Polyelectrolyte multilayers (PEMs) of poly-l-lysine (PLL) and alginic acid sodium salt (Alg) are fabricated applying the layer by layer technique and annealed at a constant temperature; 37, 50 and 80 °C, for 72 h. Atomic force microscopy reveals changes in the topography of the PEM, which is changing from a fibrillar to a smooth surface. Advancing contact angle in water varies from 36° before annealing to 93°, 77° and 95° after annealing at 37, 50 and 80 °C, respectively. Surface energy changes after annealing were calculated from contact angle measurements performed with organic solvents. Quartz crystal microbalance with dissipation, contact angle and fluorescence spectroscopy measurements show a significant decrease in the adsorption of the bovine serum albumin protein to the PEMs after annealing. Changes in the physical properties of the PEMs are interpreted as a result of the reorganization of the polyelectrolytes in the PEMs from a layered structure into complexes where the interaction of polycations and polyanions is enhanced. This work proposes a simple method to endow bio-PEMs with antifouling characteristics and tune their wettability.