Modelling, quantifying and attenuating multi-material thermal bend in atomic force microscopy cantilevers

Atomic Force Microscopy (AFM) is a technique that generates images of surfaces with a resolution down to the atomic scale through the use of a micro-cantilever. Like many machines that operate at such a scale, it is prone to thermal drift which can result in imaging artefacts. This body of work comprehensively explores the micro-cantilever’s contribution to this phenomenon, particularly in the situation where the cantilever is constructed from two materials (typically an insulator and metal). This work made use of a specific type of AFM called Scanning Thermal Microscopy (SThM), that provided simultaneous temperature and deflection quantification. Modelling demonstrated that an AFM tip can deflect up-to 166 nm/K when out-of-contact resulting in variable and erroneous measurement of deflection and topography in AFM. The latter is a highly unique insight and is a consequence of AFM’s commonly employed optical lever system that measures cantilever rotation (rather than deflection) at the laser spot focused on the cantilever. The AFM converts this signal into tip deflection, using a tip-force defined sensitivity factor, meaning the tip deflection measurement is indirect. This thermal induced deflection was similarly modelled in standard contactmode AFM cantilevers, emphasising its widespread occurrence. With AFM cantilever tips in-contact with surfaces and so their degrees of freedom limited, all cantilevers were theoretically predicted to deflect like a bridge when undergoing a temperature change. This manifested as a humped deflection profile along each cantilever’s length. As a result, this provided the conclusion that an AFM can interpret thermal induced deflection either positively or negatively depending on the longitudinal position ofthe optical lever’s laser. Both out-of-contact and in-contact experimental measurements on SThM cantilevers showed that AFM systems employing optical lever set-ups do have variable and inherently incorrect responses to thermal induced cantilever deflection. This was also seen for commercial contact-mode AFM cantilevers. Measured deflection profiles of all cantilevers when in-contact agreed with models, demonstrating inconsistent AFM measured deflection direction depending on laser location on the cantilever. This provides clear evidence of a new phenomenon not previously documented. Contact AFM scans were employed to confirm this effect’s direct impact on topographic scans with images of the same area varying positively or negatively over a 600 nm range for 2 K of temperature change. For a technique that measures sub-nanometre features, this is a significant artefact. However, with the humped cantilever deflection profile seen in-contact, it offers a point-of-inflexion where very little thermal bending induced tip-deflection would be measured by the optical lever. Measurements demonstrated an improvement of up-to 97.7 % when the laser was focused at this position on the cantilever. This presents a useful technique to mitigate thermal bending artefacts in-contact AFM scan modes, although cantilever bend itself remains present. To address this latter aspect, a simple solution was explored, where the native metal is counteracted by another metal on the backside of the cantilever. Using SThM cantilevers to study this, up-to 99 % reduction in thermal bend induced deflection of the cantilever is possible. New SThM cantilevers with evaporated aluminium on their backside to counteract the native gold were fabricated and showed complete attenuation of thermal bending for laser locations along the whole cantilever length when out-of-contact and in-contact. These were further improved by fabrication of modified SThM cantilevers with gold patterning that better complement the planar aluminium deposition to further improve the effect not only longitudinally, but also laterally in their deflection. This translated into in-contact AFM scans showing variation of only 10s nm for 2 K of temperature change in contrast to 100s nm without any design alteration. These findings were mirrored in commercial, contact-mode AFM cantilevers with a similar trend of pronounced insensitivity to thermal bending when out and in-contact with surfaces, translating into greatly reduced scan artefacts

MyD88 Is Required for Protection from Lethal Infection with a Mouse-Adapted SARS-CoV

A novel human coronavirus, SARS-CoV, emerged suddenly in 2003, causing approximately 8000 human cases and more than 700 deaths worldwide. Since most animal models fail to faithfully recapitulate the clinical course of SARS-CoV in humans, the virus and host factors that mediate disease pathogenesis remain unclear. Recently, our laboratory and others developed a recombinant mouse-adapted SARS-CoV (rMA15) that was lethal in BALB/c mice. In contrast, intranasal infection of young 10-week-old C57BL/6 mice with rMA15 results in a nonlethal infection characterized by high titer replication within the lungs, lung inflammation, destruction of lung tissue, and loss of body weight, thus providing a useful model to identify host mediators of protection. Here, we report that mice deficient in MyD88 (MyD88−/−), an adapter protein that mediates Toll-like receptor (TLR), IL-1R, and IL-18R signaling, are far more susceptible to rMA15 infection. The genetic absence of MyD88 resulted in enhanced pulmonary pathology and greater than 90% mortality by day 6 post-infection. MyD88−/− mice had significantly higher viral loads in lung tissue throughout the course of infection. Despite increased viral loads, the expression of multiple proinflammatory cytokines and chemokines within lung tissue and recruitment of inflammatory monocytes/macrophages to the lung was severely impaired in MyD88−/− mice compared to wild-type mice. Furthermore, mice deficient in chemokine receptors that contribute to monocyte recruitment to the lung were more susceptible to rMA15-induced disease and exhibited severe lung pathology similar to that seen in MyD88−/−mice. These data suggest that MyD88-mediated innate immune signaling and inflammatory cell recruitment to the lung are required for protection from lethal rMA15 infection

Semiochemical strategies for sea louse control: host location cues

The development of behavioural bioassays and electrophysiological recording techniques has enabled the role of semiochemicals to be investigated for the first time in Lepeophtheirus salmonis (Krøyer). Adult male sea lice L salmonis were activated by, and attracted to, salmon-conditioned water (SCW) and SCW extract. Non-host fish odours, turbot-conditioned water (TCW) and turbot-conditioned water extract (TCW extract), elicited activation but no attraction in the lice. Solid-phase extraction techniques were developed to extract low molecular weight components of fish odours, and biological activity was shown to be confined to these. Electrophysiological activity from aesthetascs in the first antennae was found and neural responses to odours were recorded from the antennal nerves. Preliminary experiments on individual chemicals (isophorone, 1-octen-3-ol) linked to salmon revealed behavioural activation and electrophysiological responses in adult male L salmonis. Isophorone was shown to be significantly attractive to sea lice when placed in a slow-release system in a perfused tank of seawater in a choice situation. Proof of concept has been established for successfully extracting odour cues from seawater, analysing their biological activity and applying these to slow-release technologies for field trapping of lice. Future work involving linked GC–MS techniques using behaviour, electrophysiological responses, and organolepsis will establish further host location cues specific to Atlantic salmon

Plasma Blood Levels of Tafenoquine following a Single Oral Dosage in BALBc Mice with Acute <i>Babesia microti</i> Infection That Resulted in Rapid Clearance of Microscopically Detectable Parasitemia

Previous studies of mice infected with Babesia microti have shown that a single dose of tafenoquine administered orally is extremely effective at decreasing microscopically detectable parasitemia. However, a critical limitation of studies to date is the lack of data concerning the plasma levels of tafenoquine that are needed to treat babesiosis. In the current study, we begin to address this gap by examining the plasma levels of tafenoquine associated with the rapid reduction of B. microti patent parasitemia in a mouse model of babesiosis. In the current study, we infected BALB/c mice with 1 × 107 B. microti-infected red blood cells. Two days post-infection, mice were treated with 20 mg/kg of tafenoquine succinate or vehicle control administered orally by gavage. Parasitemia and plasma levels of tafenoquine were evaluated every 24 h post-treatment for 96 h. This allowed us to correlate blood plasma levels of tafenoquine with reductions in parasitemia in treated mice. Consistent with previous studies, a single oral dose of 20 mg/kg tafenoquine resulted in a rapid reduction in parasitemia. Plasma levels of tafenoquine 24 h post-administration ranged from 347 to 503 ng/mL and declined thereafter. This blood plasma tafenoquine level is similar to that achieved in humans using the current FDA-approved dose for the prevention of malaria