Non-invasive analysis and a final destructive analysis were employed to study the fouling formation during the initial days of AS-MBR operation. The fouling layer development was quantified in-situ non-invasively with Optical Coherence Tomography (OCT). The increase in biomass thickness was related to the transmembrane pressure (TMP) and to the increase in concentration of soluble microbial products (SMP) in the reactor The OCT non-destructive analysis allowed normalizing the final autopsy values for the amount of biomass deposited on the membrane. After 8 days of operation, the cake layer presented a biomass activity of 400 pg/mm3 of intra-ATP and EPS concentration of 9.8 mg/ mm3. The microbial community analysis of sludge and biofouling on the membrane surface revealed the abundance of Proteobacteria.