Regioselective Tip-Enhanced Raman Spectroscopy of Lipid Membranes with Sub-Nanometer Axial Resolution.

Noninvasive and label-free analysis of cell membranes at the nanoscale is essential to comprehend vital cellular processes. However, conventional analytical tools generally fail to meet this challenge due to the lack of required sensitivity and/or spatial resolution. Herein, we demonstrate that tip-enhanced Raman spectroscopy (TERS) is a powerful nanoanalytical tool to analyze dipalmitoylphosphatidylcholine (DPPC) bilayers and human cell membranes with submolecular resolution in the vertical direction. Unlike the far-field Raman measurements, TERS spectra of the DPPC bilayers reproducibly exhibited a uniquely shaped C-H band. These unique spectral features were also reproducibly observed in the TERS spectrum of human pancreatic cancer cells. Spectral deconvolution and DFT simulations confirmed that the TERS signal primarily originated from vibrations of the CH3 groups in the choline headgroup of the lipids. The reproducible TERS results obtained in this study unequivocally demonstrate the ultrahigh sensitivity of TERS for nanoanalysis of lipid membranes under ambient conditions.

Nanoscale Chemical Imaging of Human Cell Membranes Using Tip-Enhanced Raman Spectroscopy.

Lack of appropriate tools for visualizing cell membrane molecules at the nanoscale in a non-invasive and label-free fashion limits our understanding of many vital cellular processes. Here, we use tip-enhanced Raman spectroscopy (TERS) to visualize the molecular distribution in pancreatic cancer cell (BxPC-3) membranes in ambient conditions without labelling, with a spatial resolution down to ca. 2.5 nm. TERS imaging reveals segregation of phenylalanine-, histidine-, phosphatidylcholine-, protein-, and cholesterol-rich BxPC-3 cell membrane domains at the nm length-scale. TERS imaging also showed a cell membrane region where cholesterol is mixed with protein. Interestingly, the higher resolution TERS imaging revealed that the molecular domains observed on the BxPC-3 cell membrane are not chemically "pure" but also contain other biomolecules. These results demonstrate the potential of TERS for non-destructive and label-free imaging of cell membranes with nanoscale resolution.

Size-Dependent Interaction of Amyloid ß Oligomers with Brain Total Lipid Extract Bilayer-Fibrillation Versus Membrane Destruction.

Amyloid ß, Aß(1-42), is a component of senile plaques present in the brain of Alzheimer's disease patients and one of the main suspects responsible for pathological consequences of the disease. Herein, we directly visualize the Aß activity toward a brain-like model membrane and demonstrate that this activity strongly depends on the Aß oligomer size. PeakForce quantitative nanomechanical mapping mode of atomic force microscopy imaging revealed that the interaction of large-size (LS) Aß oligomers, corresponding to high-molecular-weight Aß oligomers, with the brain total lipid extract (BTLE) membrane resulted in accelerated Aß fibrillogenesis on the membrane surface. Importantly, the fibrillogenesis did not affect integrity of the membrane. In contrast, small-size (SS) Aß oligomers, corresponding to low-molecular-weight Aß oligomers, created pores and then disintegrated the BTLE membrane. Both forms of the Aß oligomers changed nanomechanical properties of the membrane by decreasing its Young's modulus by ∼45%. Our results demonstrated that both forms of Aß oligomers induce the neurotoxic effect on the brain cells but their action toward the membrane differs significantly.

Probing Chemical Complexity of Amyloid Plaques in Alzheimer's Disease Mice using Hyperspectral Raman Imaging.

One of the distinctive pathological features of Alzheimer's disease (AD) is the deposition of amyloid plaques within the brain of affected individuals. These plaques have traditionally been investigated using labeling techniques such as immunohistochemical imaging. However, the use of labeling can disrupt the structural integrity of the molecules being analyzed. Hence, it is imperative to employ label-free imaging methods for noninvasive examination of amyloid deposits in their native form, thereby providing more relevant information pertaining to AD. This study presents compelling evidence that label-free and nondestructive confocal Raman imaging is a highly effective approach for the identification and chemical characterization of amyloid plaques within cortical regions of an arcAß mouse model of AD. Furthermore, this investigation elucidates how the spatial correlation of Raman signals can be exploited to identify robust Raman marker bands and discern proteins and lipids from amyloid plaques. Finally, this study uncovers the existence of distinct types of amyloid plaques in the arcAß mouse brain, exhibiting significant disparities in terms of not only shape and size but also molecular composition.

Nanoscale chemical analysis of 2D molecular materials using tip-enhanced Raman spectroscopy.

Two-dimensional (2D) molecular materials have attracted immense attention due to their unique properties, promising a wide range of exciting applications. To understand the structure-property relationship of these low-dimensional materials, sensitive analytical tools capable of providing structural and chemical characterisation at the nanoscale are required. However, most conventional analytical techniques fail to meet this challenge, especially in a label-free and non-destructive manner under ambient conditions. In the last two decades, tip-enhanced Raman spectroscopy (TERS) has emerged as a powerful analytical technique for nanoscale chemical characterisation by combining the high spatial resolution of scanning probe microscopy and the chemical sensitivity and specificity of surface-enhanced Raman spectroscopy. In this review article, we provide an overview of the application of TERS for nanoscale chemical analysis of 2D molecular materials, including 2D polymers, biomimetic lipid membranes, biological cell membranes, and 2D reactive systems. The progress in the structural and chemical characterisation of these 2D materials is demonstrated with key examples from our as well as other laboratories. We highlight the unique information that TERS can provide as well as point out the common pitfalls in experimental work and data interpretation and the possible ways of averting them.

Analyzing Morphological Properties of Early-Stage Toxic Amyloid ß Oligomers by Atomic Force Microscopy.

Protein misfolding diseases, like Alzheimer's, Parkinson's, and Huntington's disease, are associated with misfolded protein aggregation. Alzheimer's disease is related to a progressive neuronal death induced by small amyloid ß oligomers. Here, we describe the procedure to prepare and identify different types of small toxic amyloid ß oligomers by atomic force microscopy (AFM).

Amyloid ß interaction with model cell membranes - What are the toxicity-defining properties of amyloid ß?

Disruption of the neuronal membrane by toxic amyloid ß oligomers is hypothesized to be the major event associated with Alzheimer's disease's neurotoxicity. Misfolding of amyloid ß is followed by aggregation via different pathways in which structurally different amyloid ß oligomers can be formed. The respective toxic actions of these structurally diverse oligomers can vary significantly. Linking a particular toxic action to a structurally unique kind of amyloid ß oligomers and resolving their toxicity-determining feature remains challenging because of their transient stability and heterogeneity. Moreover, the lipids that make up the membrane affect amyloid ß oligomers' behavior, thus adding to the problem's complexity. The present review compares and analyzes the latest results to improve understanding of amyloid ß oligomers' interaction with lipid bilayers.

Visualizing Surface Phase Separation in PS-PMMA Polymer Blends at the Nanoscale.

Phase-separated polymer blend films are an important class of functional materials with numerous technological applications in solar cells, catalysis, and biotechnology. These technologies are underpinned by the precise control of phase separation at the nanometer length-scales, which is highly challenging to visualize using conventional analytical tools. Herein, we introduce tip-enhanced Raman spectroscopy (TERS), in combination with atomic force microscopy (AFM), confocal Raman spectroscopy, and X-ray photoelectron spectroscopy (XPS), as a sensitive nanoanalytical method to determine lateral and vertical phase-separation in polystyrene (PS)-poly(methyl methacrylate) (PMMA) polymer blend films. Correlative topographical, molecular, and elemental information reveals a vertical phase separation of the polymers within the top ca. 20 nm of the blend surface in addition to the lateral phase separation in the bulk. Furthermore, complementary TERS and XPS measurements reveal the presence of PMMA within 9.2 nm of the surface and PS at the subsurface of the polymer blend. This fundamental work establishes TERS as a powerful analytical tool for surface characterization of this important class of polymers at nanometer length scales.

Bimetallic nanocatalysts supported on graphitic carbon nitride for sustainable energy development: the shape-structure-activity relation.

The catalytic performance of metal nanoparticles (NPs), including activity, selectivity, and durability, depends on their shape and structure at the molecular level. Consequently, metal NPs of different size and shape, e.g., nanobelts, nanocubes, nanoflakes, and nanowires, demonstrate different reactivity and provide different reaction rates depending on the facet exposed. In this context, the present review aims to summarize the shape-structure-activity relation of metallic nanocatalysts. Moreover, keeping in mind that the application of noble metal catalysts is expensive, we would like to draw the reader's attention to bimetallic nanocatalysts supported on graphitic carbon nitride. One of the advantages of these systems is the possibility to minimize the use of noble metals by introducing another metal either to the parent NPs and/or modifying the support materials. The development and optimization of bimetallic nanocatalysts might provide the new class of materials with superior, tunable performance, thermal stability and reduced costs compared to presently available commercial catalysts. Therefore, further application of these bimetallic composites for sustainable development in energy, green chemicals/fuels and environmental protection will be discussed.

Correction: Bimetallic nanocatalysts supported on graphitic carbon nitride for sustainable energy development: the shape-structure-activity relation.

[This corrects the article DOI: 10.1039/D0NA01063D.].

Inhibition of Amyloid ß-Induced Lipid Membrane Permeation and Amyloid ß Aggregation by K162.

Alzheimer's disease (AD) is characterized by progressive neurodegeneration associated with amyloid ß (Aß) peptide aggregation. The aggregation of Aß monomers (AßMs) leads to the formation of Aß oligomers (AßOs), the neurotoxic Aß form, capable of permeating the cell membrane. Here, we investigated the effect of a fluorene-based active drug candidate, named K162, on both Aß aggregation and AßO toxicity toward the bilayer lipid membrane (BLM). Electrochemical impedance spectroscopy (EIS), atomic force microscopy (AFM), and molecular dynamics (MD) were employed to show that K162 inhibits AßOs-induced BLM permeation, thus preserving BLM integrity. In the presence of K162, only shallow defects on the BLM surface were formed. Apparently, K162 modifies Aß aggregation by bypassing the formation of toxic AßOs, and only nontoxic AßMs, dimers (AßDs), and fibrils (AßFs) are produced. Unlike other Aß toxicity inhibitors, K162 preserves neurologically beneficial AßMs. This unique K162 inhibition mechanism provides an alternative AD therapeutic strategy that could be explored in the future.

Alzheimer's disease-related amyloid ß peptide causes structural disordering of lipids and changes the electric properties of a floating bilayer lipid membrane.

Neurodegeneration in Alzheimer's disease is associated with disruption of the neuronal cell membrane by the amyloid ß (Aß) peptide. However, the disruption mechanism and the resulting changes in membrane properties remain to be elucidated. To address this issue, herein the interaction of amyloid ß monomers (AßMs) and amyloid ß oligomers (AßOs) with a floating bilayer lipid membrane (fBLM) was studied using electrochemical and IR spectroscopy techniques. IR measurements showed that both Aß forms interacted similarly with the hydrophobic membrane core (lipid acyl chains), causing conformational and orientational changes of the lipid acyl chains, thus decreasing acyl chain mobility and altering the lipid packing unit cell. In the presence of AßOs, these changes were more significant than those in the presence of AßMs. However, respective interactions of AßMs and AßOs with the membrane hydrophilic exterior (lipid heads) were quite different. AßMs dehydrated lipid heads without affecting their orientation while AßOs changed the orientation of lipid heads keeping their hydration level intact. Electrochemical measurements showed that only AßOs porated the fBLM, thus significantly changing the fBLM electrical properties. The present results provide new molecular-level insight into the mechanism of membrane destruction by AßOs and changes in the membrane properties.