High-speed AFM height spectroscopy reveals microsecond-dynamics of unlabeled biomolecules

In their recent publication “High-speed AFM height spectroscopy reveals μs-dynamics of unlabeled biomolecules” in Nature Communications George R. Heath and Simon Sheuring develop and apply HS-AFM height spectroscopy (HS-AFM-HS, a technique inspired by fluorescence spectroscopy), a technique whereby the AFM tip is held at a fixed x–y position and  the height fluctuations under the tip in z-direction with Angstrom spatial and 10µs temporal resolution are monitored.

They demonstrate “how this technique can be used to simultaneously measure surface concentrations, diffusion rates and oligomer sizes of highly mobile annexin-V molecules during membrane-binding and self-assembly at model membranes and derive its kinetic and energetic terms. Additionally, HS-AFM-HS at specific positions in the annexin lattice where the freedom of movement is restricted to rotation allowed determination of the interaction free energies of protein-protein contacts.”* The applicability of this technique is wide and is discussed at the end of the publication.

NanoWorld Ultra-Short Cantilevers (USC) for Fast-/High-Speed AFM  ( USC-F1.2-k0.15 ) were used.

Congratulations to the authors to this publication which pushes the speed limits of AFM even further!

Increasing the temporal resolution of HS-AFM by reducing the dimensionality of data acquisition. a HS-AFM image of a DOPC/DOPS (8:2) membrane in the presence of annexin-V and NP-EGTA-caged Ca2+. Blue arrows illustrate the slow- (vertical) and the fast-scan axis (horizontal). Images can be captured at up to 10–20 frames s−1. b HS-AFM movie frames of A5 membrane-binding, self-assembly and formation of p6 2D-crystals upon UV-illumination induced Ca2+-release. c Average height/time trace of the membrane area in b. d Averaged HS-AFM image of an A5 p6-lattice overlaid with the subsequent line scanning kymograph, obtained by scanning repeatedly the central x-direction line as illustrated by the blue arrow with a maximum rate of 1000–2000 lines s−1. e Line scanning kymograph across one protomer of the non-p6 trimer, marked by * in d and e at a rate of 417 lines s−1 (2.4 ms per line). f Histogram of state dwell-times of the molecule in e. g HS-AFM image of an A5 p6-lattice partially covering a DOPC/DOPS (8:2) SLB surface during self-assembly. HS-AFM height spectroscopy (HS-AFM-HS) is performed following halting the x- and y-piezos to capture height information at a fixed position at the center of the image (illustrated by the target). h Schematic showing the principle of HS-AFM-HS. The AFM tip is oscillated in z at a fixed x,y-position, detecting single molecule dynamics such as diffusion under the tip. i Height/time trace obtained by HS-AFM-HS with the tip positioned at the center of image (g). The height/time trace allows determination of the local A5 concentration analyzing the time fraction of the occurrence of height peaks. j Dwell-time analysis of each height peak of diffusing A5 from 60 s height/time data and subsequent fitting of the distribution to multiple Gaussians (possible molecular aggregates corresponding to the fits with distinct dwell-times (τD) are shown above the graph). All scale bars: 20 nm, NanoWorld Ultra-Short Cantilevers (USC) for Fast-/High-Speed AFM ( USC-F1.2-k0.15 ) were used.
Figure 1 from “High-speed AFM height spectroscopy reveals μs-dynamics of unlabeled biomolecules”: Increasing the temporal resolution of HS-AFM by reducing the dimensionality of data acquisition. a HS-AFM image of a DOPC/DOPS (8:2) membrane in the presence of annexin-V and NP-EGTA-caged Ca2+. Blue arrows illustrate the slow- (vertical) and the fast-scan axis (horizontal). Images can be captured at up to 10–20 frames s−1. b HS-AFM movie frames of A5 membrane-binding, self-assembly and formation of p6 2D-crystals upon UV-illumination induced Ca2+-release. c Average height/time trace of the membrane area in b. d Averaged HS-AFM image of an A5 p6-lattice overlaid with the subsequent line scanning kymograph, obtained by scanning repeatedly the central x-direction line as illustrated by the blue arrow with a maximum rate of 1000–2000 lines s−1. e Line scanning kymograph across one protomer of the non-p6 trimer, marked by * in d and e at a rate of 417 lines s−1 (2.4 ms per line). f Histogram of state dwell-times of the molecule in e. g HS-AFM image of an A5 p6-lattice partially covering a DOPC/DOPS (8:2) SLB surface during self-assembly. HS-AFM height spectroscopy (HS-AFM-HS) is performed following halting the x- and y-piezos to capture height information at a fixed position at the center of the image (illustrated by the target). h Schematic showing the principle of HS-AFM-HS. The AFM tip is oscillated in z at a fixed x,y-position, detecting single molecule dynamics such as diffusion under the tip. i Height/time trace obtained by HS-AFM-HS with the tip positioned at the center of image (g). The height/time trace allows determination of the local A5 concentration analyzing the time fraction of the occurrence of height peaks. j Dwell-time analysis of each height peak of diffusing A5 from 60 s height/time data and subsequent fitting of the distribution to multiple Gaussians (possible molecular aggregates corresponding to the fits with distinct dwell-times (τD) are shown above the graph). All scale bars: 20 nm

*George R. Heath & Simon Scheuring
High-speed AFM height spectroscopy reveals μs-dynamics of unlabeled biomolecules
Nature Communicationsvolume 9, Article number: 4983 (2018)
DOI: https://doi.org/10.1038/s41467-018-07512-3

Please follow this external link for the full article: https://rdcu.be/bdaKU

Open Access The article “High-speed AFM height spectroscopy reveals μ s-dynamics of unlabeled biomolecules” by George R. Heath & Simon Scheuring is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

Meet us at MRS Fall 2018 this week

Got questions about AFM probes that you’ve always wantend to ask? You’re welcome to pass by NanoAndMore USA booth no. 610 at the 2018 MRS Fall Exhibit this week and meet NanoWorld CEO Manfred Detterbeck there.

NanoWorld AFM probes CEO Manfred Detterbeck at NanoAndMore USA booth no. 610 at MRS Fall 2018
NanoWorld CEO Manfred Detterbeck at NanoAndMore USA booth no. 610 at MRS Fall 2018

Direct AFM-based nanoscale mapping and tomography of open-circuit voltages for photovoltaics

In the article cited below Katherine Atamanuk, Justin Luria and Bryan D. Huey present “a new approach for directly mapping VOC (open-circuit voltage) with nanoscale resolution, requiring a single, standard-speed AFM scan. This leverages the concept of the proportional-integral-derivative (PID) feedback loop that underpins nearly all AFM topography imaging.”*

NanoWorld™ Pointprobe® CDT-NCHR conductive diamond coated silicon AFM probes were used in the described CT-AFM experiment.

Supporting information for «Direct AFM-based nanoscale mapping and tomography of open-circuit voltages for photovoltaics”: Figure S1: Representative quasi-VOC* image from the measured photocurrent upon illumination during an applied voltage fixed at 700 mV. NANOSENSORS conductive diamond coated CDT-NCHR AFM probes were used in the described CT-AFM experiment
Supporting information for «Direct AFM-based nanoscale mapping and tomography of open-circuit voltages for photovoltaics”: Figure S1: Representative quasi-VOC* image from the measured photocurrent upon illumination during an applied voltage fixed at 700 mV.

“Cadmium Telluride (CdTe) is an inexpensive thin-film photovoltaic with ca. 5% of the 2017 global market share for solar cells. To optimize the efficiency and reliability of these, or any electronic devices, a thorough understanding of their composition, microstructure, and performance is necessary as a function of device design, processing, and in-service conditions. Atomic force microscopy (AFM) has been a valuable tool for such characterization, especially of materials properties and device performance at the nanoscale. In the case of thin-film solar cells, local photovoltaic (PV) properties such as the open-circuit voltage, photocurrent, and work function have been demonstrated to vary by an order of magnitude, or more, within tens of nanometers […] Recently, property mapping with high spatial resolution by AFM has been further combined with the ability to serially mill a surface, in order to reveal underlying surface structures and uniquely develop three-dimensional (3D) nanoscale property maps. The most notable examples are based on pure current detection with the AFM to resolve conduction pathways in filamentary semiconducting devices and interconnects […], and tomographic AFM of photocurrents in polycrystalline solar cells during in situ illumination […].”*

*Katherine Atamanuk, Justin Luria, Bryan D. Huey
Direct AFM-based nanoscale mapping and tomography of open-circuit voltages for photovoltaics
Beilstein Journal of Nanotechnology 2018, 9, 1802–1808.
doi: 10.3762/bjnano.9.171

The article cited above is part of the Thematic Series “Scanning probe microscopy for energy-related materials”.

Please follow this external link for the full article: https://www.beilstein-journals.org/bjnano/articles/9/171

The article “Direct AFM-based nanoscale mapping and tomography of open-circuit voltages for photovoltaics” by Atamanuk et. al is an Open Access article under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.