Fibrinogen Adsorption on Three Silica-Based Surfaces:  Conformation and Kinetics

Abstract

Using AFM (atomic force microscopy) to probe protein conformation and arrangement, and TIRF (total internal reflectance fluorescence) to monitor kinetics, fibrinogen adsorption on three different silica-based surfaces was studied: the native oxide on silicon, acid-etched microscope slides, and acid-etched polished glass. The three are chemically similar, but the microscope slide is rougher and induces AFM tip instabilities that appear as high spots on the bare surface. Fibrinogen's conformation and transport-limited adsorption kinetics are found to be quantitatively similar on all three surfaces. Further, the number of adsorbed proteins in progressive AFM micrographs quantitatively match the coverages measured by TIRF during early adsorption. Surfaces appear full, via AFM, when adsorbed amounts are about an order of magnitude below their true saturation levels (via TIRF) because, above about 0.26 mg/m(2), individual proteins cannot be discerned. The results demonstrate how the appearance of AFM micrographs can be misleading regarding surface saturation. On all three surfaces, fibrinogen is, at most, slightly aggregated, showing limited, if any, surface mobility. The complexities of the microscope slide's surface landscape minimally impact adsorption.

Toward protein imprinting with polymer brushes

“…19 Specifically, AFM topography images of the surface templated with the fibrinogen molecules are presented. The footprints geometrically corresponding to fibrinogen size 20 and shape can be clearly observed ͑Fig. 5, dotted ovals͒.…”

Total internal reflection spectroscopy for studying soft matter

“…Inorganic ions, such as phosphate and sulphate, have characteristic vibrations that can be used to study adsorption kinetics. 120 TIRF has also been used to measure the kinetics of adsorption and desorption, primarily of proteins [122][123][124][125] but also of polymers. 126,127 In all cases the target molecule was fluorescently labelled.…”

Cell-Protein-Material Interaction in Tissue Engineering

“…a technique able to provide direct observation of protein conformation on different substrates has been extensively used in recent years, mainly on model surfaces, such as silica, mica, titanium graphite, and self-assembled monolayers (SAMs), flat enough so that the height magnitude is able to reveal the trinodular structure of single-adsorbed FG molecules (Toscano & Santore, 2006;Marchin & Berrie, 2003;Agnihotri & Siedlecki, 2004;Cacciafesta et al, 2000;Tunc et al, 2005;Gettens et al, 2005;Gettens & Gilbert, 2007;Ta & McDermott, 2000;Ishizaki et al, 2007;Ortega-Vinuesa et al, 1998;Mitsakakis et al, 2007;Sit & Marchant, 1999). The effect of surface wettability, as one of the most important parameters that affects the biological response to a material, on FG adsorption has lead to different, nonconsistent conclusions.…”