NEUTRON AND X-RAY REFLECTIVITY STUDIES OF HUMAN SERUM-ALBUMIN ADSORPTION ONTO FUNCTIONALIZED SURFACES OF SELF-ASSEMBLED MONOLAYERS

Neutron and X-ray reflectivity (NR and XR) have been widely used for t he investigation of the structure of thin organic films. Here we demon strate how these sensitive techniques may be applied to the study of p rotein adsorption to well-characterized self-assembled monolayers (SAM s) with different chemical functionalities. NR can be used for in situ study, while XR provides complementary information on the initial sur faces and dried layers measured in air after protein has been adsorbed . In situ measurements of adsorption of human serum albumin onto a hyd rophilic NH2-terminated monolayer clearly show the presence of a thin layer of adsorbed protein next to the SAM. Adsorption of albumin onto a hydrophobic, deuterated, CD3-terminated SAM causes even bigger chang es in the NR. Upon replacing the protein solution with protein-free bu ffer solution, the reflectivities from both kinds of monolayers do not change, demonstrating that the albumin adsorption is irreversible aft er several hours of contact with the protein solution. X-ray reflectiv ity measurements of dried substrates performed ex situ in air provide a lower bound estimate of the amount of protein which must be at the i nterface in. situ. This combination of techniques provides a uniquely sensitive approach for studying changes that occur upon protein adsorp tion at an interface.