Application of confocal scanning laser microscopy in experimental pathology

Confocal scanning laser microscopy (CSLM) represents an exciting new tool for scientific disciplines which focus on mechanistic studies such as experimental pathology. Enhanced resolution in the specimen plane and rejection of out-of-focus fluorescence flare allow analysis of specific nucleic acid sequences, enzymes, structural macromolecules, and cellular homeostasis utilizing fluorescent probes. Four different experimental applications are discussed which utilize CSLM to evaluate pathological processes at the subcellular, cellular, and tissue levels. Programmed cell death, or apoptosis, is a natural process of significance both during development and as a response to toxic stimuli. CSLM-imaging of nuclei of human B lymphoblastoid cells following exposure to a monofunctional alkylating agent suggests that the degradation of chromatin characteristic of apoptosis may occur in asymmetric patterns. Surfactant apoprotein-A is the major non-serum protein component of pulmonary surfactant and is essential for the extracellular function of surfactant. CSLM of alveolar type II cells suggests that apoprotein-A is present in both the cytoplasm, predominantly in lamellar bodies, and in the nucleus. The tumor promoter, phorbol myristate acetate, rapidly stimulated the formation of vacuoles in human neutrophils. CSLM using Lucifer Yellow as a probe suggests that cylindrical vacuoles are formed by fluid-phase pinocytosis. The blood-nerve barrier (BNB) in peripheral nerves may be an important target during toxin-induced neuropathies. Ricin-induced permeability of the BNB in the rat was rapidly visualized by CSLM as leakage of fluorescein isothiocynate (FITC)-dextran into the endoneurial compartment.     

Force detected electron spin resonance at 94 GHz

Force detected electron spin resonance (FDESR) detects the presence of unpaired electrons in a sample by measuring the change in force on a mechanical resonator as the magnetization of the sample is modulated under magnetic resonance conditions. The magnetization is coupled to the resonator via a magnetic field gradient. It has been used to both detect and image distributions of electron spins, and it offers both extremely high absolute sensitivity and high spatial imaging resolution. However, compared to conventional induction mode ESR the technique also has a comparatively poor concentration sensitivity and it introduces complications in interpreting and combining both spectroscopy and imaging. One method to improve both sensitivity and spectral resolution is to operate in high magnetic fields in order to increase the sample magnetization and g-factor resolution. In this article we present FDESR measurements on the organic conductor (fluoranthene)(2)PF(6) at 3.2 T, with a corresponding millimeter-wave frequency of 93.5 GHz, which we believe are the highest field results for FDESR reported in the literature to date. A magnet-on-cantilever approach was used, with a high-anisotropy microwave ferrite as the gradient source and employing cyclic saturation to modulate the magnetization at the cantilever fundamental frequency.     

Print your atomic force microscope

Progress in scanning probe microscopy profited from a flourishing multitude of new instrument designs, which lead to novel imaging modes and as a consequence to innovative microscopes. Often these designs were hampered by the restrictions, which conventional milling techniques impose. Modern rapid prototyping techniques, where layer by layer is added to the growing piece either by light driven polymerization or by three-dimensional printing techniques, overcome this constraint, allowing highly concave or even embedded and entangled structures. We have employed such a technique to manufacture an atomic force microscopy (AFM) head, and we compared its performance with a copy milled from aluminum. We tested both AFM heads for single molecule force spectroscopy applications and found little to no difference in the signal-to-noise ratio as well as in the thermal drift. The lower E modulus seems to be compensated by higher damping making this material well suited for low noise and low drift applications. Printing an AFM thus offers unparalleled freedom in the design and the rapid production of application-tailored custom instruments.     

The effect of some instrument operating conditions on the x-ray microanalysis of particles in the environmental scanning electron microscope

The objective of this investigation was to evaluate the practical effects of electron beam broadening in the environmental scanning electron microscope (ESEM) on particle x-ray microanalysis and to determine some of the optimum operating conditions for this type of analysis. Four sets of experiments were conducted using a Faraday cage and particles of copper, glass, cassiterite, andrutile. The accelerating voltage and chamber pressure varied from 20 to 10 kV and from 665–66 Pa (5.0 to 0.5 torr), respectively. The standard gaseous secondary electron detectors (GSED) and the long environmental secondary dectectors (ESD) for the ESEM were evaluated at different working distances. The effect of these parameters on the presence of artifact peaks was evaluated. The particles were mounted on carbon tape on an aluminum specimen mount and were analyzed individually and as a mixture. Substrate peaks were present in almost all of the spectra. The presence of neighboring particle peaks and the number of counts in these depended upon the operating conditions. In general, few of these peaks were observed with the long ESD detector at 19 mm working distance and at low chamber pressures. More peaks and counts were observed with a deviation from these conditions. The most neighboring peaks and counts were obtained with the GSED detector at 21.5 mm working distance, 10 kV accelerating voltage, and 665 Pa (5.0 torr) chamber pressure. The results of these experiments support the idea that the optimum instrumental operating conditions for EDS analysis in the ESEM occur by minimizing the gas path length and the chamber water vapor pressure, and by maximizing the accelerating voltage. The results suggest that the analyst can expect x-ray counts from the mounting materials. These tests strongly support the recommendation of the manufacturer to use the long ESD detector and a 19 mm working distance for EDS analysis. The results of these experiments indicate that neighboring particles millimeters from the target may contribute x-ray counts to the spectrum.     

Human, rhesus macaque, and feline sequences highly similar to mouse mammary tumor virus sequences

Sequences highly similar (>95%) to the mouse mammary tumor virus (MMTV) env gene have been amplified from human DNA samples, including DNA samples from patients with breast cancer (BC) and persons who did not have BC. The sequences from human DNA were distinct from the MMTV sequences used as controls in these PCR reactions, indicating that these results are not simply due to contamination. In addition to both, mouse and human-related sequences were also amplified from some monkey and cat genomic DNA samples. These products were shown to be distinct from, but highly related to, the MMTV env gene, whereas, testing of other sources (lambda phage, snake, cockroach, sea urchin, chicken, or dog) demonstrated no specific amplification. A sequence 90% similar to the MMTV group antigen gene (gag) was amplified from cat DNA. These results indicate that DNA from vertebrate species other than rodents, including some but not all humans, monkeys, and cats, can contain sequences closely related to MMTV.     

Effect of microstructure on friability in the system Al2O3-ZrO2

This paper summarizes results on the use of quantitative microscopy in relating microstructural features to the friability of a model abrasive material, Al₂O₃-ZrO₂. Quantitative stereology was used to characterize the geometric aspects of the microstructure of fused Al₂O₃-ZrO₂ abrasives and to relate these to friability. The microstructure was controlled by varying processing parameters. The quench rate of the material from the melt was found to exhibit the most control over the microstructure. The friability was measured using three methods: the ball mill test, a DeBeers Friatester and a microfriability test developed for this study. Friability was found to be controlled by the geometric aspects of microstructure. The microstructural parameter which appeared to exert the largest influence over friability was the scale factor $\text{}\text{?}$gl^α. A fracture mechanics approach was employed to explain the observed increase in strength and toughness in Al₂O₃-ZrO₂ materials.     

Multiple-input microcantilever sensors

A surface-micromachined micro-electro-mechanical-system (MEMS) process has been used to demonstrate multiple-input chemical sensing using selectively coated cantilever arrays. Cantilever motion due to absorption-induced stress was readout using a custom-designed, eight-channel integrated circuit. Combined hydrogen and mercury vapor detection was achieved with a palm-sized, self-powered module with spread-spectrum telemetry reporting.     

The PIBETA spectrometer for studying rare and forbidden decays of muons and pions

The construction and characteristics of the PIBETA spectrometer are described. This spectrometer is designed to implement a program of precise measurement of pion decay ^{+ 0} + e ⁺ + _e at the Paul Scherrer Institute (Switzerland). A spherical calorimeter, consisting of 240 crystals of pure CsI scintillator and embracing a solid angle of 3, is the main detector of the setup. In addition, the spectrometer is composed of an active collimator (which also acts as a beam degrader), a segmented active plastic target, two multiwire cylindrical proportional chambers, a 20-element cylindrical plastic hodoscope, and veto counters of cosmic muons.__________Translated from Pribory i Tekhnika Eksperimenta, No. 2, 2005, pp. 39–48.Original Russian Text Copyright © 2005 by Baranov, Kalinnikov, Karpukhin, Khomutov, A. Korenchenko, S. Korenchenko, Kravchuk, Kuchinskii, Mzhaviya, Rozhdestvenskii, Sidorkin, Tsamalaidze, Sakhelashvili, Frlez, Poanic, Li, Minehart, Smith, Stephens, Ziock, Bertl, Horisberger, Ritt, Schnyder, Wirtz, Ritchie, Supek, Kozlowski.     

In-plane contributions to phase contrast in intermittent contact atomic force microscopy

Contrast in the phase response of intermittent-contact atomic force microscopy (IC-AFM) reveals in-plane structural and mechanical properties of polymer monolayers. This result is unexpected, as IC-AFM has previously only been considered as a probe of out-of-plane properties. Until now, AFM measurements of nanoscale in-plane properties have employed contact mode techniques. In-plane property measurements are possible with intermittent contact AFM because there is a small but significant component of tip motion parallel to the sample surface. This in-plane component of tip displacement is virtually universal in AFM, implying that oscillating-tip techniques generally are sensitive to in-plane material properties. We present a simple Hertzian model of intermittent-contact AFM that includes such an in-plane displacement.     

Normal personality assessment in clinical practice: The NEO Personality Inventory.

Personality psychologists from a variety of theoretical perspectives have recently concluded that personality traits can be summarized in terms of a 5-factor model. This article describes the NEO Personality Inventory (NEO–PI), a measure of these 5 factors and some of the traits that define them, and its use in clinical practice. Recent studies suggest that NEO–PI scales are reliable and valid in clinical samples as in normal samples. The use of self-report personality measures in clinical samples is discussed, and data from 117 "normal" adult men and women are presented to show links between the NEO–PI scales and psychopathology as measured by D. N. Jackson's (1989) Basic Personality Inventory and L. Morey's (1991) Personality Assessment Inventory. The authors argue that the NEO–PI may be useful to clinicians in understanding the patient, formulating a diagnosis, establishing rapport, developing insight, anticipating the course of therapy, and selecting the optimal form of treatment for the patient. (PsycINFO Database Record (c) 2010 APA, all rights reserved)