Practical Guides for X-Ray Photoelectron Spectroscopy (XPS): First Steps in planning, conducting and reporting XPS measurements.

Over the past three decades, the widespread utility and applicability of X-ray photoelectron spectroscopy (XPS) in research and applications has made it the most popular and widely used method of surface analysis. Associated with this increased use has been an increase in the number of new or inexperienced users which has led to erroneous uses and misapplications of the method. This article is the first in a series of guides assembled by a committee of experienced XPS practitioners that are intended to assist inexperienced users by providing information about good practices in the use of XPS. This first guide outlines steps appropriate for determining whether XPS is capable of obtaining the desired information, identifies issues relevant to planning, conducting and reporting an XPS measurement, and identifies sources of practical information for conducting XPS measurements. Many of the topics and questions addressed in this article also apply to other surface-analysis techniques.

In Situ Electrochemical-AFM and Cluster-Ion-Profiled XPS Characterization of an Insulating Polymeric Membrane as a Substrate for Immobilizing Biomolecules.

The electrochemical oxidation of ortho-aminophenol (oAP) by cyclic voltammetry (CV), on platinum substrates in neutral solution, produces a polymeric film (PoAP) that grows to a limiting thickness of about 10 nm. The insulating film has potential use as a bioimmobilizing substrate, with its specificity depending on the orientation of its molecular chains. Prior investigations suggest that the film consists of alternating quinoneimine and oAP units, progressively filling all the platinum sites during the electrosynthesis. This work concerns the evaluation of the growth orientation of PoAP chains, which until now was deduced only from indirect evidence. Atomic force microscopy (AFM) has been used in situ with an electrochemical cell so that PoAP deposition on a specific area can be observed, thus avoiding any surface reorganization during ex situ transport. In parallel with microscopy, XPS experiments have been performed using cluster ion beams to profile this film, which is exceptionally thin, without damage while retaining molecular information.

Combined effects of solvation and aggregation propensity on the final supramolecular structures adopted by hydrophobic, glycine-rich, elastin-like polypeptides.

Previous work on elastin-like polypeptides (ELPs) made of hydrophobic amino acids of the type XxxGlyGlyZzzGly (Xxx, Zzz = Val, Leu) has consistently shown that differing dominant supramolecular structures were formed when the suspending media were varied: helical, amyloid-like fibers when suspended in water and globules evolving into "string of bead" structures, poly(ValGlyGlyValGly), or cigar-like bundles, poly(ValGlyGlyLeuGly), when suspended in methyl alcohol. Comparative experiments with poly(LeuGlyGlyValGly) have further indicated that the interface energy plays a significant role and that solvation effects act in concomitance with the intrinsic aggregation propensity of the repeat sequence. Continuing our investigation on ELPs using surface (X-ray photoelectron spectroscopy, atomic force microscopy) and bulk (circular dichroism, Fourier transform infrared spectroscopy) techniques for their characterization, here we have compared the effect of suspending solvents (H(2)O, dimethylsulfoxide, ethylene glycol, and MeOH) on poly(ValGlyGlyValGly), the polypeptide most inclined to form long and well-refined helical fibers in water, searching for the signature of intermolecular interactions occurring between the polypeptide chains in the given suspension. The influence of sequence specificities has been studied by comparing poly(ValGlyGlyValGly) and poly(LeuGlyGlyValGly) with a similar degree of polymerization. Deposits on substrates of the polypeptides were characterized taking into account the differing evaporation rate of solvents, and tests on their stability in ultra high vacuum were performed. Finally, combining experimental and computational studies, we have revaluated the three-dimensional modeling previously proposed for the supramolecular assembly in water of poly(ValGlyGlyValGly). The results were discussed and rationalized also in the light of published data.

Influence of amino acid specificities on the molecular and supramolecular organization of glycine-rich elastin-like polypeptides in water.

Elastin-like polypeptides adopt complex supramolecular structures, showing either a hydrophobic or a hydrophilic surface, depending on their surrounding environment and the supporting substrate. The preferred organization is important in many situations ranging from biocompatibility to bio-function. Here we compare the n-repeat pentamer LeuGlyGlyValGly (n = 7) with the analogue ValGlyGlyValGly (n = 5), as water suspensions and as deposits on silicon substrates. These sequences contain the repeat XxxGlyGlyZzzGly (Xxx, Zzz = Val, Leu) motif belonging to the hydrophobic glycine-rich domain of elastin and represent a simplified model from which to obtain information on molecular interactions functional to elastin itself. The compounds studied differ only by the presence of the -CH(2)- spacer in the Leu moiety and thus the work was aimed at revealing the influence of this spacer element on self assembly. Both polypeptides were studied under identical conditions, using combined techniques, to identify differences in their conformational states both at molecular (CD, FTIR) and supramolecular (XPS, AFM) levels. By these means, together with a Congo Red spectroscopic assay of ß-sheet formation in water, a clear correlation between amino acid sequences (sequence specificity) and their kinetics and ordering of aggregation has emerged. The novel outcomes of this work are from the supplementary measurements, made to augment the AFM and XPS studies, showing that the significant step in the self assembly of both polypeptides takes place in the liquid phase and from the finding that the substitution of Val by Leu in the first position of the pentapeptide effectively inhibits the formation of amyloidal fibers.

Conformational study and hydrogen bonds detection on elastin-related polypeptides using X-ray photoelectron spectroscopy.

The chemical bonds of the pentapeptide sequence of elastin ValGlyGlyValGly (VGGVG), both in its monomer and polymer forms, were correlated with their XPS spectra through a well-established curve-fitting procedure. To aid in this correlation, the C1s, O1s, and N1s chemical shifts of the Boc-VGGVG-OEt, were validated by theoretical calculations, performed in the framework of the Koopman approximation of HF/6-31G molecular orbitals, leading to the "preferred" conformation of the protected monomer. Then the same curve-fitting procedure was adopted for interpreting the XPS spectra of the polypentapeptide as a powder, and the XPS results obtained both for monomer and polymer compounds were compared with those obtained by FT-IR. The polymer was then analyzed after deposition onto a silicon substrate, Si(100), either from methanol or water suspensions and the presence of hydrogen bonds was detected at the polymer/substrate interface and between the polymer chains. The "surface rearrangement" that could be inferred from XPS results strongly confirms that derived from AFM images previously obtained under the same experimental conditions. In particular, the observed amyloid conformation is stabilized by hydrogen bonds to water molecules included in the structure while the formation of the beaded string structure observed in deposits from methanolic suspension is probably mediated by hydrogen bonds to the hydrated silicon surface.