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1 2011

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Detection of fibrinogen monolayers on mica by the colloid enhancement

Nattich-Rak M., Adamczyk Z., Wasilewska M., Radziszewska M.

Environmental Biotechnology

2011

Vol. 7, No. 1

1-11

Physicochemical properties of bovine plasma fibrinogen (Fb) in electrolyte solutions were characterised, comprising the diffusion coefficient (hydrodynamic radius), electrophoretic mobility (zeta potential) and the isoelectric point, found to be at pH=5.8. Similar electrokinetic measurements were performed for the mica substrate using the streaming potential cell. The kinetics of Fb adsorption on mica under diffusion-controlled transport was also studied. The surface concentration of Fb on mica was determined directly by AFM counting. By adjusting the time of adsorption, and bulk Fb concentration, monolayers of desired coverage were produced. It was confirmed that Fb adsorbed irreversibly on mica both at pH=3.5 and pH=7.4 (physiological value). It was postulated that in the latter case, where both the substrate and fibrinogen molecules were negatively charged, adsorption was due to heterogeneous charge distribution over the protein molecule. In order to check this hypothesis, monolayers of Fb on mica were studied using the colloid enhancement (CE) method, in which negatively and positively charged latex particles were used. Results of these experiments were quantitatively interpreted in terms of the fluctuation theory assuming that adsorption sites consisted of two and three Fb molecules, for pH=3.5 and 7.4, respectively. This allowed one to determine limits of applicability of the classical DLVO theory and confirm a heterogeneous charge distribution over the Fb molecule. It was also concluded that the CE method can be used for a sensitive determination of the Fb bulk concentration for the range inaccessible for other methods, i.e., for 0.1ppm and below. Another effect of vital significance confirmed in this work was that for some range of fibrinogen coverage both the negative and positive latexes efficiently adsorbed. This indicates the formation of superadsorbing surfaces having potential significance for various filtration processes.