Experimental Methodology for Analysing Macromolecular Interactions in the Context of Marine Bacterial Adhesion to Stainless Steel

Abstract

In the frame of a long-term study of biofilm formation in the marine environment, protein-coated amine latex microbeads were used as a simplified bacterial model, in order to elucidate the role of surface protein on adhesion to 316 L stainless steel. Bovine serum albumin (BSA), chosen as the model protein, was covalently immobilized to the amine latex beads using glutaraldehyde as cross-linker. The adhesion of BSA-coated latex beads to stainless steel was then examined under a well-controlled physico-chemical environment, using a shear stress flow chamber. The shear-induced detachment of beads was analysed by testing three different microbeads/stainless steel systems: bare latex beads, activated latex beads with poly(glutaral-dehyde) and BSA-coated latex beads in adhesive contact with stainless steel. The suspending medium used for detachment was representative of seawater in terms of pH and ionic strength. Adhesion was quantified through two complementary parameters: the wall shear stress threshold needed to cause 2% bead detachment ( τ W2%) and the wall shear stress required to remove 50% of the initially adhered beads ( τ W50%). Zeta potential measurements, paired with protein quantification, indicated that the coating density was weak. Nevertheless, the BSA-coated latex beads promoted an increased adhesion compared to bare beads, which was demonstrated by higher τ W2% and τ W50% values ( τ W2% = 15.9 ± 8.1 Pa and 39.4 ± 10.7 Pa; τ W50% = 41.4 ± 7.9 and 73.5 ± 14.8 Pa for bare and BSA-coated beads, respectively). These results confirmed the role of surface proteins on adhesion to stainless steel. The shear stress flow chamber and the associated method for bead coating were demonstrated to be a relevant approach to analyse nano-scale molecular interactions between biological and physical surfaces.