Differences in the adsorption of human serum albumin (HSA) to hydrophobic self-assembled monolayers of hexadecyltrichlorosilane (HTS) with different layer packings have been observed using X-ray reflectivity. The monolayers with various thicknesses and electron density parameters were prepared using different deposition times. After the monolayers were exposed to 0.1 mg/mL solutions of HSA, a layer of albumin molecules was found adsorbed to the surface. When eluting with sodium dodecyl sulfate (SDS), all adsorbed protein was removed from the more densely packed surfaces. In contrast, a significant fraction of protein molecules adsorbed to less densely packed layers resisted elution with SDS. We conclude that more mobile, less densely packed I-ITS layers can better accommodate the albumin molecules, which results in more tenacious binding.