In this work, we studied the upgrading of a mixed gas stream having a typical biogas composition by membrane gas separation with the aim of producing a purified biomethane stream. A polyetherimide/polyimide (PEI-PI) membrane module with an area of 1500 cm2 was used to evaluate the mass transport properties and the influence of water vapour and H2S on the separation performance. CO2 permeance was depleted by the presence of CH4 contrarily to this latter, whose permeation was enhanced by CO2. The permeances of both CO2 and CH4 were significantly affected by the presence of water vapour, showing a reduction of about 24–27% with respect to dry gas. Long-term analysis lasted about 650 days, showed an overall permeance variation of about 23% for N2 and 14% for CO2.The experimental permeance and selectivity values measured in wet conditions were used to simulate a multistep membrane system able to purify a “clean” biogas stream up to 98% of methane. It was found that the increasing number of steps from 3 to 7 allows a significant saving of membrane area, paired to an increment of CH4 recovery and CO2 purity.