Entrapment of methane-producing microorganisms between semi-permeable synthetic membranes in a multi-layer membrane bioreactor (MMBR) was studied and compared to the digestion capacity of a free-cell digester, using a hydraulic retention time of one day and organic loading rates (OLR) of 3.08, 6.16, and 8.16 g COD/L day. The reactor was designed to retain bacterial cells with uprising plug flow through a narrow tunnel between membrane layers, in order to acquire maximal mass transfer in a compact bioreactor. Membranes of hydrophobic polyamide 46 (PA) and hydroxyethylated polyamide 46 (HPA) as well as a commercial membrane of polyvinylidene fluoride (PVDF) were examined. While the bacteria in the free-cell digester were washed out, the membrane bioreactor succeeded in retaining them. Cross-flow of the liquid through the membrane surface and diffusion of the substrate through the membranes, using no extra driving force, allowed the bacteria to receive nutrients and to produce biogas. However, the choice of membrane type was crucial. Synthesized hydrophobic PA membrane was not effective for this purpose, producing 50-121 mL biogas/day, while developed HPA membrane and the reference PVDF were able to transfer the nutrients and metabolites while retaining the cells, producing 1102-1633 and 1016-1960 mL biogas/day, respectively.