Improvement of biomolecular analysis in thin films using in situ matrix enhanced secondary ion mass spectrometry.

Sensitivity to molecular ions remains a limiting factor for high resolution imaging mass spectrometry of organic and biological materials. Here, we investigate a variant of matrix-enhanced secondary ion mass spectrometry in which the transfer of matrix molecules to the analyte sample is carried out in situ (in situ ME-SIMS). This approach is therefore compatible with both 2D and 3D imaging by SIMS. In this exploratory study, nanoscale matrix layers were sputter-transferred inside our time-of-flight (ToF)-SIMS to a series of thin films of biomolecules (proteins, sugars, lipids) adsorbed on silicon, and the resulting layers were analyzed and depth-profiled. For this purpose, matrix molecules were desorbed from a coated target (obtained by drop-casting or sublimation) using 10 keV Ar3000+ ion beam sputtering, followed by redeposition on a collector carrying the sample to be analyzed. After evaluating the quality of the transfer of six different matrices on bare Si collectors, α-cyano-4-hydroxycinnamic acid (CHCA) was selected for further experiments. The mass spectra and depth profiles obtained from the organic layer prior to and after the sputter-transfer of CHCA were compared, along with those obtained from regular ME-SIMS samples (dried droplets) and, finally, with MALDI data for the same matrix-analyte combinations. Signal amplification factors were calculated by dividing the integrated molecular intensities obtained with or without matrix transfer. While the amplification factors are between 0.5 and 2 for molecules already detected with high intensities in SIMS, such as cholesterol or human angiotensin, other compounds show very large integrated signal amplification, even above two orders of magnitude. This is the case for D-glucose and cardiolipin, for which the molecular ion intensity is low (or very low) under normal SIMS analysis conditions. For such low ionization probability compounds, the beneficial effect of the matrix is unquestionable. Test experiments on mouse brain tissue sections also indicate signal enhancement with the matrix, especially for high mass lipid ions.

STARGATE: A new instrument for high-resolution photodissociation spectroscopy of cold ionic species.

Spectroscopy of transient anions and radicals by gated and accelerated time-of-flight experiment is a new spectrometer developed in UCLouvain. This instrument measures high-resolution photodissociation spectra of mass-selected ions by the combination of a time-of-flight spectrometer including a specific gating, bunching, and re-referencing unit with a nanosecond pulsed dye laser, a pulsed deflection, and an energy selector. The ionic species are generated in a supersonic jet expansion by means of an electric discharge or by the impact of electrons coming from an electron gun. The versatility of the molecular systems that can be addressed by this instrument is illustrated by the presentation of mass spectra of cations, anions, and ionic clusters formed from different gas mixtures and backing pressures. The high-resolution spectrum of the A~2Σ+(002)←X~2Π3/2(000) and A~2Σ+(002)←X~2Π1/2(000) rovibronic bands of N2O+ has been measured and analyzed to provide refined molecular parameters in the A~2Σ+(002) upper state. The A~2Σ+(002)←X~2Π3/2(000) band has been used to evaluate the quality of the experimental setup in terms of rotational temperature, time of measurement for certain signal to noise ratio, and the accuracy of the determination of the wavenumber scale.

Deposition of Intact and Active Proteins In Vacuo Using Large Argon Cluster Ion Beams.

Providing inert materials with a biochemical function, for example using proteins, is a cornerstone technology underlying many applications. However, the controlled construction of protein thin films remains a major challenge. Here, an innovative solvent-free approach for protein deposition is reported, using lysozyme as a model. By diverting a time-of-flight secondary ion mass spectrometer (ToF-SIMS) from its standard analytical function, large argon clusters were used to achieve protein transfer. A target consisting of a pool of proteins was bombarded with 10 keV Ar5000+ ions, and the ejected proteins were collected on a silicon wafer. The ellipsoidal deposition pattern was evidenced by ToF-SIMS analysis, while SDS-PAGE electrophoresis confirmed the presence of intact lysozyme on the collector. Finally, enzymatic activity assays demonstrated the preservation of the three-dimensional structure of the transferred proteins. These results pave the way to well-controlled protein deposition using ion beams and to the investigation of more complex multilayer architectures.

Large cluster ions: soft local probes and tools for organic and bio surfaces.

Ionised cluster beams have been produced and employed for thin film deposition and surface processing for half a century. In the last two decades, kiloelectronvolt cluster ions have also proved to be outstanding for surface characterisation by secondary ion mass spectrometry (SIMS), because their sputter and ion yields are enhanced in a non-linear fashion with respect to monoatomic projectiles, with a resulting step change of sensitivity for analysis and imaging. In particular, large gas cluster ion beams, or GCIB, have now become a reference in organic surface and thin film analysis using SIMS and X-ray photoelectron spectroscopy (XPS). The reason is that they induce soft molecular desorption and offer the opportunity to conduct damageless depth-profiling and 3D molecular imaging of the most sensitive organic electronics and biological samples, with a nanoscale depth resolution. In line with these recent developments, the present review focuses on rather weakly-bound, light-element cluster ions, such as noble or other gas clusters, and water or alcohol nanodroplets (excluding clusters made of metals, inorganic salts or ionic liquids) and their interaction with surfaces (essentially, but not exclusively, organic). The scope of this article encompasses three aspects. The first one is the fundamentals of large cluster impacts with surfaces, using the wealth of information provided by molecular dynamics simulations and experimental observations. The second focus is on recent applications of large cluster ion beams in surface characterisation, including mass spectrometric analysis and 2D localisation of large molecules, molecular depth-profiling and 3D molecular imaging. Finally, the perspective explores cutting edge developments, involving (i) new types of clusters with a chemistry designed to enhance performance for mass spectrometry imaging, (ii) the use of cluster fragment ion backscattering to locally retrieve physical surface properties and (iii) the fabrication of new biosurface and thin film architectures, where large cluster ion beams are used as tools to transfer biomolecules in vacuo from a target reservoir to any collector substrate.