An immunogold labelling technique was used to label the pili of the bacterium Bacteroides nodosus. The labelling was distinct and highly specific, and individual pili could be recognised beneath the gold probe. The labelling of somatic antigens could be distinguished from that of pilus antigens. Furthermore, labelling of fragments of cytoplasm released by cell lysis and trapped in the pili could be distinguished from pilus labelling. An antiserum that had been raised against strain 80200 (serotype N) labelled pili of strain 215 (serotype B). Double labelling experiments with this antiserum and the antiserum against strain 215 (serotype B) showed that both antisera label the same pili bundles. The ease of detection of the immunocytochemical reaction should enable this technique to be used as a routine screen for pilus antigens. It also possesses the potential for much wider applications for immunolabelling other antigens, such as viruses, that can be obtained in suspension.
In this work we combine sample elevation masks and elemental composition masks to automatically find just those areas characterized by both a given composition and elevation. Applications include differentiating contaminants from substrates when they share common elements and evaluating manufactured materials where sample height is important as in abrasives. Both SEM and EDS data are required as EDS alone does not measure elevation and SEM alone does not measure composition. The goal is a technique for routine evaluation of materials with rough surfaces.It is possible to measure sample elevation using four images gained from the four independent segments of a four quadrant backscatter electron(BSE) detector [1] in an SEM. Figure 1 shows a BSE image of a wire attached to a bond pad of an integrated circuit. Figure 2 shows an elevation profile taken along the blue line shown in Figure 1. Figure 3 shows an image in which the grey scale encodes sample height. Brighter areas are higher above the substrate.Spectral imaging collects an x-ray map by storing a full x-ray spectrum at each point in an array which spans a portion of a sample. From this Multivariate Spectral Analysis[2] finds self consistent patterns in the data. A clustering algorithm converts this result to binary masks defining areas of the sample that have similar composition. Together these three technologies allow an analyst to automatically find areas of similar composition. The use of a silicon drift detector(SDD) allows this process to proceed very rapidly [3]. Figure 4 is the automatically determined mask showing the distribution of Al in this sample. Note that it does not differentiate the wire from the pad. The next sample consists of a mixture of NaCl and KCl resting on a bed of NaCl and carbon. Figures 7, 8 and 9 show the BSE image, height profile and the height encoded image respectively.Figures 10, 11 and 12 show the Na, Cl and K x-ray distribution maps respectively. Figure 13 shows the NaCl mask determined by MVS and clustering. Figure 14 is the threshold mask generated from figure 9. Figure 15 is the result which finds just the area of the sample consisting of NaCl and which is above the level of the substrate. The use of dual opposed detectors would improve on this result by reducing shadowing artifacts producing unambiguous data for all points.A Hitachi S-3700 SEM was used to obtain the BSE images and 3D images. A Thermo Scientific NS7 EDS system with a single Ultradry SDD was used to collect the x-ray maps and composition masks. Applescript and GraphicConverter were used to prototype the thresholding and logic operations.
Scanning electron microscopy (SEM) for imaging the interface between carbon nanofibers (CNFs) and the underlying substrate is presented. By irradiating the electron beam perpendicular to the substrate, bright contrast is observed at the region where a small gap exists between the CNF and substrate. The energy-diameter diagram for the observation of the bright contrast is derived, which can be understood by using the theory of electron penetration into solid. Monte Carlo simulation is performed to reproduce the experimental observation based on our model, and the contrast sensitivity to the gap height is discussed.
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