AFM detects binding events in cell-virus interactions
This content originally appeared in the Jan 2017 issue and is a sponsored blog
Bruker’s PeakForce Tapping® mode enables high-resolution imaging of mammalian cells under physiologically relevant conditions while simultaneously mapping their physical properties with nanometre scale resolution.
In a recent study, researchers Alsteens et al. used PeakForce Tapping to map the binding of single viruses directly on living cells (Nature Nanotechnology (2016) doi:10.1038/nnano. 2016.228). To accomplish this, PeakForce Tapping was integrated with confocal laser scanning microscopy to target MDCK cells that expressed fluorescently labelled TVA receptors. The researchers were able to quantify and topographically map their binding to single EnvA-RABV(∆G) viruses with a spatial resolution of <50 nm. The studies were conducted using a novel, environmental chamber that mimics an incubator-type local environment and thus maintains cells under homeostastic conditions over extended time periods.
The upper left image (shown above) is an overlay of the DIC and confocal mCherry fluorescence images of the MDCK cells. The mCherry channel indicates which of the cells are expressing TVA receptors on their surface. The presence of both wild-type (nonfluorescent) and TVA-mCherry labelled cells provides an internal control for verification of the specificity of the binding interactions (scale bar: 25µm). Note that the shadow of the AFM cantilever is also observed in the overlay image.
The upper right image only shows the mCherry channel image. The lower PeakForce Tapping topography (left) and adhesion (right) images were collected in the area outlined by the white squares on the optical images (scale bar: 10 µm). The AFM probe was functionalized with a single virus particle and then used to image the cell surfaces. Only cells expressing TVA receptors (mCherry fluorescence image) were found to interact with the virus, with these binding events appearing as bright pixels in the adhesion image.
Analysing the individual force curves associated with the PeakForce Tapping images not only enabled Alsteens et al. to quantify the adhesion force between an individual virus and single receptor, they were also able to extract the kinetic and thermodynamic parameters describing the binding free-energy landscape of the interaction. Their quantitative method may not only help to uncover how receptors are regulated and employed for virus entry, it could also potentially provide a tool to quantify the effects of molecules that interfere with viral entry, and therefore aid the development of novel drugs to fight against virus infection.
To learn more about Prof. Alsteen’s research and Bruker’s PeakForce Tapping technology, go to www.bruker.com/webinars-AFM
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