A representative in vitro biofilm model. A) Photograph showing a methacrylate stent with 10 mm wide and 7 mm high and an internal drilling with a diameter of 2.7 and 5 mm deep to support the implants in a fixed position allowing the exposure of the coronal third of the implant surface. B) CLSM Images obtained at 12 (I), 24 (II), 48 (III), 72 (IV), 96 (V), and 120 (VI) h of incubation of biofilms over whole dental implants which were stained using LIVE/DEAD BacLight Kit with live bacteria in green, dead bacteria in red, and implant surface in blue. C) SEM images showed biofilm growth from 48 to 120 h over whole dental implants. I): Biofilms after 48 h of incubation, with a complex morphology, in which Fusobacterium nucleatum formed networks with the adhered microcolonies of bacteria. II,III): Biofilms after 72 and 96 h of incubation, indicating the bacteria were in the expected larger stacks (growing masses of bacterial cells) and presence of broad channels (green arrow) and the cell mass and ECM surrounding bacteria in the biofilm (white arrows). IV): The biofilms after incubation from 72 to 120 h did not change in architecture. Reproduced with permission.[
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