2 research outputs found
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Ultra-flat Gold QCM Electrodes Fabricated with Pressure Forming Template Stripping for Protein Studies at the Nanoscale
Single-molecule imaging of proteins using atomic force microscopy (AFM) is crucially dependent on protein attachment to ultra-flat substrates. The technique of template stripping (TS), which can be used to create large areas of atomically flat gold, has been used to great effect
for this purpose. However, this approach requires an epoxy which can swell in solution, causing surface roughening and substantially increasing the thickness of any sample, preventing its use on acoustic resonators in liquid. Diffusion bonding techniques should circumvent this problem
but cannot be used on samples containing patterned features with mismatched heights due to cracking and poor transfer. Here, we describe a new technique called pressure forming template stripping (PTS) which permits an ultra-flat (0.35 ± 0.05 nm root-mean-square roughness) layer
of gold to be transferred to the surface of a patterned substrate at low temperature and pressure. We demonstrate this technique by modifying a quartz crystal microbalance (QCM) sensor to contain an ultra-flat gold surface. Standard QCM chips have substantial roughness, preventing
AFM imaging of proteins on the surface after measurement. With our approach there is no need to run samples in parallel: the modified QCM chip is flat enough to permit high-contrast AFM imaging after adsorption studies have been conducted. The PTS-QCM chips are then used to
demonstrate adsorption of bovine serum albumin in comparison to rough QCM chips. The ability to attach thin layers of ultra-flat metals to surfaces of heterogeneous nature without epoxy will have many applications in diverse fields where there is a requirement to observe nanoscale phenomena with multiple techniques, including surface and interfacial science, optics, and biosensing
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Activation of P2X4 receptors induces an increase in the area of the extracellular region and a decrease in receptor mobility.
The P2X4 receptor (P2X4R) is an ATP-gated cation channel. Here, we used fast-scan atomic force microscopy (AFM) to visualize changes in the structure and mobility of individual P2X4Rs in response to activation. P2X4Rs were purified from detergent extracts of transfected cells and integrated into lipid bilayers. Activation resulted in a rapid (2Â s) and substantial (10-20Â nm2 ) increase in the cross-sectional area of the extracellular region of the receptor and a corresponding decrease in receptor mobility. Both effects were blocked by the P2X4R antagonist 5-BDBD. Addition of cholesterol to the bilayer reduced receptor mobility, although the ATP-induced reduction in mobility was still observed. We suggest that the observed responses to activation may have functional consequences for purinergic signalling