Atomic Force Microscopy for the Evaluation of Corneal Surface Roughness after Femtosecond Laser Flap Creation and Excimer Ablation.

Introduction: It is well known that the femtosecond laser lamellar cut has some degree of roughness. Nevertheless, as in femtosecond laser assisted LASIK (FS-LASIK), an excimer LASIK ablation is performed, the post-ablation stromal bed should show a marked decrease in roughness. We decided to compare, using atomic force microscopy (AFM), the roughness of the corneal stromal bed, after a Femtosecond lasers device (FS) flap was created with or without an excimer myopic ablation.Methods: Using 6 freshly enucleated porcine eyes, we created in every eye a flap using a femtosecond laser. Additionally in 3 eyes an excimer laser ablation to correct -3 diopters (D) was made. AFM imaging of the remaining corneal stroma was performed. Ten different square areas of 20μm x 20μm at the central area of the stroma of each corneal sample were studied. The roughness parameters used was the root-mean-square (RMS) deviation from a perfectly flap plane.Results: The RMS deviation were 360 ± 120 nm in femtosecond laser only, and 110 ± 20 nm in those cases where excimer is also involved (p<0.0001).Conclusions: Our results show that the roughness of the surface treated with excimer is clearly lower than in the group with no excimer ablation, thus the application of laser excimer after a flap creation by femtosecond laser flap creation may soften the nano-irregularities created by this technique.

Hydration Dynamics and the Future of Small-Amplitude AFM Imaging in Air

Here, we discuss the effects that the dynamics of the hydration layer and other variables, such as the tip radius, have on the availability of imaging regimes in dynamic AFM—including multifrequency AFM. Since small amplitudes are required for high-resolution imaging, we focus on these cases. It is possible to fully immerse a sharp tip under the hydration layer and image with amplitudes similar to or smaller than the height of the hydration layer, i.e., ~1 nm. When mica or HOPG surfaces are only cleaved, molecules adhere to their surfaces, and reaching a thermodynamically stable state for imaging might take hours. During these first hours, different possibilities for imaging emerge and change, implying that these conditions must be considered and reported when imaging.

Evaluating Effect of Metallic Ions On Aggregation Behavior of Amyloid Aβ42 By AFM Imaging and SERS

Background: Excessive aggregation of β-amyloid peptides (Aβ) is regarded as the hallmark of Alzheimer’s disease. Exploring the underlying mechanism regulating Aβ aggregation remains challenging and investigating aggregation events of Aβ in the presence and absence of metal ions at molecular level would be meaningful in elucidating the role of metal cations on interactions between Aβ molecules. In this study, chemical self-assembled monolayer (SAM) method was employed to fabricate monolayer of β-amyloid peptides Aβ42 on gold substrate with a bolaamphiphile named 16-Mercaptohexadecanoic acid (MHA). Firstly, the samples of gold substrate (blank control), the MHA-modified substrate and the Aβ42-modified substrate were detected by X-ray photoelectron spectroscopy (XPS) to track the self-assembly process. Aggregation behaviors of Aβ42 before and after metallic ions (Zn2+、Ca2+、Al3+) treated were monitored by atomic force microscopy (AFM) and the interaction between Aβ42 and metallic ions (Zn2+、Ca2+、Al3+) was investigated by surface-enhanced Raman Scattering (SERS), respectively.Results: The XPS spectra of binding energy of gold substrate (blank control), the MHA-modified substrate and the Aβ42-modified substrate are well fitted with the corresponding monolayer’s composition, which indicates that Aβ42 monolayer is well formed. The recorded surface morphology of different experimental groups obtained by AFM showed markedly different nanostructures, indicating occurrence of aggregation events between Aβ42 molecules after adding metal ions to the solution. Compared to the control group, the presence of metal ions resulted in the increased size of surface structures on the observed 3D topography. Further study by SERS showed that the Raman strength of Aβ42 changes significantly after the metal cation treatment. A considerable part of the amide bonds interacts with metal cations, leading to a structural change, which is characterized by the weakened β-fold Raman peak.Conclusion: The AFM imaging results suggest that aggregation events occurred between Aβ42 molecules with the addition of metal cations. Furthermore, the effect of metallic cations on the conformational change of Aβ42 studied by SERS supported the results obtained by AFM imaging. Taken together, the results showed that the presence of substoichiometric metal cations promotes aggregation behavior between Aβ42 molecules on the substrate at pH 7.4.

Accurate Morphology Characterization Using Atomic Force Microscopy via Vertical Drift Correction and Illusory Slope Elimination

Thanks to the ability to perform imaging and manipulation at the nanoscale, atomic force microscopy (AFM) has been widely used in biology, materials, chemistry, and other fields. However, as common error sources, vertical drift and illusory slope severely impair AFM imaging quality. To address this issue, this paper proposes a robust algorithm to synchronously correct the image distortion caused by vertical drift and slope, thus achieving accurate morphology characterization. Specifically, to eliminate the damage of abnormal points and feature areas on the correction accuracy, the laser spot voltage error acquired in the AFM scanning process is first utilized to preprocess the morphology height data of the sample, so as to obtain the refined alternative data suitable for line fitting. Subsequently, this paper proposes a novel line fitting algorithm based on sparse sample consensus, which accurately simulates vertical drift and slope in the cross-sectional profile of the topographic image, thereby achieving effective correction of the image distortion. In the experiments and applications, a nanoscale optical grating sample and a biological cell sample are adopted to perform topography imaging and distortion correction, so as to verify the ability of the proposed algorithm to promote AFM imaging quality.

Simulation atomic force microscopy to predict correlated conformational dynamics in proteins from topographic imaging

Atomic force microscopy (AFM) of proteins can detect only changes within the scanned molecular surface, missing all motions in other regions and thus information about functionally relevant conformational couplings. We show that simulation AFM can overcome this drawback by reconstruction of 3D molecular structures from topographic AFM images. A proof of principle demonstration is provided for an in-silico AFM experiment visualizing the conformational dynamics of a membrane transporter. The application shows that the alternating access mechanism underlying its operation can be retrieved from only AFM imaging of one membrane side. Simulation AFM is implemented in the freely available BioAFMviewer software platform, providing the convenient applicability to better understand experimental AFM observations.