Renato Zenobi

Nanoscale chemical analysis by tip-enhanced Raman spectroscopy

Abstract A fine metal tip brought to within a few nanometers of a molecular film is found to give strong enhancement of Raman scattered light from the sample. This new principle can be used for molecular analysis with excellent spatial resolution, only limited by the tip apex size and shape. No special sample preparation is required, and the enhancement is identical at every sample location, allowing for quantitative surface-enhanced Raman spectroscopy measurements. When scanning the tip over the sample surface, topographic information is obtained simultaneously and can be directly correlated with the spectroscopic data.

Ion formation in MALDI mass spectrometry

The origin of ions in matrix-assisted laser desorption/ionization (MALDI) mass spectrometry is currently a matter of active research. A number of chemical and physical pathways have been suggested for MALDI ion formation, including gas-phase photoionization, ion–molecule reactions, disproportionation, excited-state proton transfer, energy pooling, thermal ionization, and desorption of preformed ions. These pathways and others are critically reviewed, and their varying roles in the wide variety of MALDI experiments are discussed. An understanding of ionization pathways should help to maximize ion yields, control analyte charge states and fragmentation, and gain access to new classes of analytes.

Scanning near-field optical microscopy with aperture probes: Fundamentals and applications

In this review we describe fundamentals of scanning near-field optical microscopy with aperture probes. After the discussion of instrumentation and probe fabrication, aspects of light propagation in metal-coated, tapered optical fibers are considered. This includes transmission properties and field distributions in the vicinity of subwavelength apertures. Furthermore, the near-field optical image formation mechanism is analyzed with special emphasis on potential sources of artifacts. To underline the prospects of the technique, selected applications including amplitude and phase contrast imaging, fluorescence imaging, and Raman spectroscopy, as well as near-field optical desorption, are presented. These examples demonstrate that scanning near-field optical microscopy is no longer an exotic method but has matured into a valuable tool.

Quantitative determination of noncovalent binding interactions using soft ionization mass spectrometry

Abstract For a number of years, soft ionization mass spectrometry has been used for studying noncovalently bound complexes. An intriguing question in this context is whether MS experiments can be used to measure the interaction strength. A number of recent studies have addressed this question. The results of these studies, as well as the methods employed are reviewed here. We distinguish between liquid-phase methods such as mass spectrometrically detected melting curves, titration experiments, or competition experiments, and gas-phase methods such as cone voltage-driven dissociation, collision-induced dissociation, blackbody infrared radiative dissociation, or thermal dissociation of gas-phase complex ions. With a few exceptions, no agreement exists between solution-phase and gas-phase binding energies. The main reason is that electrostatic and dipolar noncovalent interactions are strengthened in the absence of solvent shielding, while other noncovalent interactions, in particular hydrophobic interactions, become less important in the absence of solvent. The possibility to quantitatively measure solution-phase as well as gas-phase noncovalent interaction strengths by mass spectrometry opens fascinating perspectives for very high sensitivity screening assays as well as for improved fundamental understanding of the nature of noncovalent interactions.

High-quality near-field optical probes by tube etching

A method called tube etching for the fabrication of near-field optical probes is presented. Tip formation occurs inside a cylindrical cavity formed by the polymer coating of an optical fiber which is not stripped away prior to etching in hydrofluoric acid. The influence of temperature, etchant concentration, and fiber type on the tip quality is studied. A tip formation mechanism for the given geometry is proposed. The procedure overcomes drawbacks of the conventional etching techniques while still producing large cone angles: (i) tips with reproducible shapes are formed in a high yield, (ii) the surface roughness on the taper is drastically reduced, and (iii) the tip quality is insensitive to vibrations and temperature fluctuations during the etching process. After aluminum coating, optical probes with well-defined apertures are obtained. Due to the smooth glass surface the aluminum coating is virtually free of pinholes.

Single-Cell Metabolomics: Analytical and Biological Perspectives

Background In recent years, there has been a surge in the development and application of single-cell genomics, transcriptomics, proteomics, and metabolomics. The metabolome is defined as the full complement of small-molecule metabolites found in a specific cell, organ, or organism. The most interesting potential application of single-cell metabolomics may be in the area of cancer—for example, identification of circulating cancer cells that lead to metastasis. Other fields where single-cell metabolomics is expected to have an impact are systems biology, stem cell research, aging, and the development of drug resistance; more generally, it could be used to discover cells’ chemical strategies for coping with chemical or environmental stress. Relative to other single-cell “-omics” measurements, metabolomics provides a more immediate and dynamic picture of the functionality (i.e., of the phenotype) of a cell, but is arguably also the most difficult to measure. This is because the metabolome can dynamically react to the environment on a very short time scale (seconds or less), because of the large structural diversity and huge dynamic range of metabolites, because it is not possible to amplify metabolites, and because tagging them with fluorescent labels would distort their normal function. Single-cell analysis uses a wide variety of imaging and chemical analysis methods to study vastly different cell types and sizes. (A) Closterium acerosum (algal cells, ~300 μm × 40 μm; optical micrograph). (B) Euglena gracilis (algal cells, diameter ~20 μm); Raman image of β-carotene distribution (left) and fluorescence emission from proplastids (right)

What Can We Learn from Ambient Ionization Techniques

Ambient mass spectrometry—mass spectrometric analysis with no or minimal effort for sample preparation—has experienced a very rapid development during the last 5 years, with many different methods now available for ionization. Here, we review its range of applications, the hurdles encountered for its quantitative use, and the proposed mechanisms for ion formation. Clearly, more effort needs to be put into investigation of matrix effects, into defining representative sampling of heterogeneous materials, and into understanding and controlling the underlying ionization mechanisms. Finally, we propose a concept to reduce the number of different acronyms describing very similar embodiments of ambient mass spectrometry.

Nanoscale Chemical Imaging Using Tip‐Enhanced Raman Spectroscopy: A Critical Review

Methods for chemical analysis at the nanometer scale are crucial for understanding and characterizing nanostructures of modern materials and biological systems. Tip-enhanced Raman spectroscopy (TERS) combines the chemical information provided by Raman spectroscopy with the signal enhancement known from surface-enhanced Raman scattering (SERS) and the high spatial resolution of atomic force microscopy (AFM) or scanning tunneling microscopy (STM). A metallic or metallized tip is illuminated by a focused laser beam and the resulting strongly enhanced electromagnetic field at the tip apex acts as a highly confined light source for Raman spectroscopic measurements. This Review focuses on the prerequisites for the efficient coupling of light to the tip as well as the shortcomings and pitfalls that have to be considered for TERS imaging, a fascinating but still challenging way to look at the nanoworld. Finally, examples from recent publications have been selected to demonstrate the potential of this technique for chemical imaging with a spatial resolution of approximately 10 nm and sensitivity down to the single-molecule level for applications ranging from materials sciences to life sciences.

The Matrix Suppression Effect and Ionization Mechanisms in Matrix‐assisted Laser Desorption/Ionization

At appropriate matrix:analyte mixing ratios, small to moderate sized analyte ions (1000–20 000 u) can fully suppress positively charged matrix ions in matrix-assisted laser desorption/ionization (MALDI) mass spectra. This is true for all matrix species, including radical cations and adducts with protons or alkali-metal ions. Full matrix suppression is also observed, regardless of the preferred analyte ion form, be it protonated or an alkali adduct. These facts lead us to propose a mechanism for prompt, primary (not secondary gas-phase) MALDI ionization in which excited matrix molecules are the key species. At least two such excited molecules are believed to be necessary for free ion generation. This model is found to be consistent with the available data, as well as making several predictions which are confirmed by new observations. The model also predicts that the matrix suppression effect will not be observable with heavy analytes because their large excluded volume precludes desorption at the necessary mixing ratios.

Nanoscale chemical imaging using top-illumination tip-enhanced Raman spectroscopy.

We present a new top-illumination scheme for tip-enhanced Raman spectroscopy (TERS) in a gap-mode configuration with illumination and detection in a straightforward fashion perpendicular to the sample surface. This illumination focuses the light tightly around the tip end, which effectively diminishes far-field background contributions during TERS measurements. The setup maintains the entire functionality range of both the scanning probe microscopy and the confocal optical microscopy of the setup. For the first time, we show large (64 × 64 up to 200 × 200 pixels), high-resolution TERS imaging with full spectral information at every pixel, which is necessary for the chemical identification of sample constituents. With a scanning tunneling microscope tip and feedback, these TERS maps can be recorded with a resolution better than 15 nm (most likely even less, as discussed with Figure 6). An excellent enhancement (∼10(7)×, sufficient for detection of few molecules) allows short acquisition times (<<1 s/pixel) and reasonably low laser power (in the microwatt regime) yielding spectroscopic images with high pixel numbers in reasonable time (128 × 128 pixels in <25 min). To the best of our knowledge, no Raman maps with similar pixel numbers and full spectral information have ever been published.