Secondary Ion Mass Spectrometry

SUMMARY The right and left gluteus medius muscles of 4 mature Andalusian stallions were examined by repeated needle biopsy over a specific area and depth, as well as at different depths, to determine whether the cross-sectional area and capillary supply of the various fiber types are homogeneous throughout the muscle. The muscle biopsy specimens were histochemically analyzed for fiber types (myofibrillar adenosine triphosphatase) and capillaries (amylase-periodic acid-Schiff method). Differences between contralateral sites were not identifiable for any of the analyzed variables. Differences between sampling depths were larger than those between sample sites. The percentage of type-1 fibers increased, and that of type-2B fibers decreased as a function of increased sampling depth, but the relative frequency of type- 2A fibers did not change with depth. The mean crosssectional area of type-1 and type-2A fibers increased significantly, and that of type-2B fibers decreased between the most superficial and the deepest sampling sites. The capillary density and the mean number of capillaries per fiber increased as a function of sampling depth, but significant changes were not recorded between sampling depths for overall mean fiber area. The mean number of capillaries in contact with fibers of each histochemical type increased significantly with increasing sampling depth only for type- 1 fibers. However, the number of capillaries in contact with fibers changed significantly, relative to the fiber area, as a function of sampling depth; for type- 28 fibers this variable increased, whereas for type-2A and type-1 fibers, it decreased. These results indicate that fiber size and capillary supply of the various fiber types in the gluteus medius muscle of horses vary as a function of depth. These patterns of variation most likely indicate the functional difference between various portions of the muscle, and support the general opinion that the functional demands placed on the fiber population are an important factor in the development of muscle fiber properties.

High detection sensitivity in bulk analysis or depth profiling by secondary ion mass spectrometry (SIMS) can only be achieved, for positively charged ions, if the nearsurface regions of the sputter eroded sample are fully oxidized. Using oxygen primary ions, a stationary oxidation state is established after some time of bombardment during which period the sputtering yield decreases and the ionization probability increases. The physical and chemical processes occurring during the transient period are reviewed with emphasis on the results for impurity analysis in silicon, i.e. the matrix material that has been studied most thoroughly in the past. The transient decrease in sputtering yield gives rise to a depth scale offset and an associated apparent shift of impurity profiles towards the surface. The effect is largest at normal beam incidence, ca. 1 nm /keV (O + 2 ), in which case silicon is fully oxidized. The transition depth, i.e. the depth sputtered before achieving a stable ion yield is about twice as large and increases as the impact angle is turned away from normal. The shift and the depth scale offset can be measured safely using thin (delta) layers of isotopically pure tracers, for example 30 Si in 28 Si. Boron delta layers can serve as secondary standards because this impurity behaves almost the same as the host silicon atoms. The profile shift observed with other common dopants may contain contributions due to unidirectional relocation, often driven by segregation away from the surface. At O + 2 energies below about 0.7 keV the transition depths fall below the thickness of typical native oxides, so that the transient changes in sputtering yield and ionization probability can disappear. The transient phenomena may be described by a simple sputtering-oxidation model that connects the depth scale offset to other observable parameters like the initial and final sputtering yield, the transition fluence and the oxide thickness.

The complementary metal-oxide-semiconductor (CMOS) industry continues to push the boundaries of what is possible. Along with this, secondary ion mass spectrometry (SIMS) depth profiling continues to support CMOS R&D and high volume manufacturing (due to its unparalleled sensitivity and detection limits over predefined volumes and within acceptable time periods). New developments in SIMS are also being realized to support the move of the CMOS industry from planar structures to three-dimensional (3D) structures. This article presents a perspective of existing SIMS research areas for beyond one-dimensional structural analysis along with potential future SIMS implementation and data processing scenarios. The topics covered include: (1) existing SIMS depth profiling approaches for 3D structures, (2) in-fab SIMS deployment, (3) data analysis of in-fab SIMS derived depth profiles from 3D structures via pattern recognition, and (4) hybrid characterization approaches. With SIMS depth profiling of optimized (for SIMS) 3D structures already demonstrated, the remaining topics may serve to extend these and other capabilities and open new application areas for SIMS.

Heavily n-doped epitaxially grown Si layers are of great importance for source/drain (S/D) application in advanced node nMOS devices. For contact resistivity reduction, the dopant activation level is very important. Various techniques are being used to evaluate dopant activation in Si:P layers. Among these two methods are Scanning Spreading Resistance Microscopy (SSRM) and Differential Hall Effect Metrology (DHEM). SSRM uses an atomic force microscope equipped with a hard conductive probe that is scanned in contact mode on the cross-sectioned sample’s surface and measures spreading resistance. Measured resistance values as a function of depth are converted into resistivity and carrier concentration depth profiles using calibration measurements and conversion relationships. DHEM provides depth profiles of mobility, resistivity and carrier concentration through a semiconductor layer by making successive sheet resistance (Rs) and Hall voltage measurements using Hall effect/Van der Pauw techniques, as the electrically active thickness of the layer is reduced through successive oxidation steps. Controlled oxidation is achieved by electrochemical anodization. Data collected can then be processed to yield the depth profiles. In this contribution we have carried out SSRM and DHEM measurements on Si:P epi layers subjected to different processing conditions including annealing and ohmic contact fabrication. Secondary Ion Mass Spectrometry (SIMS) was used to measure the total (active + inactive) dopant profiles through the films. Effects of these processes on dopant diffusion and activation were studied and the results from DHEM and SSRM were compared.In-situ phosphorus (P) doped Si epi-layers were grown over 300mm diameter boron doped monitor wafers. While one set was kept as the reference, a second set was treated by a spike-annealing process at 1000 °C. In a third set a Ti/TiN contact fabrication process was carried out and the contact was removed before analysis. In the fourth set contact process was applied to the spike annealed wafer before removal of the Ti/TiN layers. Bulk sheet resistance measurements were made using 4-point probe (4PP) for all the samples. SSRM measurements were carried out at IMEC. Cross-shaped Van der Pauw test-patterns were formed on 8mmx8mm areas on the samples and DHEM measurements were performed at ALP. Figure below shows the dopant depth profiles obtained by SIMS and the carrier concentration profiles from DHEM and SSRM techniques for samples D02 (as deposited Si:P) and D03 (spike annealed). One can make some general observations from the data in this figure. The total dopant concentration as measured by SIMS is ~ 1.4E21/cm3. There is only a small difference in the total dopant distribution profiles (SIMS) between the as deposited sample and the spike annealed sample. However, the spike annealed sample shows much higher dopant activation as measured by DHEM. Carrier concentration is ~2.5x higher in sample D03 (~5E20/cm3) compared to sample D02 (~2E20/cm3). Activation levels measured by SSRM, however, are lower for both samples, and the peak carrier concentration value increases only slightly upon spike annealing, going from ~2E20/cm3 in sample D02 to ~2.2E20/cm3 in sample D03. DHEM clearly indicates the sharp interface between the p-type substrate and the n-type epi-layer and its depth calibration agrees well with the expected thicknesses of the epi layers. The tail of the SSRM data is much more graded. Data from other samples will be presented and discussed in the final manuscript. Figure 1

Mass spectrometry imaging (MSI) is a label free technique capable of providing simultaneous identification and localization of biomolecules. A multimodal approach is required that allows for the study of the complexity of biological tissue samples to overcome the limitations of a single MSI technique. Secondary ion mass spectrometry (SIMS) allows for high spatial resolution imaging while matrix-assisted laser desorption (MALDI) offers a significantly wider mass range. The combination of coregistered SIMS and MALDI images results in detailed and unique biomolecular information. In this Technical Note, we describe how gold sputtered/implanted fiducial markers (FM) are created and can be used to ensure a proper overlay and coregistration of the two-dimensional images provided by the two MSI modalities.

Loss of depth resolution with depth in secondary ion mass spectrometry (SIMS) due to variations in ion dose density across the rastered area

A method is proposed for the quantitative estimation of the carbon ionization coefficient followed by the determination of its concentration in silicate glasses by secondary ion mass spectrometry (SIMS). The method is based on the calculation of the sputtering ratio of carbon from the surface of silicate glasses. The dependence of the ionization coefficient on the NBO/T parameter (ratio of the number of nonbridging oxygen atoms to the number tetrahedrally coordinated silicon and aluminum ions), corresponding to the structure and composition of the matrix, is shown. The dependences obtained are calibration graphs for the determination of the ionization coefficient and for the subsequent quantitative estimation of carbon in a silicate sample for a particular SIMS instrument and experimental conditions.

Depth profiles and secondary ion mass spectrometry (SIMS) relative sensitivity factors (RSF) for quantification (inverse relative ion yields) are reported for oxygen ion bombardment and positive secondary ion mass spectrometry and for cesium bombardment and negative secondary ion mass spectrometry measured for implants of more than 40 elements in PMMA/polymethyl methacrylate/Lucite/AZ111, polypyromellitimide/polyimide/Kapton, and epoxy, using magnetic sector SIMS instruments (CAMECA IMS 3f or 4f). The RSFs are plotted versus ionization potential (positive secondary ions) and versus electron affinity (negative secondary ions), and the resulting patterns are compared among themselves and with those for semiconductors reported previously. Charge compensation using electron flooding was employed in most cases for both oxygen (oblique incidence flooding) and cesium (normal incidence flooding) primary beams. Profiles in these low atomic number materials are relatively deep. Crater depths are sometimes difficult to measure; some are distorted and some are not. More work in this area of SIMS quantification and depth profiling in organic polymers appears to be needed, but these results are promising.

Secondary ion mass spectrometry (SIMS) is a technique capable of imaging tissues, single cells, and microbes revealing chemical species with sub-micrometer spatial resolution. The recently developed Fourier transform ion cyclotron resonance (FTICR) SIMS instrument provides high mass resolving power and mass accuracy, ToF-SIMS can generate chemical maps with an order of magnitude better lateral resolution than the FTICR-SIMS, and the NanoSIMS instrument offers sub-100 nm spatial resolution in chemical imaging. Many commercial ToF-SIMS instruments are also capable of depth profiling that allows three-dimensional reconstructions of cell and tissue structure.

Recently, a report raised the possibility of shrapnel-induced chronic beryllium disease from long-term exposure to the surface of retained aluminum shrapnel fragments in the body. Since the shrapnel fragments contained trace beryllium, methodological developments were needed for beryllium quantification and to study its spatial distribution in relation to other matrix elements, such as aluminum and iron, in metallurgic samples. In this work, we developed methodology for quantification of trace beryllium in samples of shrapnel fragments and other metallurgic sample-types with main matrix of aluminum (aluminum cans from soda, beer, carbonated water and aluminum foil). Sample preparation procedures were developed for dissolving beryllium for its quantification with the fluorescence detection method for homogenized measurements. The spatial distribution of trace beryllium on the sample surface and in 3D was imaged with a dynamic secondary ion mass spectrometry instrument, CAMECA IMS 3f secondary ion mass spectrometry ion microscope. The beryllium content of shrapnel (∼100 ppb) was the same as the trace quantities of beryllium found in aluminum cans. The beryllium content of aluminum foil (∼25 ppb) was significantly lower than cans. SIMS imaging analysis revealed beryllium to be distributed in the form of low micron-sized particles and clusters distributed randomly in X-Y- and Z dimensions, and often in association with iron, in the main aluminum matrix of cans. These observations indicate a plausible formation of Be-Fe or Al-Be alloy in the matrix of cans. Further observations were made on fluids (carbonated water) for understanding if trace beryllium in cans leached out and contaminated the food product. A direct comparison of carbonated water in aluminum cans and plastic bottles revealed that beryllium was below the detection limits of the fluorescence detection method (∼0.01 ppb). These observations indicate that beryllium present in aluminum matrix was either present in an immobile form or its mobilization into the food product was prevented by a polymer coating on the inside of cans, a practice used in food industry to prevent contamination of food products. The lack of such coating in retained shrapnel fragments renders their surface a possible source of contamination for long-term exposure of tissues and fluids and induction of disease, as characterized in a recent study. Methodological developments reported here can be extended to studies of beryllium in electronics devices and components.

The advantages of applying multivariate analysis to mass spectrometry imaging (MSI) data have been thoroughly demonstrated in recent decades. The identification and visualization of complex relationships between pixels in a hyperspectral data set can provide unique insights into the underlying surface chemistry. It is now recognized that most MSI data contain nonlinear relationships, which has led to increased application of machine learning approaches. Previously, we exemplified the use of the self-organizing map (SOM), a type of artificial neural network, for analyzing time-of-flight secondary ion mass spectrometry (TOF-SIMS) hyperspectral images. Recently, we developed a novel methodology, SOM-relational perspective mapping (RPM), which incorporates the algorithm RPM to improve visualization of the SOM for 2D TOF-SIMS images. Here, we use SOM-RPM to characterize and interpret 3D TOF-SIMS depth profile data, voxel-by-voxel. An organic Irganox™ multilayer standard sample was depth profiled using TOF-SIMS, and SOM-RPM was used to create 3D similarity maps of the depth-profiled sample, in which the mass spectral similarity of individual voxels is modeled with color similarity. We used this similarity map to segment the data into spatial features, demonstrating that the unsupervised method meaningfully differentiated between Irganox-3114 and Irganox-1010 nanometer-thin multilayer films. The method also identified unique clusters at the surface associated with environmental exposure and sample degradation. Key fragment ions characteristic of each cluster were identified, tying clusters to their underlying chemistries. SOM-RPM has the demonstrable ability to reduce vast data sets to simple 3D visualizations that can be used for clustering data and visualizing the complex relationships within.

A detailed protocol for quantitative single cell mass spectrometry imaging (MSI) analysis is described in this chapter with examples of the treatment of cells with anticancer drug, cisplatin. Cisplatin, cis-diamminedichloridoplatinum ii (CDDP), is widely used for the treatment of many malignancies, including testicular, ovarian, bladder, cervical, head and neck, and small cell and non-small cell lung cancers. The possibility of renal injury by cisplatin treatment is a major dose-limiting factor in this cancer therapy. At present, the mechanisms of cisplatin-induced renal cytotoxicity are poorly understood. In this work, secondary ion mass spectrometry (SIMS) was used for investigating cisplatin-induced alterations in intracellular chemical composition in a well-established model (LLC-PK(1) cell line) for studying renal injury. The cells were cryogenically prepared by the sandwich freeze-fracture method for subcellular imaging analysis of chemical composition (total concentrations of K(+), Na(+), and Ca(2+)) in individual cells. The single cell analysis of these diffusible ions necessitates the use of reliable cryogenic sample preparations for SIMS. The sandwich freeze-fracture method offers a simple approach for cryogenically preserving diffusible ions and molecules inside the cells for SIMS analysis. A CAMECA IMS-3f SIMS ion microscope instrument capable of producing chemical images of single cells with 500-nm spatial resolution was used in the study. In cisplatin-treated cells, SIMS imaging showed the presence of detectable amount of platinum at mass 195, as (195)Pt(+) secondary ions in individual cells. SIMS observations also revealed that individual cells differed in their response to cisplatin. While the chemical composition of some cells was unaffected by cisplatin, others showed a reduction in cytoplasmic calcium stores that was not associated with changes in their intracellular K or Na concentrations. Another population of cells displayed an increase in cytoplasmic calcium concentration that was associated with higher levels of intracellular Na and a reduction in K concentration of the same cells. Since the loss of intracellular K and the gain of Na and Ca are typical symptoms of cell injury, it is plausible that the initial response of the cell to cisplatin treatment is the reduction in cytoplasmic calcium pool in stores. If, somehow, the calcium stores are compromised with cisplatin, then maintenance of free Ca(2+) homeostasis would become uncontrollable in the cell. These observations open new avenues of research for understanding of the mode of action of cisplatin in cell injury. This study also demonstrates the need and vast potential of single cell imaging mass spectrometry techniques in cell biology and medicine.

Secondary ion mass spectrometry (SIMS) has become a powerful tool for a chemical imaging of surfaces. In particular, cluster ion beams have been widely implemented with SIMS using spatial distributions of molecules as an analysis and imaging method for a sample composed of organic molecules. In this study, we developed a home-built time-of-flight (ToF)-SIMS instrument and generated organic molecular cluster ion beams ionized by multiphoton ionization (MPI) as a primary ion beam. Further, the sample stage loaded with rhodamine 6G was bombarded with size-controlled toluene cluster ions to produce secondary ions. Mass spectra of rhodamine 6G were acquired as a function of the size of toluene cluster ions. We plotted the fragments-to-parent ion ratio against each size of cluster ion, and we found that an intact rhodamine 6G signal would be observed with less fragmentations using (Tol)n > 18+. We expect this combination of the organic molecular cluster ion beam generated by MPI and ToF-SIMS to greatly expand the field of surface analysis.

A micromolar concentration of zinc has been shown to significantly change the dynamics of exocytosis as well as the vesicle contents in a model cell line, providing direct evidence that zinc regulates neurotransmitter release. To provide insight into how zinc modulates these exocytotic processes, neurotransmitter release and vesicle content were compared with single cell amperometry and intracellular impact vesicle cytometry with a range of zinc concentrations. Additionally, time-of-flight secondary ion mass spectrometry (ToF-SIMS) images of lipid distributions in the cell membrane after zinc treatment correlate to changes in exocytosis. By combining electrochemical techniques and mass spectrometry imaging, we proposed a mechanism by which zinc changes the fusion pore and the rate of neurotransmitter release by changing lipid distributions and results in the modulation of synaptic strength and plasticity.

New methods to analyze cells and tissues in ambient condition without any harsh chemical fixation or physical freezing and drying are summarized in this report. The first approach, an atmospheric pressure mass spectrometry imaging method, is based on laser ablation in atmospheric pressure assisted by atmospheric plasma and nanomaterials such as nanoparticles and graphene to enhance laser ablation. The second one is based on secondary ion mass spectrometry (SIMS) imaging of live cells in solution capped with single-layer graphene to preserve intact and hydrated biological samples even under ultrahigh vacuum for SIMS bio-imaging in solution.