The application of immobilized-cell technology in photobioreactor for hydrogen production could offer improvements in photo-hydrogen production rate and light utilization efficiency. Indigenous Rhodopseudomonas palustris CQK 01 was attached to the surface of a cover glass slide in a flat-panel photobioreactor, to form biofilm under illumination with a range of intensities and wavelengths. The morphology and structure of mature photosynthetic bacterial (PSB) biofilm were determined to elucidate the relationship between biofilm formation and hydrogen production performance. The effects of operation conditions on hydrogen production performance of the biofilms formed under various illumination conditions were experimentally investigated. The results showed that illumination wavelength and intensity substantially influenced the morphology and structure of the biofilm, and the hydrogen production performance of mature biofilm varied significantly with the illumination conditions that were used for biofilm formation. Biofilm formed under 590 nm and 5000 lx illumination showed the highest hydrogen production performance. (C) 2010 Elsevier Ltd. All rights reserved.