As major structural EPS of the cystic fibrosis biofilm produced by mucoid Pseudomonas aeruginosa, alginate and its role in the tenacity of lung infection is a hot research topic.[1] It has been shown and proven that alginate plays a major role in shielding of antibiotics, such as aminoglycosides like Gentamycin [2] or Tobramycin (TS) [3]. However, to investigate in vitro the interaction between this EPS and drugs, the most reported models rely on either direct isolation of bacterial alginate or use native alginate derived from seaweed. As it is known that P. aeruginosa produces acetylated alginate, bacterial alginate seems more appropriate to use, but it comes with the drawback of tedious isolation processes and the need of extensive bacterial culturing [4]. One the other hand, native seaweed alginate, despite being commercially available, lacks acetylation and, hence, does not mimic CF-like biofilm environment. In this work we aim to investigate the influence of acetylation on the interaction between alginate and the aminoglycoside TS. Therefore, seaweed derived alginate was modified according to literature [5] and its interaction with TS was investigated at different scales. Bio-printed alginate microbeads were used to investigate macroscopic interaction, whereas interactions on a molecular level were assessed using isothermal titration calorimetry (ITC). Our results showed, for the first time, that the intensity of the interaction between alginate and TS highly depends on its degree of acetylation. Indeed, it was found, that acetylation strongly influences the shrinking behaviour of alginate beads once incubated in 0.1 wt% TS solution. We discovered also that not only the relative size after incubation but also the speed at which it shrinks were strongly affected by acetylation, as shown by Figure 1. First preliminary results in swelling and ITC experiments have proven that acetylation plays a key role in “shielding” drugs, like TS.