Silicone coatings with enhanced antifouling behavior towards bacteria, diatoms, and a diatom dominated slime were prepared by incorporating PEO-silane amphiphiles with varied siloxane tether lengths (a–c): α-(EtO)₃Si(CH₂)₂-oligodimethylsiloxane_n-block-poly(ethylene oxide)₈-OCH₃ [n = 0 (a), 4 (b), and 13 (c)]. Three modified silicone coatings (A–C) were prepared by the acid-catalyzed sol–gel cross-linking of a–c, respectively, each with a stoichiometric 2:3 M ratio of α, ω-bis(Si–OH)polydimethylsiloxane (M_n = 3,000 g mol⁻¹). The coatings were exposed to the marine bacterium Bacillus sp.416 and the diatom (microalga) Cylindrotheca closterium, as well as a mixed community of Bacillus sp. and C. closterium. In addition, in situ microfouling was assessed by maintaining the coatings in the Atlantic Ocean. Under all test conditions, biofouling was reduced to the highest extent on coating C which was prepared with the PEO-silane amphiphile having the longest siloxane tether length (c).