Abstract The objective of this work was to evaluate the performance of mesophilic continuous anaerobic co-digestion using oil-extracted microalgae (M) and glycerol (G) in co-digestion with chicken litter (CL). The process included the starting up and stabilization of continuous anaerobic bioreactors using CL as a feedstock and the corresponding adaptation to M-CL and M-G-CL feedstocks. The treatments were selected based on a previous report of our research team on Biochemical Methane Potential (BMP) evaluation, taking in consideration only the best M-G-CL feedstock ratios. The performance was evaluated by the Specific Methanogenic Activity (SMA); the best response (270.0 mL CH4 gVS added−1) was obtained with a triple co-digestion M-G-CL 30:3:67, which was 39.0% above the CL treatment. These findings have shown that the two main residuals from microalgae biodiesel production (G and M) can be used as a feedstock to improve methane production through anaerobic digestion.