Sodium dodecyl sulfate adsorption onto positively charged surfaces: monolayer formation with opposing headgroup orientations.

The adsorption and structure of sodium dodecyl sulfate (SDS) layers onto positively charged films have been monitored in situ with vibrational sum-frequency-generation (SFG) spectroscopy and surface plasmon resonance (SPR) sensing. Substrates with different charge densities and polarities used in these studies include CaF2 at different pH values as well as allylamine and heptylamine films deposited onto CaF2 and Au substrates by radio frequency glow discharge deposition. The SDS films were adsorbed from aqueous solutions ranging in concentration from 0.067 to 20 mM. In general the SFG spectra exhibited well resolved CH and OH peaks. However, at SDS concentrations between 1 and 8 mM the SFG CH and OH intensities decreased close to background levels. Combined data sets from molecular conformation, orientation, and order sensitive SFG with mass sensitive SPR suggest that the observed changes in SFG intensities above 0.2 mM are related to structural arrangements in the SDS layer. A model is proposed where the SFG intensity minimum between 1 and 8 mM is associated with a monolayer containing two headgroup orientations, one pointing toward the substrate and one pointing toward the solution phase. The SFG peaks observed at concentrations below 0.2 mM are dominated by the presence of adsorbed contaminants such as fatty alcohols (e.g., dodecanol), which are more surface active than SDS. As SDS solution concentration is increased above 1 mM SDS molecules are incorporated in the surface layer, with dodecanol continuing to be present in the surface layer for solution concentrations up to at least the critical micelle concentration.

ToF-SIMS Analysis of Adsorbed Proteins: Principal Component Analysis of the Primary Ion Species Effect on the Protein Fragmentation Patterns.

In time-of-flight secondary ion mass spectrometry (ToF-SIMS), the choice of primary ion used for analysis can influence the resulting mass spectrum. This is because different primary ion types can produce different fragmentation pathways. In this study, analysis of single-component protein monolayers were performed using monatomic, tri-atomic, and polyatomic primary ion sources. Eight primary ions (Cs(+), Au(+), Au(3) (+), Bi(+), Bi(3) (+), Bi(3) (++), C(60) (+)) were used to examine to the low mass (m/z < 200) fragmentation patterns from five different proteins (bovine serum albumin, bovine serum fibrinogen, bovine immunoglobulin G and chicken egg white lysozyme) adsorbed onto mica surfaces. Principal component analysis (PCA) processing of the ToF-SIMS data showed that variation in peak intensity caused by the primary ions was greater than differences in protein composition. The spectra generated by Cs(+), Au(+) and Bi(+) primary ions were similar, but the spectra generated by monatomic, tri-atomic and polyatomic primary ion ions varied significantly. C(60) primary ions increased fragmentation of the adsorbed proteins in the m/z < 200 region, resulting in more intense low m/z peaks. Thus, comparison of data obtained by one primary ion species with that obtained by another primary ion species should be done with caution. However, for the spectra generated using a given primary ion beam, discrimination between the spectra of different proteins followed similar trends. Therefore, a PCA model of proteins created with a given ion source should only be applied to datasets obtained using the same ion source. The type of information obtained from PCA depended on the peak set used. When only amino acid peaks were used, PCA was able to identify the relationship between proteins by their amino acid composition. When all peaks from m/z 12-200 were used, PCA separated proteins based on a ratio of C(4)H(8)N(+) to K(+) peak intensities. This ratio correlated with the thickness of the protein films and Bi(1) (+) primary ions produced the most surface sensitive spectra.

Investigating activated sludge flocs using microanalytical techniques: demonstration of environmental scanning electron microscopy and time-of-flight secondary ion mass spectrometry for wastewater applications.

Environmental scanning electron microscopy (ESEM) with an energy-dispersive X-ray spectrometer (EDS) and time-of-flight secondary ion mass spectrometry (ToF-SIMS) were demonstrated to be useful analytical tools for investigating surface and bulk components of individual floc particles from both full- and bench-scale activated sludge systems. Detailed surface imaging of various hydrated biological floc particles by ESEM revealed substantial differences in surface features between treatment systems, while EDS identified spatial differences in the iron and the aluminum distributions. The ToF-SIMS spectra had signature fragments of protein and polysaccharide material from the floc surface, suggesting that this technique is capable of surface profiling extracellular polymeric substances. Principal-component analysis of the positive ion ToF-SIMS spectra from the mixed-liquor-suspended solid (MLSS) samples and reference aquatic organic materials found slight differences between the full- and bench-scale MLSS surface properties but substantial differences among MLSS and treated effluent from the same facility.

Surface chemical and mechanical properties of plasma-polymerized N-isopropylacrylamide.

Surface-immobilized poly(N-isopropyl acrylamide) (pNIPAM) is currently used for a wide variety of biosensor and biomaterial applications. A thorough characterization of the surface properties of pNIPAM thin films will benefit those applications. In this work, we present analysis of a plasma-polymerized NIPAM (ppNIPAM) coating by multiple surface analytical techniques, including time-of-flight secondary-ion mass spectrometry (ToF-SIMS), contact angle measurement, atomic force microscopy (AFM), and sum frequency generation (SFG) vibrational spectroscopy. ToF-SIMS data show that the plasma-deposited NIPAM polymer on the substrate is cross-linked with a good retention of the monomer integrity. Contact angle results confirm the thermoresponsive nature of the film as observed by a change of surface wettability as a function of temperature. Topographic and force-distance curve measurements by AFM further demonstrate that the grafted film shrinks or swells depending on the temperature of the aqueous environment. A clear transition of the elastic modulus is observed at 31-32 degrees C. The change of the surface wettability and mechanical properties vs temperature are attributed to different conformations taken by the polymer, which is reflected on the outmost surface as distinct side chain groups orienting outward at different temperatures as measured by SFG. The results suggest that a ppNIPAM thin film on a substrate experiences similar mechanical and chemical changes to pNIPAM bulk polymers in solution. The SFG result provides evidence supporting the current theory of the lower critical solution temperature (LCST) behavior of pNIPAM.

PEO-like plasma polymerized tetraglyme surface interactions with leukocytes and proteins: in vitro and in vivo studies.

Polyethylene oxide (PEO) surfaces reduce non-specific protein and cell interactions with implanted biomaterials and may improve their biocompatibility. PEO-like polymerized tetraglyme surfaces were made by glow discharge plasma deposition onto fluorinated ethylene propylene copolymer (FEP) substrates and were shown to adsorb less than 10 ng/cm2 of fibrinogen in vitro. The ability of the polymerized tetraglyme surfaces to resist leukocyte adhesion was studied in vitro and in vivo. Polymerized tetraglyme and FEP were implanted subcutaneously in mice and removed after 1 day or 4 weeks. Histological analysis showed a similar degree of fibrous encapsulation around all of the 4-week implants. Darkly stained wells were present in the fibrous tissues at the tissue-material interface of both FEP and tetraglyme. Scanning electron micrographs showed that in vivo macrophage adhesion to polymerized tetraglyme was much higher than to FEP. After 2-hour contact with heparinized whole blood, polymorphonuclear leukocyte (PMN) adhesion to polymerized tetraglyme was much higher than to FEP, while platelet adhesion to polymerized tetraglyme was lower than to FEP. When PMNs isolated from blood were suspended in 10% autologous plasma, cell adhesion to polymerized tetraglyme was higher than to FEP; however when the cells were suspended in heat inactivated serum, cell adhesion to FEP was higher than to polymerized tetraglyme. The surface chemistry of polymerized tetraglyme did not change after 2-hour blood contact, but displayed nitrogen functional groups after 1-day implantation and became slightly degraded after 4-week implantation. The surface chemistry of FEP did not change significantly after blood contact or implantation. Loosely bound proteins such as fibrinogen on polymerized tetraglyme may contribute to the adhesion of PMNs and macrophages and ultimately to fibrous encapsulation (the foreign body response) around the implants.

Limits of detection for time of flight secondary ion mass spectrometry (ToF-SIMS) and X-ray photoelectron spectroscopy (XPS): detection of low amounts of adsorbed protein.

Characterization of biomaterial surfaces requires analytical techniques that are capable of detecting a wide concentration range of adsorbed protein. This range includes detection of low amounts of adsorbed protein (<10 ng/cm2) that may be present on non-fouling biomaterials. X-ray Photoelectron Spectroscopy (XPS) and Time of Flight Secondary Ion Mass Spectrometry (ToF-SIMS) are surface sensitive techniques capable of detecting adsorbed proteins. We have investigated the lower limits of detection of both XPS and ToF-SIMS on four model substrates each presenting unique challenges for analysis by XPS and ToF-SIMS: mica, poly(tetrafluoroethylene), allyl amine plasma polymer and heptyl amine plasma polymer. The detection limit for XPS ranged from 10 ng/cm2 of fibrinogen (on mica) to 200 ng/cm2 (on allyl amine plasma polymers). The detection limit for ToF-SIMS ranged from 0.1 ng/cm2 of fibrinogen to 100 ng/cm2, depending on the substrate and data analysis. Optimal conditions provided detection limits between 0.1 ng/cm2 and 15 ng/cm2 on all of the substrates used in this study. While both techniques were shown to be effective in detecting protein, the sensitivity of both XPS and ToF-SIMS was shown to be dependent on substrate surface chemistry and the organization of the adsorbed protein film. This study specifically highlights the applicability of ToF-SIMS in the characterization of low level protein adsorption.