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Aizome (Japanese indigo dyeing) is a unique dyeing method using microbial activity under anaerobic alkaline conditions. In indigo-dye fermenting suspensions, microorganisms reduce indigo into leuco-indigo with acetaldehyde as a reductant. In this study, we constructed a semi-microbial biofuel cell using an indigo-dye fermenting suspension. Carbon fiber and Pt mesh were used as the anode and cathode materials, respectively. The open-circuit voltage (OCV) was 0.6 V, and the maximum output power was 32 µW cm−2 (320 mW m−2). In addition, the continuous stability was evaluated under given conditions starting with the highest power density, the power density rapidly decreased in 0.5 h due to the degradation of the anode. Conversely, at the OCV, the anode potential exhibited high stability for two days. However, the OCV decreased by approximately 80 mV after 2 d compared with the initial value, which was attributed to the performance degradation of the gas-diffusion-cathode system caused by the evaporation of the dispersion solution. This is the first study to construct a semi-microbial biofuel cell using an indigo-dye fermenting suspension.