| Abstract
| - In complex adsorbed protein films, the biological reactivity of the adsorbed proteins depends on theirrelative concentrations, organization, conformation, and orientation. Time-of-flight secondary ion massspectrometry (ToF-SIMS) samples the outermost composition of an adsorbed protein film, providingbiologically relevant information about the organization and availability of the adsorbed proteins in thefilm. We have previously shown that ToF-SIMS can quantitatively measure the composition of binaryadsorbed protein films. However, for one particular binary system studied (immunoglobulin G−fibrinogen),a difference between the ToF-SIMS and 125I-radiolabeled protein adsorption measurements was reported.It was hypothesized that the composition of the protein film at its outermost (and, therefore, most biologicallyrelevant) surface was different than that of the overall protein film due to the protrusion of the adsorbedfibrinogen over the adsorbed immunoglobulin G. This study provides further evidence supporting thishypothesis by incorporating electron spectroscopy for chemical analysis (ESCA) to examine the overallthickness of the adsorbed protein layers. Binary mixtures of three important plasma proteins (albumin,fibrinogen, and immunoglobulin G) and a ternary mixture of all three proteins were studied using radiolabeledprotein adsorption measurements, ESCA, and ToF-SIMS. The different types of information generatedusing these techniques are shown to be complementary in describing the structure (i.e., organization,composition, conformation, and orientation) of the adsorbed protein films. Models of the proteins in thebinary and ternary films are proposed based on the experimental data that reflect the apparent preferentiallocation of fibrinogen at the surface for mixtures of fibrinogen and immunoglobulin G.
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