A novel application of time-of-flight secondary ion mass spectrometry (ToF-SIMS) with continuous Ar cluster beams to peptide analysis was investigated. In order to evaluate peptide structures, it is necessary to detect fragment ions related to multiple neighbouring amino acid residues. It is, however, difficult to detect these using conventional ToF-SIMS primary ion beams such as Bi cluster beams. Recently, C60 and Ar cluster ion beams have been introduced to ToF-SIMS as primary ion beams and are expected to generate larger secondary ions than conventional ones. In this study, two sets of model peptides have been studied: (des-Tyr)-Leu-enkephalin and (des-Tyr)-Met-enkephalin (molecular weights are approximately 400 Da), and [Asn(1) Val(5)]-angiotensin II and [Val(5)]-angiotensin I (molecular weights are approximately 1,000 Da) in order to evaluate the usefulness of the large cluster ion beams for peptide structural analysis. As a result, by using the Ar cluster beams, peptide molecular ions and large fragment ions, which are not easily detected using conventional ToF-SIMS primary ion beams such as Bi3 (+), are clearly detected. Since the large fragment ions indicating amino acid sequences of the peptides are detected by the large cluster beams, it is suggested that the Ar cluster and C60 ion beams are useful for peptide structural analysis.

Previous studies have shown that the use of a 20 keV water cluster beam as a primary beam for the analysis of organic and bio-organic systems resulted in a 10−100 times increase in positive molecular ion yield for a range of typical analytes compared to C 60 and argon cluster beams. This resulted in increased sensitivity to important lipid molecules in the bioimaging of rat brain. Building on these studies, the present work compares 40 and 70 keV water cluster beams with cluster beams composed of pure argon, argon and 10%CO 2 , and pure CO 2 . First, as previously, we show that for E/nucleon about 0.3 eV/nucleon water and nonwater containing cluster beams generate very similar ion yields, but below this value, the water beams yields of BOTH negative and positive "molecular" ions increase, in many cases reaching a maximum in the <0.2 region, with yield increases of ∼10−100. Ion fragment yields in general decrease quite dramatically in this region. Second, for water cluster beams at a constant E/nucleon, "molecular" ion yield increases with beam energy and hence cluster size due to increased sputter yield (ionization probability is constant). Third, as a consequence of the increased ion yield and the improved focusability using high-energy cluster beams, imaging in the 1 μm spatial resolution region is demonstrated on HeLa cells and rat brain tissue, monitoring molecules that were previously difficult to detect with other primary beams. Finally, the suggestion that the secondary ion emission zone has quasi-aqueous character seems to be sustained.

 Understanding the mechanisms of iron trafficking in plants is key to enhancing the nutritional quality of crops. Because it is difficult to image iron in transit, we currently have an incomplete picture of the route(s) of iron translocation in developing seeds and how the tissue-specific distribution is established.  We have used a novel approach, combining 57 Fe isotope labelling and Nanoscale Secondary Ion Mass Spectrometry (NanoSIMS), to visualize iron translocation between tissues and within cells in immature wheat grain, Triticum aestivum L.  This enabled us to track the main route of iron transport from maternal tissues to the embryo through the different cell types. Further evidence for this route was provided by genetically diverting iron into storage vacuoles, with confirmation provided by histological staining and TEM-EDS. Almost all iron in both control and transgenic grains was found in intracellular bodies, indicating symplastic rather than apoplastic transport. Furthermore, a new type of iron body, highly enriched in 57 Fe, was observed in aleurone cells and may represent iron being delivered to phytate globoids.  Correlation of the 57 Fe enrichment profiles obtained by NanoSIMS with tissue-specific gene expression provides an updated model of iron homeostasis in cereal grains with relevance for future biofortification strategies.

RationaleTo discover the degree to which water‐containing cluster beams increase secondary ion yield and reduce the matrix effect in time‐of‐flight secondary ion mass spectrometry (TOF‐SIMS) imaging of biological tissue.MethodsThe positive SIMS ion yields from model compounds, mouse brain lipid extract and mouse brain tissue together with mouse brain images were compared using 20 keV C60 +, Ar2000 +, water‐doped Ar2000 + and pure (H2O)6000 + primary beams.ResultsWater‐containing cluster beams where the beam energy per nucleon (E/nucleon) ≈ 0.2 eV are optimum for enhancing ion yields dependent on protonation. Ion yield enhancements over those observed using Ar2000 + lie in the range 10 to >100 using the (H2O)6000 + beam, while with water‐doped (H2O)Ar2000 + they lie in the 4 to 10 range. The two water‐containing beams appear to be optimum for tissue imaging and show strong evidence of increasing yields from molecules that experience matrix suppression under other primary beams.ConclusionsThe application of water‐containing primary beams is suggested for biological SIMS imaging applications, particularly if the beam energy can be raised to 40 keV or higher to further increase ion yield and enhance spatial resolution to ≤1 µm. © 2015 The Authors. Rapid Communications in Mass Spectrometry Published by John Wiley & Sons Ltd.

a Low ionization yields in time of flight secondary ion mass spectrometry (ToF-SIMS) particularly from single cells and tissues are proving to be a significant limitation in allowing this technique to reach its full potential. A number of approaches including embedding the sample in water or spraying water above sample surface has shown great prospective for increasing the ionization yield by a factor of 10 to 100 through 'proton mediated' reaction. Based on this hypothesis, a water cluster primary ion source has been developed in collaboration with Ionoptika Ltd to generate giant water cluster ions (H 2 O) n + (n = 1À10 000) using a similar supersonic jet expansion methodology as for argon cluster beams. The ion yields of arginine, cholesterol, angiotensin II and a lipid mix have been measured under static and high ion dose conditions using (H 2 O) 5000 + , (H 2 O) 3000 + , Ar 3000 + and C 60 + primary ion beams at 20 keV. An enhancement in yields up to a factor of around 4 is observed under water cluster impact, in comparison with C 60 + at 1 × 10 11 ions/cm 2 ion dose, whereas this increases by around 10-50 times at high ion dose conditions.