Protein adsorption is a delicate process, which results from the balance between the properties of proteins and their solid supports. Although the relevance of some of these parameters has been already unveiled, the precise involvement of electrostatics and other weaker intermolecular forces requires further comprehension. Aiming to contribute to this task, this work explores the attachment, rearrangement, and surface aggregation of a model spheroprotein, such as bovine β-lactoglobulin (β-LG), onto hydrophilic substrates prefunctionalized with different alkylthiol films. Thereby, a variety of electrostatic scenarios for the adsorption of β-LG could be recreated through the variation of the pH and the functional chemistry of the surfaces. The changes in surface mass density (plus associated water) and film flexibility were followed in situ with quartz crystal microbalance with dissipation monitoring. Film packing and aggregation were assessed by faradaic electrochemical measurements and ex situ atomic force microscopy and field effect scanning electron microscopy. In contrast to previous hypotheses arguing that electrostatic interactions between charged substrates and proteins would be the only driving force, a complex interplay between Coulombic and non-Coulombic intermolecular forces (which would depend upon the experimental conditions) has been suggested to explain the results.