Atomic force microscopy (AFM) and X-ray reflectivity (XR) have been used together to provide a detailed and direct look at the structure of human serum albumin protein adsorbed onto well-characterized selfassembled monolayer (SAM) surfaces at several protein concentrations. The duration of SAM deposition was also varied to investigate the influence of the density of hydrocarbon chains in the SAM on protein binding tenacity. Concurrent study of adsorption to bare silicon wafers with native oxide surfaces provided a comparison with a hydrophilic surface similar to widely studied glass and quartz surfaces. Both AFM and XR measurements showed that after adsorption, rinsing, and drying, the surfaces of all substrates were covered with no more than a single layer of adsorbed protein. Thin dense protein layers were seen for the substrates exposed to protein concentrations of 0.1 and 0.5 mg/mL. Partial surface coverage by protein aggregates having larger thicknesses was seen for substrates exposed to lower concentrations. The tenacity of the protein adsorption on different substrates was tested by eluting the adsorbed protein with a 1% solution of sodium dodecyl sulfate surfactant. This treatment removed almost all protein from the bare silicon surface and from the fully formed, dense SAMs. A significant amount of adsorbed protein remained on the surface of the less dense, "incomplete" monolayers, suggesting that protein adsorbed more tenaciously on that surface.

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We report the synthesis of novel hyperbranched amphiphilic poly(ethylene oxide)−polystyrene (PEO−PS) n copolymers obtained by controlled radical polymerizations:  nitroxide mediated polymerization (NMP) and reversible addition−fragmentation chain transfer (RAFT). The macroinimers used to synthesize copolymers have the general structure AB*, where A stands for PEO with a terminal double bond and B* stands for a PS block with a terminal initiating group (TEMPO or RAFT CTA). Bulk NMP yielded copolymers with higher molecular weight and higher polydispersity. RAFT polymerization in solution gave hyperbranched copolymers with higher molecular weight but lower polydispersity. Langmuir monolayers displayed reversible amphiphilic behavior at the air−water interface. The random, mixed character of short hydrophilic and hydrophobic fragments results in peculiar surface behavior:  unlike regular linear and star block copolymers, the amphiphilic hyperbranched macromolecules with higher PEO content are spread at the air−water interface and short PEO fragments are not submerged into the water subphase even at high compression.

The role that alkyl chain packing density in a self-assembled monolayer (SAM) plays in the adsorption of protein to the SAM surface was investigated using in situ neutron reflectivity (NR) and total internal reflection fluorescence (TIRF) measurements of the adsorption behavior of human serum albumin (HSA) and human immunoglobulin G (HGG). The proteins differ particularly in the fact that HSA has specific binding pockets for alkyl chains while HGG does not. NR results show that HSA adsorbs from a 1.0 mg/mL solution as a single layer on the SAMs, with the protein interpenetrating into a less densely packed SAM, but not into a more densely packed SAM. Likewise, the kinetics of the HSA adsorption to the SAMs varies markedly with the alkyl chain packing. In contrast, both the structure of the adsorbed layer and adsorption kinetics vary little with the alkyl chain packing density in the case of HGG adsorbing from solution. HGG also does not penetrate into loosely packed SAMs. NR results reveal that the HGG adsorbs as two layers, with the layer closest to the SAM being more tightly packed. When HSA and HGG compete for adsorption sites on a SAM, HGG effectively displaces HSA from a tightly packed SAM, but does not compete effectively with HSA adsorbed tenaciously onto a loosely packed SAM.