Atomic Force Microscopy Height Research Articles

Atomic force microscopy 3D structural reconstruction of individual particles in the study of amyloid protein assemblies.

Recent developments in atomic force microscopy (AFM) image analysis have made three-dimensional (3D) structural reconstruction of individual particles observed on 2D AFM height images a reality. Here, we review the emerging contact point reconstruction AFM (CPR-AFM) methodology and its application in 3D reconstruction of individual helical amyloid filaments in the context of the challenges presented by the structural analysis of highly polymorphous and heterogeneous amyloid protein structures. How individual particle-level structural analysis can contribute to resolving the amyloid polymorph structure-function relationships, the environmental triggers leading to protein misfolding and aggregation into amyloid species, the influences by the conditions or minor fluctuations in the initial monomeric protein structure on the speed of amyloid fibril formation, and the extent of the different types of amyloid species that can be formed, are discussed. Future perspectives in the capabilities of AFM-based 3D structural reconstruction methodology exploiting synergies with other recent AFM technology advances are also discussed to highlight the potential of AFM as an emergent general, accessible and multimodal structural biology tool for the analysis of individual biomolecules.

Live-Cell SOFI Correlation with SMLM and AFM Imaging.

Standard optical imaging is diffraction-limited and lacks the resolving power to visualize many of the organelles and proteins found within the cell. The advent of super-resolution techniques overcame this barrier, enabling observation of subcellular structures down to tens of nanometers in size; however these techniques require or are typically applied to fixed samples. This raises the question of how well a fixed-cell image represents the system prior to fixation. Here we present the addition of live-cell Super-Resolution Optical Fluctuation Imaging (SOFI) to a previously reported correlative process using Single Molecule Localization Microscopy (SMLM) and Atomic Force Microscopy (AFM). SOFI was used with fluorescent proteins and low laser power to observe cellular ultrastructure in live COS-7 cells. SOFI-SMLM-AFM of microtubules showed minimal changes to the microtubule network in the 20 min between live-cell SOFI and fixation. Microtubule diameters were also analyzed through all microscopies; SOFI found diameters of 249 ± 68 nm and SMLM was 71 ± 33 nm. AFM height measurements found microtubules to protrude 26 ± 13 nm above the surrounding cellular material. The correlation of SMLM and AFM was extended to two-color SMLM to image both microtubules and actin. Two target SOFI was performed with various fluorescent protein combinations. rsGreen1-rsKAME, rsGreen1-Dronpa, and ffDronpaF-rsKAME fluorescent protein combinations were determined to be suitable for two target SOFI imaging. This correlative application of super-resolution live-cell and fixed-cell imaging revealed minimal artifacts created for the imaged target structures through the sample preparation procedure and emphasizes the power of correlative microscopy.

Differences in erythrocytes size and shape in prediabetes and diabetes assessed by two microscopy techniques and its association with dietary patterns. Pilot study.

Hemorheology and microcirculation alterations are caused by erythrocyte size and shape (ESS) modifications. People´s diets can alter erythrocyte functions and membrane fluidity by changing cell membrane components. The aim was to identify differences in ESS obtained by scanning electron (SEM) and atomic force microscopy (AFM) in people with prediabetes and type 2 diabetes (T2DM) and assess their relationship with dietary patterns. The study population included 31 participants (14 healthy, 11 with prediabetes, and 6 with T2DM). Dietary intake was assessed by a food frequency questionnaire, and dietary patterns were obtained using principal component analysis. ESS (diameter, height, axial ratio, thickness, and concave depth) were obtained by SEM and AFM. Differences in ESS between groups were observed with SEM (height) and AFM (height, axial ratio, and concave depth). T2DM presented smaller erythrocytes, more elongated and more altered forms. Two dietary patterns were identified: (1) Unhealthy: more refined cereals, high-fat dairy, fast food, sugary beverages, and fewer fruits, fish, seafood, low-fat dairy, and water. (2) Prudent: higher consumption of refined cereals, vegetables, poultry, low-fat dairy and nuts, and lower tortillas, eggs, high-fat dairy, and legumes. Tertile 3 of the Unhealthy dietary pattern had 80% of healthy participants. A difference in diameter and height (0.44 and 0.32 μm, respectively) obtained by SEM was observed when comparing tertile 2 (smaller erythrocytes) versus tertile 3 in the Unhealthy dietary pattern. SEM and AFM are excellent tools to assess ESS. Unhealthy dietary patterns might be associated with altered ESS. HIGHLIGHTS: SEM and AFM are excellent tools to assess erythrocyte size and shape modifications. Two dietary patterns were identified: healthy and prudent. Smaller erythrocytes were observed in the second tertile of the unhealthy pattern.

On the nature and propagation of errors in roughness parameters obtained from spectral analysis of atomic force microscopy topographic images

Scale-dependent surface roughness strongly affects critical surface properties of materials, including adhesion, wettability, and optical/thermal properties. As a particular example, tuning the ratio of the true to nominal area—a parameter that depends on the root mean square (RMS) local slope of the finest scales of topography—is an effective approach to tailor the wetting characteristics of solid surfaces. While power spectral density (PSD) analysis of atomic force microscopy (AFM) topographic images allows for directly assessing the scale-dependence of surface roughness, this approach to analyze AFM height maps requires power-law modeling and extrapolation of a PSD with inherently non-normal error distributions. Here, we use a Monte Carlo approach based on synthetic AFM images of known input power-law parameters to (1) evaluate the accuracy of fitting techniques based on the expected distribution of the PSD; (2) evaluate the error propagation from the standard errors of the fitted power-law parameters to the computed RMS slope and area ratio; and (3) evaluate the statistical power of various PSD regression techniques when differentiating surfaces. The results indicated that standard error for ordinary least squares on a log-log PSD (log OLS) underpredicts the observed variance by ∼50%. This underprediction can be eliminated by implementing a log-link gamma regression. Moreover, when propagating the standard error to derived parameters (e.g., the RMS slope), the propagated error is generally conservative relative to the observed variance and closely predicts the observed variance when extrapolating to the finest scale. This result demonstrates the possibility of accurately estimating roughness parameters that are critical for evaluating surface phenomena on the basis of fitting and extrapolating AFM data using self-affine models. Finally, our results provided evidence for the possibility of statistically differentiating surfaces with similar power-law parameters when using weighted gamma regression with a mean of 10 images, as opposed to unweighted log-OLS that requires as many as 10 000 images to differentiate images.

Detachable all-carbon-linked 3D covalent organic framework films for semiconductor/COF heterojunctions by continuous flow synthesis

Detachable all-carbon-linked 3D covalent organic framework films for semiconductor/COF heterojunctions by continuous flow synthesis

18.55% Efficiency Polymer Solar Cells Based on a Small Molecule Acceptor with Alkylthienyl Outer Side Chains and a Low-Cost Polymer Donor PTQ10

18.55% Efficiency Polymer Solar Cells Based on a Small Molecule Acceptor with Alkylthienyl Outer Side Chains and a Low-Cost Polymer Donor PTQ10

In Situ Supramolecular Polymerization via Organometallic-Catalyzed Macromolecular Metamorphosis

In Situ Supramolecular Polymerization via Organometallic-Catalyzed Macromolecular Metamorphosis

Polymer-solvent interaction and conformational changes at a molecular level: Implication to solvent-assisted deformation and aggregation at the polymer surface

HypothesisWe hypothesize that varying the chemical structure of the monomeric unit in a polymer will affect the surface structure and interfacial molecular group orientations of the polymer film leveraging its response to solvents of different chemical affinities. ExperimentsPoly (2-methoxy ethyl methacrylate) and poly (2-tertbutoxy ethyl methacrylate) thin films exposed to either deuterated water (D2O) or deuterated chloroform (CDCl3) were studied by sum frequency generation (SFG) spectroscopy, contact angle goniometry, and atomic force microscopy (AFM) at the polymer-solvent interface, supported with molecular simulation studies. FindingsSFG spectral analysis of the polymer thin films corroborated molecular re-organization at the surface when exposed to different chemical environments. The AFM height images of the polymer surfaces were homogeneously flat under CDCl3 and showed swollen regions under D2O. Following the removal of D2O, the exposed areas have imprinted, recessed locations and exposure to CDCl3 resulted in the formation of aggregates. The chemical affinity and characteristics of the solvents played a role in conformational change at the polymer surface. It had direct implications to interfacial processes involving adsorption, permeation which eventually leads to swelling, deformation or aggregation, and possibly dissolution.

Sub-micro scale cell segmentation using deep learning.

Automated cell segmentation is key for rapid and accurate investigation of cell responses. As instrumentation resolving power increases, clear delineation of newly revealed cellular features at the submicron through nanoscale becomes important. Reliance on the manual investigation of myriad small features retards investigation; however, use of deep learning methods has great potential to reveal cell features both at high accuracy and high speed, which may lead to new discoveries in the near term. In this study, semantic cell segmentation systems were investigated by implementing fully convolutional neural networks called U-nets for the segmentation of astrocytes cultured on poly-l-lysine-functionalized planar glass. The network hyperparameters were determined by changing the number of network layers, loss functions, and input image modalities. Atomic force microscopy (AFM) images were selected for investigation as these are inherently nanoscale and are also dimensional. AFM height, deflection, and friction images were used as inputs separately and together, and the segmentation performances were investigated on five-fold cross-validation data. Transfer learning methods, including VGG16, VGG19, and Xception, were used to improve cell segmentation performance. We find that AFM height images inherit more discriminative features than AFM deflection and AFM friction images for cell segmentation. When transfer-learning methods are applied, statistically significant segmentation performance improvements are observed. Segmentation performance was compared to classical image processing algorithms and other algorithms in use by considering both AFM and electron microscopy segmentation. An accuracy of 0.9849, Matthews correlation coefficient of 0.9218, and Dice's similarity coefficient of 0.9306 were obtained on the AFM test images. Performance evaluations show that the proposed system can be successfully used for AFM cell segmentation with high precision.

How high is a MoSe2 monolayer?

Transition metal dichalcogenides (TMDCs) have attracted significant attention for optoelectronic, photovoltaic and photoelectrochemical applications. The properties of TMDCs are highly dependent on the number of stacked atomic layers, which is usually counted post-fabrication, using a combination of optical methods and atomic force microscopy height measurements. Here, we use photoluminescence spectroscopy, Raman spectroscopy, and three different AFM methods to demonstrate significant discrepancies in height measurements of exfoliated MoSe2 flakes on SiO2 depending on the method used. We also highlight the often overlooked effect that electrostatic forces can be misleading when measuring the height of a MoSe2 flake using AFM.

Use of Atomic Force Microscopy (AFM) to monitor surface crystallization in caffeine-oxalic acid (CAFOXA) cocrystal compacts

Our objective was to monitor the surface crystallization in disordered caffeine-oxalic acid (CAFOXA) cocrystals following exposure to elevated water vapor pressure. This was accomplished using atomic force microscopy (AFM). Disorder was induced in the cocrystal particles by the common pharmaceutical unit operations of milling and compaction. The ‘activated’ solid, upon exposure to elevated water vapor pressure, had a high propensity to sorb water. This led to a rise in molecular mobility and the surface underwent rapid crystallization to form needle shaped crystals of CAFOXA. Using AFM height and phase imaging, we were able to directly visualize phase transformations on the compact surface. The milled compacts exhibited higher processing induced disorder than the unmilled compacts, thereby accelerating the surface recrystallization.

Systematic Investigation of Solution-State Aggregation Effect on Electrical Conductivity in Doped Conjugated Polymers

Systematic Investigation of Solution-State Aggregation Effect on Electrical Conductivity in Doped Conjugated Polymers

P-type InxGa1−xN semibulk templates (0.02 < x < 0.16) with room temperature hole concentration of mid-1019 cm−3 and device quality surface morphology

Using the semibulk approach, p-InxGa1−xN semibulk (p-SB) templates were grown with an indium content ranging from 2.4% to 15.2% via metalorganic chemical vapor deposition. When compared to optimized bulk p-GaN, the hole concentration in p-SB with an In content of ∼15.2% increased by two orders of magnitude from 5.22 × 1017 to 5.28 × 1019 cm−3. The resistivity and mobility of the templates decreased gradually from 3.13 Ω · cm and 3.82 cm2/V s for p-GaN to 0.24 Ω · cm and 0.48 cm2/V s for p-SB with an In content of 15.2%. Temperature dependent Hall measurements were conducted to estimate the activation energy of the p-SB template. The p-SB with the In content of ∼15.2% is estimated to have an activation energy of 29 meV. These heavily doped p-SB templates have comparable material qualities to that of GaN. The atomic force microscopy height retraces of p-SB films show device quality surface morphology, with root mean square roughness ranging from 2.53 to 4.84 nm. The current results can impact the performances of several nitride-based devices, such as laser diodes, LEDs, solar cells, and photodetectors.

Defect height estimation via model-less TSOM under optical resolution

We propose a new method of through-focus scanning optical microscopy (TSOM) without a reference database, i.e., a model-less TSOM method. Building a TSOM reference database is time-consuming or even impractical in some TSOM applications that involve complex structures, such as 3D NAND, or irregular shapes such as defects. The proposed model-less TSOM method was used to determine just the height of defect particles, for the first time as far as we are aware. Defect height is the only relevant dimension for the display panel application. Specifically, we analyzed 40 organic light-emitting diode (OLED) surface defects using a lab-developed motion-free TSOM tool consisting of a 50× objective lens (numerical aperture (NA) 0.55), a 532-nm light source, an imaging detector with a 7.5-µm pitch, and a deformable mirror. The tool is in-line and capable of achieving high throughput non-destructively, both relevant features for industrial applications. We investigated linear regression relations between newly defined TSOM parameters (TSOM height, TSOM area and TSOM volume) and the defect heights, which were first measured by atomic force microscopy (AFM). Following defect classification based on in-focus images, we successfully found that the AFM height has a linear correlation with 50% TSOM height (H50%) within ± 20.3 nm (1σ) error over the range of 140 to 950 nm. The one-sigma error, i.e., 20.3 nm, was approximately λ/26 or 1/43 of the depth of focus (DOF) of the applied microscope.

Patterning of transition metal dichalcogenides catalyzed by surface plasmons with atomic precision

Summary Plasmon-induced charge transfer has drawn considerable interests and spurred rapid developments of photovoltaics and photocatalysis. Various plasmonic metal/transition metal dichalcogenide (TMD) heterostructures have been developed to promote charge separation. For plasmon-induced catalysis, the transformation of TMDs is rare, and the erosion of TMDs has not yet been reported. Here, we report the etching of two-dimensional TMDs by planar surface plasmon polaritons (SPPs). We found that SPPs can etch various TMDs into desired layers and lateral size through controlling the light power and incident direction. In aqueous media, the strong plasmonic coupling at Au/MoS2 interface generates holes in the valence band of MoS2 that can weaken its interlayer interaction. As a synergistic effect, the plasmonic hot electrons produce oxidizing H2O2 to dissolve MoS2 from the top layers. Our results provide a new perspective for understanding the plasmonic coupling at metal-TMD interfaces and a reliable route toward fabricating well-defined TMDs.

Particle size distributions for cellulose nanocrystals measured by atomic force microscopy: an interlaboratory comparison

Particle size measurements of cellulose nanocrystals (CNCs) are challenging due to their broad size distribution, irregular shape and propensity to agglomerate. Particle size is one of the key parameters that must be measured for quality control purposes and to differentiate materials with different properties. We report the results of an interlaboratory comparison (ILC) which examined atomic force microscopy (AFM) data acquisition and data analysis protocols. Samples of CNCs deposited on poly-L-lysine coated mica were prepared in the pilot laboratory and sent to 10 participating laboratories including academic, government and industrial organizations with varying levels of experience with imaging CNCs. The participant data sets indicated that the central location, width and asymmetry varied considerably for both length and height distributions. To deal with this variability we used a skew normal distribution to model the data from each laboratory and to obtain the consensus distribution that describes the CNC particle size. The skew normal distribution has 3 parameters: a central location (mean), distribution width (standard deviation) and asymmetry (shape) factor. This approach gave consensus distributions with mean, standard deviation and asymmetry factor of 94.9 nm, 37.3 nm and 6.0 for length and 3.4 nm, 1.2 nm and 2.8 for height, respectively. The use of multiple probes and/or deterioration of the probe with increased use are significant contributing factors to the variability in mean length between laboratories. There is less variability in height across participating laboratories and tests of applied imaging force indicate that it is possible to image without significant compression of the CNCs. The number of CNCs necessary to obtain a reliable data set depends on the probes and operating conditions, but with careful control of various parameters analysis of 250 and 300 CNCs should provide consistent data sets for height and length, respectively for one sample. Comparison of AFM with transmission electron microscopy (TEM) data obtained in the same ILC demonstrated excellent agreement between measured lengths for the 2 methods. By contrast AFM height was approximately one half the TEM width, a result that indicates the presence of a significant number of laterally agglomerated particles, consistent with literature data.

Monolayer MoS2 on sapphire: an azimuthal reflection high-energy electron diffraction perspective

Molybdenum disulfide (MoS2) on the c-plane sapphire has been a very popular system to study in the two-dimensional (2D) materials community. Bottom-up synthesis of monolayer (ML) MoS2 with excellent electrical properties has been achieved on sapphire by various methods, making it a very promising candidate to be used in the next generation nano-electronic devices. However, large-area ML MoS2 with comparable quality as the relatively small size exfoliated ML remains quite a challenge. To overcome this bottle neck, a comprehensive understanding of the structure of the as-grown ML material is an essential first step. Here, we report a detailed structural characterization of wafer-scale continuous epitaxial ML MoS2 grown by metalorganic chemical vapor deposition on sapphire using an azimuthal reflection high-energy electron diffraction (ARHEED) technique. With ARHEED we can map not only 2D but also 3D reciprocal space structure of the ML statistically. From the oscillation in the ARHEED intensity profile along the vertical direction of the ML, we derived a real space distance of ~3 Å at the interface of ML and sapphire. Quantitative diffraction spot broadening analyses of the 3D reciprocal space map reveals low density defects and a small angular misalignment of orientation domains in ML MoS2. Based on atomic force microscopy height distribution analysis, cross-section scanning transmission electron microscopy, and density functional theory calculations, we suggest that there exists a passivation layer between MoS2 ML and sapphire substrate. This ARHEED methodology also has been applied to ML WS2 and is expected to be applicable to other ML transition metal dichalcogenides on arbitrary crystalline or non-crystalline substrates.

Light‐Induced and Oxygen‐Mediated Degradation Processes in Photoactive Layers Based on PTB7‐Th

Low‐bandgap polymers are sensitive to various degradation processes, which strongly decrease their lifetime. The chemical and physical changes occurring in the low‐bandgap polymer with benzodithiophene units poly[4,8‐bis(5‐(2‐ethylhexyl)thiophene‐2‐yl)benzo[1,2‐b:4,5‐b′]dithiophene‐2,6‐diyl‐alt‐(4‐(2‐ethylhexyl)‐3‐fluorothieno[3,4‐b]thiophene‐2‐carboxylate] (PTB7‐Th) and its blend with the fullerene derivative [6,6]‐phenyl‐C71‐butyric acid methyl ester (PC71BM) are followed during irradiation‐induced aging by combination of various characterization methods. The active layer morphology is investigated using atomic force microscopy (AFM) as well as in‐operando grazing incidence small angle X‐ray scattering (GISAXS), indicating morphological alterations and material loss due to chemical modifications. Optical spectroscopy gives insights into these chemical processes which lead to significant absorption losses under ambient conditions. Independent of the energy of the absorbed photons, but only in combination with oxygen, the excitation of the polymer leads to a fatal increase in oxidation probability. Fourier transform infrared (FTIR) data highlight the sensitivity of the conjugated polymer backbone to oxidation, a result of lost conjugation and therefore absorption capability. With combined AFM height and infrared (IR) mapping, the chemical degradation and material loss is confirmed on a nanoscale. Although the chemical structure is seriously damaged, the blend morphology is not undergoing major changes.

Formation of Higher Structural Levels in λ-Carrageenan Induced by the Antimalarial Drug Chloroquine.

The linear polysaccharide λ-carrageenan is the only one among the carrageenans not forming secondary, tertiary, and quaternary structures in the presence of inorganic ions. Chloroquine (CQ) is a well-established antimalaria drug also recently discussed in therapeutics against the COVID-19 pandemic. The interaction of this polysaccharide-ionic drug pair was investigated by combining UV-vis spectrophotometry and atomic force microscopy (AFM) imaging. A decrease of the UV peak assigned to free CQ and the occurrence of isosbestic points indicate the formation of complexes. High-resolution AFM height images revealed an increasing height of the single polysaccharide chains in the random coil state upon addition of CQ, indicating the formation of a secondary structure, followed by higher hierarchical aggregates. The disappearance of higher-ordered structures and the recovery of polysaccharide chains with primary structure were observed by introducing inorganic cations (Na+, K+, Ca2+), replacing the condensed CQ and paving the way to reversible ion-induced drug release.

Water Molecule-Triggered Anisotropic Deformation of Carbon Nitride Nanoribbons Enabling Contactless Respiratory Inspection

The exploitation of the interaction between nanostructured matter and small molecules, such as H2O at interfaces via dynamic hydrogen bonding, is essentially the key for smart, responsive nanodevic...