Macromolecular Mass Transport to a Surface: Effects of Shear Rate, pH, and Ionic Strength

Abstract

Macromolecular mass transport from a homogeneous solution to a glass planar surface was investigated under diffusion-limited conditions in a rectangular flow cell. A total internal reflection fluorescence (TIRF) method was used to determine the diffusivity of bovine serum albumin (BSA) based on the measurement of its adsorption rate to a glass surface under diffusion-limited conditions. Adsorption of BSA, which was covalently bound to fluorescein isothiocyanate (FITC), was measured over the shear rate range of 200 to 1000 s−1using TIRF. Adsorbed concentrations of tritium-labeled BSA–FITC conjugate on the glass surface were compared to the measured surface fluorescence intensity to determine the calibration constant under various conditions. Adsorption was determined as a function of shear rate, pH, ionic strength, and buffer system. All adsorption experiments were run at 25 ± 1°C. Results in this study showed that the ratio of surface fluorescence intensity to adsorbed surface concentration of BSA was nearly constant and not a function of the various conditions investigated. The diffusivity of BSA measured showed no shear rate dependence and appeared to be a Brownian diffusivity. The range of diffusivity values was 3.0 × 10−7to 7.1 × 10−7cm2/s, with the highest diffusivity value occurring at the isoelectric point. The value of the BSA diffusivity calculated using the Stokes–Einstein equation was 3.1 × 10−7cm2/s.