CORRELATION BETWEEN SURFACE-MORPHOLOGY AND SURFACE FORCES OF PROTEIN-A ADSORBED ON MICA

We have investigated the morphology and surface forces of protein A ad sorbed on mica surface in the protein solutions of various concentrati ons. The force-distance curves, measured with a surface force apparatu s (SFA), were interpreted in terms of two different regimens: a ''larg e-distance'' regimen in which an electrostatic double-layer force domi nates, and an ''adsorbed layer'' regimen in which a force of steric or igin dominates. To further clarify the forces of steric origin, the su rface morphology of the adsorbed protein layer was investigated with a n atomic force microscope (AFM) because the steric repulsive forces ar e strongly affected by the adsorption mode of protein A molecules on m ica. At lower protein concentrations (2 ppm, 10 ppm), protein A molecu les were adsorbed ''side-on'' parallel to the mica surfaces, forming a monolayer of similar to 2.5 nm. AFM images at higher concentrations ( 30 ppm, 100 ppm) showed protruding structures over the monolayer, whic h revealed that the adsorbed protein A molecules had one end oriented into the solution, with the remainder of each molecule adsorbed side-o n to the mica surface. These extending ends of protein A overlapped ea ch other and formed a ''quasi-double layer'' over the mica surface. Th ese AFM images proved the existence of a monolayer of protein A molecu les at low concentrations and a ''quasi-double layer'' with occasional protrusions at high concentrations, which were consistent with the ad sorption mode observed in the force-distance curves.