate optimally at the 10e30 keV e-beam acceleration energy range with detectors in the chamber, whereas new conventional high-vacuum SEM configurations provide high resolution also at lower energies (below 1 keV) and use inlens detectors. The ESEM configuration operates in the following three vacuum regimes: n
High vacuum (typically 105 torr) for imaging and microanalysis of conducting specimens or samples after conductive coating preparation
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n
n
Low vacuum (<1 torr) for imaging and microanalysis of nonconducting specimens without preparation ESEM mode (<10 torr, typically) for specimens that were previously impossible to investigate with high- or low-vacuum SEM modes. The pressure in this mode can in principle exceed 20 torr.
The SEM operation is mainly controlled by the e-beam probe current, e-beam acceleration voltage, and sample working distance (the distance between the lower pole piece in the column and the sample surface). In addition, lowvacuum mode requires the control of the chamber pressure, whereas for wet mode, both pressure and sample temperature are controlled.
3.3 ELECTRONeSPECIMEN INTERACTION, GENERATED SIGNALS AND DETECTORS Various signals are generated by interaction of an incident e-beam with the specimen (Fig. 3.1). The three main detected signals in most SEM configurations are SEs for surface characterization, X-rays for microanalysis, and backscattered electrons (BSEs) for atomic number contrast information. Additional signals such as cathodoluminescence are used in specific material applications, whereas the generated Auger electrons are measured in specific Auger analysis systems. Most SEM detectors (such as for SE, BSE, and X-ray signals) are positioned above the sample for imaging the reflected signals and thus provide imaging of bulk samples as well as nanoscale materials on various substrates. Low-dimensional structures such as thin films, CNTs, and NPs could be imaged by a scanning transmitted electron microscope (STEM) detector in SEM. The STEM detector is positioned beneath the sample for imaging
n FIGURE 3.1 Electronespecimen interactiondsignal generation in scanning electron microscopy (SEM).
3.3 ElectroneSpecimen Interaction, Generated Signals and Detectors 89
the transmitted electrons, whereas the nanosize materials are typically mounted on a TEM grid. The STEM detector provides bright field (BF) or dark field (DF) signals with high sensitivity to atomic number as well as to the material density and thickness (also named “mass-thickness”). The EverharteThornley (ET) detector is widely used in VP-SEM or ESEM for SE surface imaging at high-vacuum mode, but cannot function in gaseous environments. There are several approaches among the manufacturers for SE detection at low vacuum, for which the gas environment plays a central role. ESEM low-vacuum and wet modes use SE detectors, which are based [4] on gas ionization by the SEs. Under external bias of a few hundred volts, the generated electrons are accelerated and collected by the electrode, whereas the positive ions drift to the sample producing charge neutralization. The detector electrode and the sample can be considered as two plates of a capacitor, which under bias forms a cascade SE ionization process and finally provides the external detector current that is dependent on the sample SE yield. Increasing the gas pressure will improve the signal to a limit at which signal degradation occurs due to excessive incident e-beam scattering with the gas medium. The low-vacuum ESEM uses a large field detector (LFD) that can be mounted in the chamber opposite the EverharteThornley (ET) detector. Therefore, switching between high- and low-vacuum modes does not require any instrumental modifications during operation and simultaneous usage of SE, BSE, and X-ray signals. The BSE signal at high- and lowvacuum modes uses solid-state detector (SSD), which is usually mounted below the pole piece. The X-ray signal is usually [1] detected by a dedicated silicon drifted detector (SDD) or Si(Li) detector for energy dispersive spectroscopy microanalysis of element content, whereas wavelength-dispersive spectroscopy (WDS) analysis can be added as well. The ESEM wet mode utilizes an externally water-cooled Peltier stage instead of the ordinary sample stage. Both sample temperature and chamber pressure are controlled to achieve high levels of humidity. According to the equilibrium water-vapor phase diagram [4], the partial water pressure at room temperature is about 20 torr. Better imaging is achieved under lower pressures of typically 5.3e6.5 torr, which requires cooling the sample to 2e5 C. A patented gaseous secondary-electron detector (GSED), which is mounted below the pole piece in place of the standard SSD detector, is used for ESEM imaging. Additional detectors and accessories are used for in situ experiments [5] in ESEM. It includes heating stages, tensile [6] and indentation stages for in situ mechanical analysis, various manipulators, and electrical probe
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station [7]. Electron backscattered diffraction (EBSD) systems [8e9] use elastic backscattered electrons and transmission Kikuchi diffraction (TKD) configurations for texture information at micron and nanoscales. In situ experiments also refer to liquid phase electron beameinduced deposition [10] and wetting experiments as described in following sections.
3.4 RESOLUTION AT THE VARIOUS VACUUM MODES The VP-SEM or ESEM resolution, similarly to SEM, can be measured by the operator on the SE picture itself because the SE signal has the highest resolution among all SEM detected signals. The resolution is practically defined by the minimum distance, which can still be resolved, between two adjacent particles (usually gold particles on carbon) or sometimes by the particle edge width. The obtained resolution at all vacuum modes is thus a convolution between main e-beam spot size and SE mean free path. The SEM resolution is typically of the order of one to a few nanometers and can even reach subnanometers at certain conditions and specific SEM configurations. The STEM signal resolution of transmitted electrons depends on e-beam spot size as well as on beam broadening through the sample. In electron microscopy, the electron optics and sample chamber are usually kept at high vacuum to avoid interaction of gas molecules with the incident electron beam. At low-vacuum and wet modes, the gaseous medium in the sample chamber enhances primary e-beam scattering before landing on the sample. A basic question is how this affects the primary e-beam spot size and overall microscope resolution. The average number of elastic scattering events in the gas environment per incident electron is given by m ¼ L/lEL, when L is the gas path length (GPL) and lEL the elastic mean free path. According to Poisson distribution, the probability that an electron is not scattered obeys exp(-m). Ordinary high-vacuum conditions obey m < 0.05, which provides imaging with 95% unscattered electrons. Under lowvacuum and wet modes, m varies from 0.05 to 3 for imaging with 95% e5% unscattered electrons, whereas m > 3 does not provide imaging due to almost complete scattering. The values of L and lEL can be adjusted by the operator: the L value by controlling the GPL and lEL value according to the gas type, gas pressure, and e-beam acceleration voltage. As has been explained [5], although the primary beam at low vacuum and ESEM modes is partly scattered tens to thousands of microns away from its main trajectory, a significant number of incident electrons can remain in focus. The coming beam is thus composed of two components: the
3.5 Imaging at Various Vacuum Modes and Corresponding Applications 91
unscattered component, which remains in focus, and a scattered component providing a “skirt” effect. As long as the m value is minimized (by adjusting the GPL and lEL), the signal-to-noise (S/N) ratio of the unscattered component is kept high enough and the image resolution is similar to that of the high-vacuum mode.
3.5 IMAGING AT VARIOUS VACUUM MODES AND CORRESPONDING APPLICATIONS Possible ESEM applications for both biology and materials sciences have been outlined in the introduction. Examples for research at each vacuum mode are shown in the following subsections with emphasis on micronand nanoscale applications. The benefits and limitations for nanoscale imaging are discussed.
3.5.1 Imaging at High-Vacuum Mode SE imaging in ESEM is performed at high-vacuum mode in similarity to conventional high-vacuum SEM systems. SE imaging uses ET detector and is typically carried out at high voltages (of 10e20 KV) using conducting samples or conductive coated samples. A comparable SE and STEM characterization is recommended for nanomaterials or thin films on TEM grids, as has been shown [11] for drug-encapsulating micron-size particles. The following high-vacuum mode demonstration refers to energy storage technology and more specifically to electric-vehicle lithium-ion batteries [12]. Emphasis is given in this research to the production of alloy anodes of high capacitance and safety characteristics. We thus explain the characterization of the new anode materials, made of tin powder supported by multiwall carbon nanotubes (MWCNTs). The anode material is prepared on a TEM grid. We compare SE information using the ET detector with BF STEM information using the SSD in attempt to reveal the benefit of each imaging mode. The anode surface structure is shown at the SE image (Fig. 3.2A). The location of tin powder NPs within the MWCNTs matrix fits dark spots of tens of nanometers, which are provided by the atomic number contrast at the BF STEM image (Fig. 3.2B). The nonuniform background is due to thickness variations within the carbon matrix thus providing massethickness information. For comparison, dispersed MWCNTs are shown at the BF STEM image revealing NPs over MWCNTs, whereas the background is of uniform intensity (Fig. 3.2C). This high-vacuum SEM analysis has the benefit of scanning large anode areas and rapid image accumulation using simultaneously surface, atomic number, and massethickness information.
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n FIGURE 3.2 Nano-tin anode alloy for lithium battery: (A) secondary electron signal, (B)e(C) bright-field scanning transmitted electron microscope signal. Samples of Prof. E. Peled and Prof. D. Golodnitsky group.
3.5.2 Imaging at Low-Vacuum Mode Sample charging by incident e-beam is due to accumulation of net charge on the sample surface because of electronespecimen interaction. The net accumulated charge is due to nonzero balance between incoming and outgoing electrons. It is typical to nonconducting samples but appears also in ungrounded conducting samples and composite (conducting/ nonconducting) materials. The sample charge is either positive or negative (except for two crossover e-beam energies of zero net charge), which
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depends on conditions such as e-beam voltage, sample composition, and surface topography. Operation at high-acceleration voltages (typically above 5 kV) results in negative charge on nonconducting sample surfaces. Charging effects are frequently observed at the SE image. The removal of charge depends on e-beam dose, sample conductivity, and specific vacuum conditions in VP-SEM or ESEM. At high vacuum, the operation at specific crossover acceleration voltages can result in zero net charge for homogeneous smooth surface. However, in case that the nonconducting sample is of nonhomogeneous composition or of high roughness, charge neutralization cannot be achieved using a single acceleration voltage. Instead, VPSEM and ESEM provide charge neutralization from the entire sample surface by using leaking gas (typically air, nitrogen, or water vapor) into the sample chamber. Gas molecules, which are positively ionized by the SEs, compensate the negative charge on the sample surface. The following demonstration refers to imaging of Ni powder particles produced by pulsed-arc synthesis [13] between Ni electrodes submerged in pure ethanol. The pulsed-arc parameters control the particle size distribution. Powder samples were obtained by extracting liquid from the treatment vessel after a predetermined sedimentation and drying on a glass substrate. We compare high-vacuum mode with low-vacuum mode SE images for sample of Ni particles on glass substrate (Fig. 3.3). High-vacuum mode, at 10 KV acceleration voltage, results with negative-charge accumulation on sample surface. Image distortions appear as bright regions due to SE repulsion by the surface electric field and as scan distortions due to repulsion of main e-beam (Fig. 3.3A). Low vacuum of 100 Pa at 10 kV eliminates charging effects and provides charge neutralization over the entire sample surface (Fig. 3.3B). Higher magnification of the same sample provides imaging of submicron particles under 130Pa low-vacuum conditions (Fig. 3.3C).
3.5.3 Imaging at Wet Mode Using Reflected Secondary Electron Signal For imaging in wet environments two directions in electron microscopy (EM) have evolved: n n
Window technique Aperture-limited technique
In window technique, a pair of electron transparent windows is placed above and below the specimen allowing inlet- and outlet-side horizontal flow, while maintaining high vacuum outside the liquid cell device. It is usually designed [14] for TEM (or STEM) imaging using transmitted electrons of hundreds of kiloelectronvolts. Dedicated Wet-SEM capsules [15]
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n FIGURE 3.3 Ni particles on a glass substrate: (A) high-vacuum mode (BeC) low-vacuum mode. Samples of Prof. M. Boxman and Dr. N. Parkansky group.
have been produced for imaging wet materials in a high-vacuum SEM using BSEs typically above 10 keV. The alternative aperture-limited technique corresponds to environmental TEM (ETEM) or ESEM at wet mode, providing the benefit of an “open” liquid environment. The ESEM wet mode is of extended vacuum range relative to low-vacuum mode and, in addition, it has the advantage of “real” surface imaging by SEs. Wet samples and in particularly biological samples, can be imaged using water-vapor phase diagram equilibrium conditions. Thus, it reduces the rate of water evaporation from the sample, eliminates the need for dehydration step, facilitates the preparation procedure, and thus fastens clinical
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n FIGURE 3.4 Fungal spores: (A) low vacuum, (B) wet mode. Samples of Prof. N. Osherov group.
diagnosis. The detailed role of ESEM in cell biology has been reported elsewhere [16e17]. We here refer to ESEM imaging of fungal spores. Fungal infections [18] contribute substantially to human suffering and mortality. SE imaging at low-vacuum mode is compared with wet mode (Fig. 3.4). The sample has been air-dried before inspection at low-vacuum mode, showing shrinkage of surface structure (Fig. 3.4A). For wet mode, the fungal spores have been immersed in water and slowly in situ dehydrated in the ESEM until their surface structure is revealed (Fig. 3.4B). Imaging at 5 C and 6.4 torr provides their native surface spherical structure. A thin water layer on top of the surface is shown just before complete dehydration causing some dark contrast and blurring at Fig. 3.4B. It thus demonstrates the benefit of imaging biological samples at wet mode without any preparation or dehydration step. As explained in the next sections, further concern is required for eliminating radiation damage artifacts in images of biological and polymer materials under gaseous or liquid environments. ESEM wet mode is widely used for the study of various wettability aspects at the micron scale and nanoscale. It includes studying the triple-line structure [19], surface tension [20], line tension, and hydrophobic/hydrophilic [21] and rough surfaces [22e24]. The condensation and evaporation processes over the sample surface are provided by increasing or reducing the ESEM chamber pressure at constant sample temperature around dew point or alternatively by controlling the temperature at fixed pressure.
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n FIGURE 3.5 Wet mode: (A) Peltier cooling stage with liquid marble on top and GSED below the pole piece, (B) marble surface imaging in wet mode.
Samples of Prof. E. Bormashenko group.
The following demonstration of ESEM wet mode corresponds to the understanding of the mechanism of floating and rolling of liquid marbles [25]. In particular, it supports the assumption that the marble is isolated from the supporting surface by an air layer. The studied marbles are composed of polyvinylidene fluoride (PVDF) particles with average size of 130 nm encapsulating a 10-mL NaOH water drop (middle arrow at Fig. 3.5A). The sample is mounted on a Peltier cooling stage (bottom arrow at Fig. 3.5A), and is imaged at wet mode (2 C and 5.4 torr) using SE GSED (top arrow Fig. 3.5A). The ESEM image of part of the liquid marble surface shows that the PVDF particles do not form a monolithic layer, but are of hierarchical structure (Fig. 3.5B). The center of the image shows the water-clearing area (indicated by arrow at Fig. 3.5B). The hierarchical structure is typical to “lotus leaf” surfaces, known for their superhydrophobic properties. The absence of direct contact between the liquid and solid substrate is explained by the clearing areas surrounded by the particles’ hierarchical coating structure, which provides low-friction rolling and a nonstick air layer.
3.5.4 Imaging at Wet Mode Using Transmitted Electron Signal The wet scanning transmission electron microscope (wet-STEM) detector in ESEM has been established as a novel method for imaging nanoparticles in the liquid volume with a few nanometer resolution [26e27]. Furthermore, it has been utilized for measuring miniemulsions [28], biological samples [29],
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self-assembly NP dynamics [30], and water dynamics in carbon nanopipes [31]. Applications for wettability study at nanoscale are shown [32e36] for self-supported liquid films. The following demonstration refers to a quantitative wettability study [34] of nanodroplet nucleation and growth using wet-STEM. The wet-STEM mode uses Peltier cooling stage with SSD for imaging transmitted electrons. Thin self-supported water films have been produced inside the holes of a holey carbon TEM grid. Condensation and evaporation are provided by increasing or reducing the ESEM chamber pressure at constant sample temperature. Dynamic in situ imaging shows that irregularities at the water film boundaries constitute nucleation sites for both dropwise and filmwise condensation. Referring to dropwise condensation, the growth of two nanodroplets (signed as “1” and “2”) are studied under constant relative humidity (Fig. 3.6). Initial stages of nanodroplet growth are shown (Fig. 3.6A and B) for two selected time intervals (t ¼ 13s and t ¼ 27s). The power law dependence of droplet radius growth is shown as well (Fig. 3.6C). The fitting provides power law values of m ¼ 0.55 0.02 for the upper droplet (for “1”) and 0.30 0.02 for lower drop (for “2”). The distribution of power law values indicates the role of pinning centers for nanodroplet growth.
3.6 BENEFITS AND LIMITATIONS OF IMAGING IN GASEOUS/LIQUID ENVIRONMENT The SEM method provides a versatile tool due to its multiple advantages, which include relative ease of operation and wide application. In addition, the SEM provides wide lateral range for imaging, which scales from nanometer to centimeter (seven orders of magnitude) and a large depth of focus relative to optical microscopy. Simultaneous topography and energydispersiveewavelength-dispersive spectroscopy (EDS-WDS) chemical analysis is an important advantage for correlative SEM information. SEM can be used as a bulk method (using X-ray signal) as well as surface method (using SE signal) in dependence of the measured signal and acceleration voltage. The SEM provides a platform for e-beam-based systems such as dual-beam SEM-FIB (focused ion beam) for micro- and nanomachining applications. SEM is a nondestructive method for inorganic materials. It provides fast imaging and is thus suitable for in situ dynamic experiments. In situ dynamic imaging in VP-SEM or ESEM can be carried out under low-vacuum or wet environmental conditions, which extends both the material analysis methods and applications. There are possible several radiation-damage effects, which are particularly significant for organic or soft-matter materials. These include bond scission,
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n FIGURE 3.6 Wet-STEM: (A) t ¼ 13s, (B) t ¼ 27s, (C) dynamic droplet radius growth for two nanodroplets (signed as “1” and “2”).
cross-linking, mass loss, e-beam contamination, radiology, charging, and heating. E-beam heating has been reported for a few ESEM applications such as peptide nanotubes [37] and condensed micron-size droplets [38]. The solutions for the charging effects have been shown and discussed in previous sections. We mainly concentrate on two sources of radiation damage under a water-vapor environment: e-beam contamination and e-beam radiology. E-beam contamination occurs in electron microscopy due to carbonization of the sample surface at the e-beam-irradiated area because of sample surface contaminates or residual vacuum hydrocarbons. It is typically observed as darkening of the irradiated area under conventional SEM vacuum conditions
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of 105 torr. Contamination in gaseous ESEM environments is considered less significant than under conventional high-vacuum conditions. Radiology is a result of inelastic e-beam scattering in liquid samples or vapor environment, which forms ionized and excited water molecular states. It thus refers to both “window technique” (using liquid cells or capsules) and “aperture limited technique” (ESEM and ETEM). The excited water-vapor molecules can decay into free radicals or ions. The high yield of the hydroxyl free radical OH suggests that it dominates the radiology effect leading to degradation of polymer surface at low-vacuum ESEM. The effect is even more significant while in situ ESEM studying liquids over polymers or hydrated biological samples. Radiology effects due to electronewater interactions have recently been reported [39] for liquid-cell electron microscopy. As has been explained, organic materials such as polymers without conductive coating are sensitive to e-beam damage. A typical demonstration is shown at SE images of Fig. 3.7. Polystyrene NPs, of 100-nm diameter, at high vacuum with conductive Au/Pd coating (Fig. 3.7A) are compared with the uncoated sample at low vacuum of 100 Pa (Fig. 3.7B). Degradation is observed for the uncoated polystyrene NPs at low vacuum during a few minutes of e-beam imaging. As has been explained elsewhere [40e41], direct exposure of the sample to e-beam results in material mass loss, whereas additional free radicals (due to e-beam interaction with the vapor) further lead to radiolysis and oxidation of the polymer. Nanoscale imaging requires high magnifications, which increase the e-beam dose and thus heating, mass loss, and radiology effects become dominant.
n FIGURE 3.7 Polystyrene particles (A) coated sample at high-vacuum mode, (B) uncoated sample at low-vacuum mode.
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For soft-matter nanotechnology, the elimination of radiation damage during ESEM research is thus an important task for polymer, organic materials, and liquid samples.
3.7 SUMMARY The VP-SEM and ESEM provide high-resolution imaging at all vacuum modes. These methods are thus used in nanoscience research for broad disciplines and applications. In particular, SEM imaging under gaseous conditions is applicable for low-dimensional structures including NPs, CNTs, and nanodroplets. As is shown, the choice of exact vacuum conditions depends on sample properties. The benefits of multi SEM imaging modes are explicitly shown for both surface and transmission information. Characterization of chemical bonding at micron and nano-scales is important as well for image interpretation and for studying the properties of in situ processed materials under gaseous environments. Thus, besides the known traditional EDS and WDS methods, additional tools will be required in the coming years for nanoscale analysis. Theoretical understanding of radiation damage for each type of studied material is important to eliminate these effects at micron- and nanoscale levels. Future progress in instrumentation is expected to provide lower dose imaging to overcome the mentioned limitations of e-beam radiation damage in organic and biological materials. New detectors and SEM configurations in the coming decade are expected to improve image resolution and S/N ratio under gaseous environments. Further attempts may also be done to carry experiments closer to “real” environmental conditions, i.e., at room temperatures and pressures [42e43]. In addition, it has been shown [44] that better control of environmental conditions (both humidity and temperature) has to be done by adding sensors at the sample position. The availability of in situ probe stations and dedicated sample stages will expand the amount of in situ dynamic experiments in the following years. Therefore, besides lateral resolution, the issue of high temporal resolution is going to play an important role. The nanotechnology research under lowdose imaging, high lateral and temporal resolutions, and high S/N ratio will thus benefit from VP-SEM and ESEM progress in the coming years.
ACKNOWLEDGMENTS Acknowledgment is given to the following researchers at Tel-Aviv University (TAU) for their fruitful collaboration and permission to publish selected pictures:
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Prof. E. Peled and Prof. D. Golodnitsky from the chemical department at TAU, Dr. N. Parkansky and Prof. R. L. Boxman from electrical discharge and plasma laboratory at TAU, and Prof. N. Osherov from microbiology and immunology, school of medicine at TAU. Acknowledgment is given to Prof. E. Bormashenko from Ariel University for his collaboration and image contribution. In addition, the author thanks the Wolfson Applied Materials Research Center (WAMRC) and the center of Nanoscience and Nanotechnology at TAU for supporting the ESEM laboratory.
REFERENCES [1] J. Goldstein, D. Newbury, D. Joy, C. Lyman, P. Echlin, E. Lifshin, L. Sawyer, J. Michael, Scanning Electron Microscopy and X-ray Microanalysis, Plenum, NY, 2003. [2] L. Riemer, Scanning electron microscopy, Springer Ser. Opt. Sci. 45 (1984). [3] L. Reimer, Image Formation in Low-voltage Scanning Electron Microscopy, 1993. Bellingham, WA. [4] D.J. Stokes, Principles and Practice of Variable Pressure/Environmental Scanning Electron Microscopy, Wiley Press, West Sussex, 2008. [5] R. Podor, J. Ravaux, H.P. Brau, In situ experiments in the scanning electron microscope chamber, in: V. Kazmiruk (Ed.), Scanning Electron Microscopy, InTech, Europe, 2012, pp. 31e54. [6] S. Ifergane, Z. Barkay, O. Beeri, N. Eliaz, Study of fracture evolution in copper sheets by in situ tensile test and EBSD analysis, J. Mater. Sci. 45 (2010) 6345e6352. [7] D. Azulai, T. Belenkova, H. Gilon, Z. Barkay, G. Markovich, Transparent metal nanowire thin films prepared in mesostructured templates, Nano Lett. 9 (2009) 4246e4249. [8] V. Randle, Microtexture Determination and Its Applications, Maney Publishing, 2003. [9] Z. Barkay, E. Grunbaum, G. Leitus, S. Reich, Study of the superconducting Cs-doped WO3 crystal surface by electron backscattered diffraction, J. Supercond. Novel Magn. 21 (2008) 145e150. [10] M. Bresin, A. Botman, S.J. Randolph, M. Straw, J.T. Hastings, Liquid phase electron induced deposition on bulk substrates using environmental scanning electron microscopy, Microsc. Microanal. 20 (2014) 376e384. [11] Z. Barkay, I. Rivkin, R. Margalit, Three dimensional characterization of drugencapsulating particles using STEM detector in FEG-SEM, Micron 40 (2009) 480e485. [12] S. Menkin, Z. Barkay, D. Golodnitsky, E. Peled, Nanotin alloys supported by multiwall carbon nanotubes as high-capacity and safer anode materials for EV lithium batteries, J. Power Sources 245 (2014) 345e351. [13] N. Parkansky, N. Frenkel, G. Alterkop, R.L. Boxman, S. Goldsmith, Z. Barkay, Y. Rosenberg, O. Goldstein, Influence of pulsed arc parameters on powder production in ethanol, Powder Technol. 162 (2006) 121e125. [14] J.M. Gogan, L. Rotkina, H. Bau, In situ liquid-cell electron microscopy of colloid aggregation and growth dynamics, Phys. Rev. E 83 (2011) 1e5, 061405.
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[15] S. Thiberge, A. Nechushtan, D. Sprinzak, O. Gileadi, V. Behar, O. Zik, Y. Chowers, S. Michaeli, J. Schlessinger, E. Moses, Scanning electron microscopy of cells and tissues under fully hydrated conditions, PNAS 101 (2004) 3346e3351. [16] J.E. McGregor, L.T.L. Staniewicz, S.E. Guthrie, A.M. Donald, Imaging Techniques: Methods and Protocols, Methods in Molecular Biology, Springer, 2013 (Chapter 26). [17] S.E. Kirk, J.N. Skepper, A.M. Donald, Applications of environmental scanning electron microscopy to determine biological surface structure, J. Microsc. 233 (2009) 205e224. [18] N. Osherov, Modulation of host-cell MAP-kinase signaling during fungal infection, MAP Kinase 4 (2009) 5389. [19] E. Bormashenko, A. Musin, G. Whyman, Z. Barkay, M. Zinigrad, Revisiting the fine structure of the triple line, Langmuir 29 (2013) 14163e14167. [20] E. Bormashenko, A. Musina, G. Whymana, Z. Barkay, A. Starostinc, V. Valtsiferc, V. Strelnikovc, Revisiting the surface tension of liquid marbles: measurement of the effective surface tension of liquid marbles with the pendant marble method, Colloids Surf., A 425 (2013) 15e23. [21] D. Aronov, G. Rosenman, Z. Barkay, Wettability study of modified silicon oxide surface using environmental scanning electron microscope, J. Appl. Phys. 101 (2007) 84901e84905. [22] M. Nosonovsky, B. Bhushan, Biomimetic superhydrophobic surfaces: multiscale approach, Nano Lett. 7 (2007) 2633e2637. [23] K.J. Rykaczewski, J.H. Scott, Methodology for imaging nano-to-microscale water condensation dynamics on complex nanostructures, ACS Nano 5 (2011) 5962e5968. [24] Z. Barkay, R. Golombick, Y. Gabay, Y. Benayahu, Droplet contact angle measurement on microstructures of octocoral sclerites-ESEM image analysis, JCCE 1 (2016) 42e47. [25] E. Bormashenko, Y. Bormashenko, A. Musin, Z. Barkay, On the mechanism of floating and sliding of liquid marbles, Chemphyschem 10 (2009) 654e656. [26] A. Bogner, P.H. Jouneau, G. Thollet, D. Basset, C. Gauthier, A history of scanning electron microscopy developments: towards “wet-STEM” imaging, Micron 38 (2007) 390e401. [27] A. Bogner, G. Thollet, D. Basset, P.H. Jouneau, C. Gauthier, Wet STEM: a new development in environmental SEM for imaging nano-objects included in a liquid phase, Ultramicroscopy 104 (2005) 290e301. [28] M. Amaral, A. Borger, C. Gauthie, G. Thollet, P.H. Jouneau, J.Y. Cavaille, J.M. Asua, Novel experimental technique for the determination of monomer droplet size distribution in miniemulsion, Macromol. Rapid Commun. 26 (2005) 365e368. [29] D.B. Peckys, N. de Jonge, Liquid scanning transmission electron microscopy: imaging protein complexes in their native environment in whole eukaryotic cells, Microsc. Microanal. 20 (2014) 346e365. [30] F. Novotny, P. Wandrol, J. Proska, M. Slouf, In situ WetSTEM observations of gold nanorod self-assembly dynamics in a drying colloidal droplet, Microsc. Microanal. 20 (2014) 385e393. [31] M. Pia Rossi, Y. Haihui, Y. Gogotsi, S. Babu, P. Ndungu, J.C. Bradley, Environmental scanning electron microscopy study of water in carbon nanopipes, Nano Lett. 4 (2004) 989e993.
References 103
[32] Z. Barkay, Dynamic study of nanodroplet nucleation and growth using transmitted electrons in ESEM, in: Z.M. Wang, A. Waag, G. Salamo, N. Kishimoto, S. Bellucci, Y.J. Park (Eds.), Lecture Notes in Nanoscale Science and Technology, Nanodroplets, vol. 18, Springer, 2013, pp. 51e72. [33] Z. Barkay, Wettability study using transmitted electrons in environmental scanning electron microscope, Appl. Phys. Lett. 96 (2010) 1e3, 183109. [34] Z. Barkay, Dynamic study of nanodroplet nucleation and growth on self-supported nano-thick liquid films, Langmuir 26 (2010) 18581e18584. [35] Z. Barkay, Quantitative wettability study at nanometer scale based on wet-STEM in ESEM, Microsc. Microanal. 18 (2010) 1134e1135. [36] Z. Barkay, In situ imaging of nanodroplet condensation and coalescence on thin liquid films, Microsc. Microanal. 20 (2014) 317e322. [37] N. Amdursky, P. Beker, I. Koren, B. Bank-Srour, E. Mishina, S. Semin, T. Rasing, Y. Rosenberg, Z. Barkay, E. Gazit, G. Rosenman, Structural transition in peptide nanotubes, Biomacromolecules 12 (2011) 1349e1354. [38] K. Rykaczewski, J.H.J. Scott, A.G. Fedorov, Electron beam heating effects during environmental scanning electron microscopy imaging of water condensation on superhydrophobic surfaces, Appl. Phys. Lett. 98 (2011) 1e3, 093106. [39] N.M. Schneider, M.M. Norton, B.J. Mendel, J.M. Grogan, F.M. Ross, H.H. Bau, Electron-water interactions and implications for liquid cell electron microscopy, J. Phys. Chem., C 118 (2014) 22373e22382. [40] S. Kitching, A.M. Donald, Beam damage of polypropylene in environmental scanning electron microscope: an FTIR study, J. Microsc. 190 (1998) 357e365. [41] C.P. Royall, B.L. Thiel, A.M. Donald, Radiation damage of water in environmental scanning electron microscope, J. Microsc. 204 (2001) 185e195. [42] M. Toth, M. Uncovsky, R. Knowles, F.S. Baker, Secondary electron imaging at gas pressures in excess of 1 kPa, Appl. Phys. Lett. 91 (2007) 1e3, 053122. [43] I. Solomonov, D. Talmi-Frank, Y. Milstein, S. Addadi, A. Aloshin, I. Safi, Introduction of correlative light and airSEM microscopy imaging for tissue research under ambient conditions, Sci. Rep. 4 (5987) (2014) 1e7. [44] R. Leart, R. Brydson, Characterization of ESEM conditions for specimen hydration control, J. Phys. Conf. Ser. 241 (2010) 1e4, 012024.
3
Scanning Electron Microscopy Under Gaseous Environment
Zahava Barkay Tel-Aviv University, Tel-Aviv, Israel
CHAPTER OUTLINE
3.1 3.2 3.3 3.4 3.5
Introduction 85 Basics of Electron Optics and Vacuum Conditions 87 ElectroneSpecimen Interaction, Generated Signals and Detectors 88 Resolution at the Various Vacuum Modes 90 Imaging at Various Vacuum Modes and Corresponding Applications 91 3.5.1 Imaging at High-Vacuum Mode 91 3.5.2 Imaging at Low-Vacuum Mode 92 3.5.3 Imaging at Wet Mode Using Reflected Secondary Electron Signal 93 3.5.4 Imaging at Wet Mode Using Transmitted Electron Signal
3.6 Benefits and Limitations of Imaging in Gaseous/Liquid Environment 3.7 Summary 100 Acknowledgments 100 References 101
96
97
3.1 INTRODUCTION Variable-pressure scanning electron microscopy (VP-SEM) and environmental scanning electron microscopy (ESEM) differ from conventional scanning electron microscopy (SEM) because the sample can be viewed not just under high vacuum but also in a gaseous high-pressure environment. ESEM operates at an extended pressure relative to VP-SEM, but the principles are similar. VP-SEM and ESEM under low-vacuum conditions are suitable for imaging poor-conducting samples without grounding or preconductive coating. Microscopy Methods in Nanomaterials Characterization. http://dx.doi.org/10.1016/B978-0-323-46141-2.00003-1 Copyright © 2017 Elsevier Inc. All rights reserved.
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Low-vacuum imaging in materials sciences refers to large disciplines in which conductive coating is not desirable or sample postprocessing is required. It includes the fields of geology, optical devices, papers, plastics, ceramics, fabrics, composite materials, soft condensed matter (polymers, lithography resist patterns/structures), and various porous or outgassing materials. Research in biology at low vacuum includes imaging of uncoated dehydrated samples such as tissues, cells, inner-cell structures, drug encapsulation in cells, and immunolabeling. The applications include the fields of zoology, botanic, biotechnology, and medical devices such as nonconducting stents and implants. Low-vacuum imaging at nanoscale refers to bulk material surfaces as well as to nanomaterials such as nanoparticles (NPs) and carbon nanotubes (CNTs), which are frequently deposited on silicon dioxide or other nonconducting substrates. We will further focus on NPs and CNTs, which have major applications in biology, fuel catalytic reactions, electronic devices, and energy research fields. The ESEM, which operates at three vacuum modes, enables the investigation of (1) conductive samples at high vacuum, (2) nonconductive or highvacuum incompatible materials at low vacuum, and (3) wet samples. The research in ESEM is compatible for both biological and materials sciences by combining two main advantages: n
n
Nanometer resolution in multivacuum range: high-vacuum, low-vacuum, and wet modes. Real “wet” mode in the sense of 100% humidity in the specimen chamber.
Wet mode, often-named “environmental mode”, corresponds to biology and pathology research on hydrated cells and tissues as well as emulsions, cosmetics, oils, paints, gels, and food industries. In situ dynamic experiments in ESEM refer to wetting experiments, hydration and dehydration, corrosion, oxidation and reduction experiments, materials growth (tubes, wires, films, and coatings), gas-mediated reactions, and beaminduced chemical reactions. Several books have been published on SEM [1e3], VP-SEM, and ESEM [4], providing comprehensive overview on various instrumental and analytic aspects. In addition, a large variety of book chapters, review articles, journal articles, and internet media have been devoted to SEM research and development in the past 70 years. This chapter thus cannot replace that large amount of literature. It rather intends to highlight the relevance of SEM under gaseous environment for various applications in both biology and materials sciences. It further emphasizes the benefits and limitations of the various vacuum and imaging modes for nanoscale applications.
3.2 Basics of Electron Optics and Vacuum Conditions 87
The chapter is organized in sections, which refer to (1) basics of electron optics and vacuum conditions (Section 3.2); (2) electronespecimen interaction, generated signals, and detectors (Section 3.3); (3) resolution at the various vacuum modes (Section 3.4); (4) imaging at various vacuum modes and corresponding applications (Section 3.5); and (5) benefits and limitations of imaging in gaseous/liquid environments (Section 3.6) followed by summary (Section 3.7). Section 3.5 consists of subsections, which include imaging at high-vacuum, low-vacuum, and wet modes using the reflected secondary electron (SE) signal and the transmitted electron signal. All demonstrations in this chapter have been carried out using FEI Company (FEI) Quanta 200 FEG ESEM. However, the concepts are typical to imaging in gaseous environments using other systems and manufacturers of VP-SEM or low-vacuum SEM.
3.2 BASICS OF ELECTRON OPTICS AND VACUUM CONDITIONS All SEM systems consist [1e4] of an electron column with electron gun and magnetic lenses for e-beam generation, acceleration, and focusing. In most configurations, the electron column is at a vertical position, whereas the electron gun is at the top and the sample chamber is adjacent below the electron column. At the bottom of the column, a set of scan coils deflect the beam to generate a scanning electron beam (e-beam) on the sample. Electrone specimen interaction and signal generation take place at the sample chamber and the signals are collected by detectors above the sample or by inlens detectors, which provide the analysis of bulk samples. The SEM configuration differs from the transmission electron microscope (TEM) configuration, which requires thin samples that are usually mounted on a TEM grid. Unlike the high-vacuum SEM, which holds high vacuum through the column and sample chamber, the VP-SEM or ESEM use low vacuum at the sample chamber, whereas the e-beam column is maintained at high vacuum. The VP-SEM or ESEM configurations use differential pumping with a series of pressure-limiting apertures to maintain the pressure gradient through the system. The exact vacuum scheme depends on the SEM manufacturer. VP-SEM or ESEM are designed to operate optimally at the 10e30 keV e-beam acceleration energy range with detectors in the chamber, whereas new conventional high-vacuum SEM configurations provide high resolution also at lower energies (below 1 keV) and use inlens detectors. The ESEM configuration operates in the following three vacuum regimes: n
High vacuum (typically 105 torr) for imaging and microanalysis of conducting specimens or samples after conductive coating preparation
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n
n
Low vacuum (<1 torr) for imaging and microanalysis of nonconducting specimens without preparation ESEM mode (<10 torr, typically) for specimens that were previously impossible to investigate with high- or low-vacuum SEM modes. The pressure in this mode can in principle exceed 20 torr.
The SEM operation is mainly controlled by the e-beam probe current, e-beam acceleration voltage, and sample working distance (the distance between the lower pole piece in the column and the sample surface). In addition, lowvacuum mode requires the control of the chamber pressure, whereas for wet mode, both pressure and sample temperature are controlled.
3.3 ELECTRONeSPECIMEN INTERACTION, GENERATED SIGNALS AND DETECTORS Various signals are generated by interaction of an incident e-beam with the specimen (Fig. 3.1). The three main detected signals in most SEM configurations are SEs for surface characterization, X-rays for microanalysis, and backscattered electrons (BSEs) for atomic number contrast information. Additional signals such as cathodoluminescence are used in specific material applications, whereas the generated Auger electrons are measured in specific Auger analysis systems. Most SEM detectors (such as for SE, BSE, and X-ray signals) are positioned above the sample for imaging the reflected signals and thus provide imaging of bulk samples as well as nanoscale materials on various substrates. Low-dimensional structures such as thin films, CNTs, and NPs could be imaged by a scanning transmitted electron microscope (STEM) detector in SEM. The STEM detector is positioned beneath the sample for imaging
n FIGURE 3.1 Electronespecimen interactiondsignal generation in scanning electron microscopy (SEM).
3.3 ElectroneSpecimen Interaction, Generated Signals and Detectors 89
the transmitted electrons, whereas the nanosize materials are typically mounted on a TEM grid. The STEM detector provides bright field (BF) or dark field (DF) signals with high sensitivity to atomic number as well as to the material density and thickness (also named “mass-thickness”). The EverharteThornley (ET) detector is widely used in VP-SEM or ESEM for SE surface imaging at high-vacuum mode, but cannot function in gaseous environments. There are several approaches among the manufacturers for SE detection at low vacuum, for which the gas environment plays a central role. ESEM low-vacuum and wet modes use SE detectors, which are based [4] on gas ionization by the SEs. Under external bias of a few hundred volts, the generated electrons are accelerated and collected by the electrode, whereas the positive ions drift to the sample producing charge neutralization. The detector electrode and the sample can be considered as two plates of a capacitor, which under bias forms a cascade SE ionization process and finally provides the external detector current that is dependent on the sample SE yield. Increasing the gas pressure will improve the signal to a limit at which signal degradation occurs due to excessive incident e-beam scattering with the gas medium. The low-vacuum ESEM uses a large field detector (LFD) that can be mounted in the chamber opposite the EverharteThornley (ET) detector. Therefore, switching between high- and low-vacuum modes does not require any instrumental modifications during operation and simultaneous usage of SE, BSE, and X-ray signals. The BSE signal at high- and lowvacuum modes uses solid-state detector (SSD), which is usually mounted below the pole piece. The X-ray signal is usually [1] detected by a dedicated silicon drifted detector (SDD) or Si(Li) detector for energy dispersive spectroscopy microanalysis of element content, whereas wavelength-dispersive spectroscopy (WDS) analysis can be added as well. The ESEM wet mode utilizes an externally water-cooled Peltier stage instead of the ordinary sample stage. Both sample temperature and chamber pressure are controlled to achieve high levels of humidity. According to the equilibrium water-vapor phase diagram [4], the partial water pressure at room temperature is about 20 torr. Better imaging is achieved under lower pressures of typically 5.3e6.5 torr, which requires cooling the sample to 2e5 C. A patented gaseous secondary-electron detector (GSED), which is mounted below the pole piece in place of the standard SSD detector, is used for ESEM imaging. Additional detectors and accessories are used for in situ experiments [5] in ESEM. It includes heating stages, tensile [6] and indentation stages for in situ mechanical analysis, various manipulators, and electrical probe
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station [7]. Electron backscattered diffraction (EBSD) systems [8e9] use elastic backscattered electrons and transmission Kikuchi diffraction (TKD) configurations for texture information at micron and nanoscales. In situ experiments also refer to liquid phase electron beameinduced deposition [10] and wetting experiments as described in following sections.
3.4 RESOLUTION AT THE VARIOUS VACUUM MODES The VP-SEM or ESEM resolution, similarly to SEM, can be measured by the operator on the SE picture itself because the SE signal has the highest resolution among all SEM detected signals. The resolution is practically defined by the minimum distance, which can still be resolved, between two adjacent particles (usually gold particles on carbon) or sometimes by the particle edge width. The obtained resolution at all vacuum modes is thus a convolution between main e-beam spot size and SE mean free path. The SEM resolution is typically of the order of one to a few nanometers and can even reach subnanometers at certain conditions and specific SEM configurations. The STEM signal resolution of transmitted electrons depends on e-beam spot size as well as on beam broadening through the sample. In electron microscopy, the electron optics and sample chamber are usually kept at high vacuum to avoid interaction of gas molecules with the incident electron beam. At low-vacuum and wet modes, the gaseous medium in the sample chamber enhances primary e-beam scattering before landing on the sample. A basic question is how this affects the primary e-beam spot size and overall microscope resolution. The average number of elastic scattering events in the gas environment per incident electron is given by m ¼ L/lEL, when L is the gas path length (GPL) and lEL the elastic mean free path. According to Poisson distribution, the probability that an electron is not scattered obeys exp(-m). Ordinary high-vacuum conditions obey m < 0.05, which provides imaging with 95% unscattered electrons. Under lowvacuum and wet modes, m varies from 0.05 to 3 for imaging with 95% e5% unscattered electrons, whereas m > 3 does not provide imaging due to almost complete scattering. The values of L and lEL can be adjusted by the operator: the L value by controlling the GPL and lEL value according to the gas type, gas pressure, and e-beam acceleration voltage. As has been explained [5], although the primary beam at low vacuum and ESEM modes is partly scattered tens to thousands of microns away from its main trajectory, a significant number of incident electrons can remain in focus. The coming beam is thus composed of two components: the
3.5 Imaging at Various Vacuum Modes and Corresponding Applications 91
unscattered component, which remains in focus, and a scattered component providing a “skirt” effect. As long as the m value is minimized (by adjusting the GPL and lEL), the signal-to-noise (S/N) ratio of the unscattered component is kept high enough and the image resolution is similar to that of the high-vacuum mode.
3.5 IMAGING AT VARIOUS VACUUM MODES AND CORRESPONDING APPLICATIONS Possible ESEM applications for both biology and materials sciences have been outlined in the introduction. Examples for research at each vacuum mode are shown in the following subsections with emphasis on micronand nanoscale applications. The benefits and limitations for nanoscale imaging are discussed.
3.5.1 Imaging at High-Vacuum Mode SE imaging in ESEM is performed at high-vacuum mode in similarity to conventional high-vacuum SEM systems. SE imaging uses ET detector and is typically carried out at high voltages (of 10e20 KV) using conducting samples or conductive coated samples. A comparable SE and STEM characterization is recommended for nanomaterials or thin films on TEM grids, as has been shown [11] for drug-encapsulating micron-size particles. The following high-vacuum mode demonstration refers to energy storage technology and more specifically to electric-vehicle lithium-ion batteries [12]. Emphasis is given in this research to the production of alloy anodes of high capacitance and safety characteristics. We thus explain the characterization of the new anode materials, made of tin powder supported by multiwall carbon nanotubes (MWCNTs). The anode material is prepared on a TEM grid. We compare SE information using the ET detector with BF STEM information using the SSD in attempt to reveal the benefit of each imaging mode. The anode surface structure is shown at the SE image (Fig. 3.2A). The location of tin powder NPs within the MWCNTs matrix fits dark spots of tens of nanometers, which are provided by the atomic number contrast at the BF STEM image (Fig. 3.2B). The nonuniform background is due to thickness variations within the carbon matrix thus providing massethickness information. For comparison, dispersed MWCNTs are shown at the BF STEM image revealing NPs over MWCNTs, whereas the background is of uniform intensity (Fig. 3.2C). This high-vacuum SEM analysis has the benefit of scanning large anode areas and rapid image accumulation using simultaneously surface, atomic number, and massethickness information.
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n FIGURE 3.2 Nano-tin anode alloy for lithium battery: (A) secondary electron signal, (B)e(C) bright-field scanning transmitted electron microscope signal. Samples of Prof. E. Peled and Prof. D. Golodnitsky group.
3.5.2 Imaging at Low-Vacuum Mode Sample charging by incident e-beam is due to accumulation of net charge on the sample surface because of electronespecimen interaction. The net accumulated charge is due to nonzero balance between incoming and outgoing electrons. It is typical to nonconducting samples but appears also in ungrounded conducting samples and composite (conducting/ nonconducting) materials. The sample charge is either positive or negative (except for two crossover e-beam energies of zero net charge), which
3.5 Imaging at Various Vacuum Modes and Corresponding Applications 93
depends on conditions such as e-beam voltage, sample composition, and surface topography. Operation at high-acceleration voltages (typically above 5 kV) results in negative charge on nonconducting sample surfaces. Charging effects are frequently observed at the SE image. The removal of charge depends on e-beam dose, sample conductivity, and specific vacuum conditions in VP-SEM or ESEM. At high vacuum, the operation at specific crossover acceleration voltages can result in zero net charge for homogeneous smooth surface. However, in case that the nonconducting sample is of nonhomogeneous composition or of high roughness, charge neutralization cannot be achieved using a single acceleration voltage. Instead, VPSEM and ESEM provide charge neutralization from the entire sample surface by using leaking gas (typically air, nitrogen, or water vapor) into the sample chamber. Gas molecules, which are positively ionized by the SEs, compensate the negative charge on the sample surface. The following demonstration refers to imaging of Ni powder particles produced by pulsed-arc synthesis [13] between Ni electrodes submerged in pure ethanol. The pulsed-arc parameters control the particle size distribution. Powder samples were obtained by extracting liquid from the treatment vessel after a predetermined sedimentation and drying on a glass substrate. We compare high-vacuum mode with low-vacuum mode SE images for sample of Ni particles on glass substrate (Fig. 3.3). High-vacuum mode, at 10 KV acceleration voltage, results with negative-charge accumulation on sample surface. Image distortions appear as bright regions due to SE repulsion by the surface electric field and as scan distortions due to repulsion of main e-beam (Fig. 3.3A). Low vacuum of 100 Pa at 10 kV eliminates charging effects and provides charge neutralization over the entire sample surface (Fig. 3.3B). Higher magnification of the same sample provides imaging of submicron particles under 130Pa low-vacuum conditions (Fig. 3.3C).
3.5.3 Imaging at Wet Mode Using Reflected Secondary Electron Signal For imaging in wet environments two directions in electron microscopy (EM) have evolved: n n
Window technique Aperture-limited technique
In window technique, a pair of electron transparent windows is placed above and below the specimen allowing inlet- and outlet-side horizontal flow, while maintaining high vacuum outside the liquid cell device. It is usually designed [14] for TEM (or STEM) imaging using transmitted electrons of hundreds of kiloelectronvolts. Dedicated Wet-SEM capsules [15]
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n FIGURE 3.3 Ni particles on a glass substrate: (A) high-vacuum mode (BeC) low-vacuum mode. Samples of Prof. M. Boxman and Dr. N. Parkansky group.
have been produced for imaging wet materials in a high-vacuum SEM using BSEs typically above 10 keV. The alternative aperture-limited technique corresponds to environmental TEM (ETEM) or ESEM at wet mode, providing the benefit of an “open” liquid environment. The ESEM wet mode is of extended vacuum range relative to low-vacuum mode and, in addition, it has the advantage of “real” surface imaging by SEs. Wet samples and in particularly biological samples, can be imaged using water-vapor phase diagram equilibrium conditions. Thus, it reduces the rate of water evaporation from the sample, eliminates the need for dehydration step, facilitates the preparation procedure, and thus fastens clinical
3.5 Imaging at Various Vacuum Modes and Corresponding Applications 95
n FIGURE 3.4 Fungal spores: (A) low vacuum, (B) wet mode. Samples of Prof. N. Osherov group.
diagnosis. The detailed role of ESEM in cell biology has been reported elsewhere [16e17]. We here refer to ESEM imaging of fungal spores. Fungal infections [18] contribute substantially to human suffering and mortality. SE imaging at low-vacuum mode is compared with wet mode (Fig. 3.4). The sample has been air-dried before inspection at low-vacuum mode, showing shrinkage of surface structure (Fig. 3.4A). For wet mode, the fungal spores have been immersed in water and slowly in situ dehydrated in the ESEM until their surface structure is revealed (Fig. 3.4B). Imaging at 5 C and 6.4 torr provides their native surface spherical structure. A thin water layer on top of the surface is shown just before complete dehydration causing some dark contrast and blurring at Fig. 3.4B. It thus demonstrates the benefit of imaging biological samples at wet mode without any preparation or dehydration step. As explained in the next sections, further concern is required for eliminating radiation damage artifacts in images of biological and polymer materials under gaseous or liquid environments. ESEM wet mode is widely used for the study of various wettability aspects at the micron scale and nanoscale. It includes studying the triple-line structure [19], surface tension [20], line tension, and hydrophobic/hydrophilic [21] and rough surfaces [22e24]. The condensation and evaporation processes over the sample surface are provided by increasing or reducing the ESEM chamber pressure at constant sample temperature around dew point or alternatively by controlling the temperature at fixed pressure.
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n FIGURE 3.5 Wet mode: (A) Peltier cooling stage with liquid marble on top and GSED below the pole piece, (B) marble surface imaging in wet mode.
Samples of Prof. E. Bormashenko group.
The following demonstration of ESEM wet mode corresponds to the understanding of the mechanism of floating and rolling of liquid marbles [25]. In particular, it supports the assumption that the marble is isolated from the supporting surface by an air layer. The studied marbles are composed of polyvinylidene fluoride (PVDF) particles with average size of 130 nm encapsulating a 10-mL NaOH water drop (middle arrow at Fig. 3.5A). The sample is mounted on a Peltier cooling stage (bottom arrow at Fig. 3.5A), and is imaged at wet mode (2 C and 5.4 torr) using SE GSED (top arrow Fig. 3.5A). The ESEM image of part of the liquid marble surface shows that the PVDF particles do not form a monolithic layer, but are of hierarchical structure (Fig. 3.5B). The center of the image shows the water-clearing area (indicated by arrow at Fig. 3.5B). The hierarchical structure is typical to “lotus leaf” surfaces, known for their superhydrophobic properties. The absence of direct contact between the liquid and solid substrate is explained by the clearing areas surrounded by the particles’ hierarchical coating structure, which provides low-friction rolling and a nonstick air layer.
3.5.4 Imaging at Wet Mode Using Transmitted Electron Signal The wet scanning transmission electron microscope (wet-STEM) detector in ESEM has been established as a novel method for imaging nanoparticles in the liquid volume with a few nanometer resolution [26e27]. Furthermore, it has been utilized for measuring miniemulsions [28], biological samples [29],
3.6 Benefits and Limitations of Imaging in Gaseous/Liquid Environment 97
self-assembly NP dynamics [30], and water dynamics in carbon nanopipes [31]. Applications for wettability study at nanoscale are shown [32e36] for self-supported liquid films. The following demonstration refers to a quantitative wettability study [34] of nanodroplet nucleation and growth using wet-STEM. The wet-STEM mode uses Peltier cooling stage with SSD for imaging transmitted electrons. Thin self-supported water films have been produced inside the holes of a holey carbon TEM grid. Condensation and evaporation are provided by increasing or reducing the ESEM chamber pressure at constant sample temperature. Dynamic in situ imaging shows that irregularities at the water film boundaries constitute nucleation sites for both dropwise and filmwise condensation. Referring to dropwise condensation, the growth of two nanodroplets (signed as “1” and “2”) are studied under constant relative humidity (Fig. 3.6). Initial stages of nanodroplet growth are shown (Fig. 3.6A and B) for two selected time intervals (t ¼ 13s and t ¼ 27s). The power law dependence of droplet radius growth is shown as well (Fig. 3.6C). The fitting provides power law values of m ¼ 0.55 0.02 for the upper droplet (for “1”) and 0.30 0.02 for lower drop (for “2”). The distribution of power law values indicates the role of pinning centers for nanodroplet growth.
3.6 BENEFITS AND LIMITATIONS OF IMAGING IN GASEOUS/LIQUID ENVIRONMENT The SEM method provides a versatile tool due to its multiple advantages, which include relative ease of operation and wide application. In addition, the SEM provides wide lateral range for imaging, which scales from nanometer to centimeter (seven orders of magnitude) and a large depth of focus relative to optical microscopy. Simultaneous topography and energydispersiveewavelength-dispersive spectroscopy (EDS-WDS) chemical analysis is an important advantage for correlative SEM information. SEM can be used as a bulk method (using X-ray signal) as well as surface method (using SE signal) in dependence of the measured signal and acceleration voltage. The SEM provides a platform for e-beam-based systems such as dual-beam SEM-FIB (focused ion beam) for micro- and nanomachining applications. SEM is a nondestructive method for inorganic materials. It provides fast imaging and is thus suitable for in situ dynamic experiments. In situ dynamic imaging in VP-SEM or ESEM can be carried out under low-vacuum or wet environmental conditions, which extends both the material analysis methods and applications. There are possible several radiation-damage effects, which are particularly significant for organic or soft-matter materials. These include bond scission,
98 CHAPTER 3 Scanning Electron Microscopy Under Gaseous Environment
n FIGURE 3.6 Wet-STEM: (A) t ¼ 13s, (B) t ¼ 27s, (C) dynamic droplet radius growth for two nanodroplets (signed as “1” and “2”).
cross-linking, mass loss, e-beam contamination, radiology, charging, and heating. E-beam heating has been reported for a few ESEM applications such as peptide nanotubes [37] and condensed micron-size droplets [38]. The solutions for the charging effects have been shown and discussed in previous sections. We mainly concentrate on two sources of radiation damage under a water-vapor environment: e-beam contamination and e-beam radiology. E-beam contamination occurs in electron microscopy due to carbonization of the sample surface at the e-beam-irradiated area because of sample surface contaminates or residual vacuum hydrocarbons. It is typically observed as darkening of the irradiated area under conventional SEM vacuum conditions
3.6 Benefits and Limitations of Imaging in Gaseous/Liquid Environment 99
of 105 torr. Contamination in gaseous ESEM environments is considered less significant than under conventional high-vacuum conditions. Radiology is a result of inelastic e-beam scattering in liquid samples or vapor environment, which forms ionized and excited water molecular states. It thus refers to both “window technique” (using liquid cells or capsules) and “aperture limited technique” (ESEM and ETEM). The excited water-vapor molecules can decay into free radicals or ions. The high yield of the hydroxyl free radical OH suggests that it dominates the radiology effect leading to degradation of polymer surface at low-vacuum ESEM. The effect is even more significant while in situ ESEM studying liquids over polymers or hydrated biological samples. Radiology effects due to electronewater interactions have recently been reported [39] for liquid-cell electron microscopy. As has been explained, organic materials such as polymers without conductive coating are sensitive to e-beam damage. A typical demonstration is shown at SE images of Fig. 3.7. Polystyrene NPs, of 100-nm diameter, at high vacuum with conductive Au/Pd coating (Fig. 3.7A) are compared with the uncoated sample at low vacuum of 100 Pa (Fig. 3.7B). Degradation is observed for the uncoated polystyrene NPs at low vacuum during a few minutes of e-beam imaging. As has been explained elsewhere [40e41], direct exposure of the sample to e-beam results in material mass loss, whereas additional free radicals (due to e-beam interaction with the vapor) further lead to radiolysis and oxidation of the polymer. Nanoscale imaging requires high magnifications, which increase the e-beam dose and thus heating, mass loss, and radiology effects become dominant.
n FIGURE 3.7 Polystyrene particles (A) coated sample at high-vacuum mode, (B) uncoated sample at low-vacuum mode.
100 CHAPTER 3 Scanning Electron Microscopy Under Gaseous Environment
For soft-matter nanotechnology, the elimination of radiation damage during ESEM research is thus an important task for polymer, organic materials, and liquid samples.
3.7 SUMMARY The VP-SEM and ESEM provide high-resolution imaging at all vacuum modes. These methods are thus used in nanoscience research for broad disciplines and applications. In particular, SEM imaging under gaseous conditions is applicable for low-dimensional structures including NPs, CNTs, and nanodroplets. As is shown, the choice of exact vacuum conditions depends on sample properties. The benefits of multi SEM imaging modes are explicitly shown for both surface and transmission information. Characterization of chemical bonding at micron and nano-scales is important as well for image interpretation and for studying the properties of in situ processed materials under gaseous environments. Thus, besides the known traditional EDS and WDS methods, additional tools will be required in the coming years for nanoscale analysis. Theoretical understanding of radiation damage for each type of studied material is important to eliminate these effects at micron- and nanoscale levels. Future progress in instrumentation is expected to provide lower dose imaging to overcome the mentioned limitations of e-beam radiation damage in organic and biological materials. New detectors and SEM configurations in the coming decade are expected to improve image resolution and S/N ratio under gaseous environments. Further attempts may also be done to carry experiments closer to “real” environmental conditions, i.e., at room temperatures and pressures [42e43]. In addition, it has been shown [44] that better control of environmental conditions (both humidity and temperature) has to be done by adding sensors at the sample position. The availability of in situ probe stations and dedicated sample stages will expand the amount of in situ dynamic experiments in the following years. Therefore, besides lateral resolution, the issue of high temporal resolution is going to play an important role. The nanotechnology research under lowdose imaging, high lateral and temporal resolutions, and high S/N ratio will thus benefit from VP-SEM and ESEM progress in the coming years.
ACKNOWLEDGMENTS Acknowledgment is given to the following researchers at Tel-Aviv University (TAU) for their fruitful collaboration and permission to publish selected pictures:
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Prof. E. Peled and Prof. D. Golodnitsky from the chemical department at TAU, Dr. N. Parkansky and Prof. R. L. Boxman from electrical discharge and plasma laboratory at TAU, and Prof. N. Osherov from microbiology and immunology, school of medicine at TAU. Acknowledgment is given to Prof. E. Bormashenko from Ariel University for his collaboration and image contribution. In addition, the author thanks the Wolfson Applied Materials Research Center (WAMRC) and the center of Nanoscience and Nanotechnology at TAU for supporting the ESEM laboratory.
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