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Preventing bacterial attachment to surfaces is the most efficient approach to controlling biofilm proliferation. The aim of this study was to compare anti-adhesion potentials of 5 and 50 mmol/L polyelectrolyte multilayers of poly(allylamine hydrochloride)/poly(sodium 4–styrenesulfonate), poly(4-vinyl-N-ethylpyridinium bromide)/ poly(sodium 4–styrenesulfonate), and poly(4-vinyl-N-isobutylpyridinium bromide)/poly(sodium 4–styrenesulfonate) against Escherichia coli. Glass surface was covered with five polyelectrolyte layers and exposed to bacterial suspensions. Poly(4-vinyl-N-ethylpyridinium bromide)/poly(sodium 4–styrenesulfonate) was the most effective against bacterial adhesion, having reduced it by 60 %, followed by poly(4-vinyl-N-isobutylpyridinium bromide)/poly(sodium 4– styrenesulfonate) (47 %), and poly(allylamine hydrochloride)/poly(sodium 4–styrenesulfonate) (38 %). Polyelectrolyte multilayers with quaternary amine groups have a significant anti-adhesion potential and could find their place in coatings for food, pharmaceutical, and medical industry.