Human Serum Albumin Monolayers on Mica: Electrokinetic Characteristics

Abstract

Adsorption of human serum albumin (HSA) on mica from 0.15 M NaCl solutions and various pH values was studied using in situ streaming potential measurements, AFM imaging, and XPS. The results obtained by the streaming potential were consistent with AFM measurements and theoretical predictions based on the random sequential adsorption model. This allowed one to determine both the kinetics of adsorption and the maximum coverage of HSA as a function of pH. At pH 3.5, the maximum coverage of HSA was 0.45 (which corresponds to 1.4 mg m(-2) neglecting hydration). This decreased monotonically with the increase in pH, attaining 0.30 (pH 5.1) and 0.25 (pH 7.4). At pH >10.5, the adsorption of HSA on mica was negligible. Further experimental studies performed for HSA monolayers of well-controlled coverage revealed their stability against pH cycling. It was found in these experiments that at pH <4 and >8 the electrokinetic properties of HSA monolayers approached the reference data pertinent to the bulk. However, for an intermediate pH range, deviations from the bulk reference data were observed, suggesting a dipolar (heterogeneous) charge distribution over adsorbed HSA molecules. This caused a slight shift in the isoelectric point of the monolayer determined to be 4.7 compared to the bulk value of 5.1. However, for the HSA coverage below 0.2, the effect of the substrate was significant, making the zeta potential more negative and shifting the apparent isoelectric point to more acidic values. It was suggested that these results obtained for planar and smooth interfaces could be used as reference data for interpreting albumin adsorption on colloid carrier particles.