A two-phase protocol for ambient hydrogen production using Chlamydomonas reinhardtii

H₂ production from green-microalgae, for energy purposes, is the ultimate goal of large-scale production. Here, we present a two-phase protocol for hydrogen production assay under ambient conditions using Chlamydomonas reinhardtii, which eliminates steps used previously, including centrifugation and resuspension with sulfur-deprived media. We detail steps for Chlamydomonas reinhardtii culture, acetate supply replenishment, anaerobic induction, and H₂ quantification. This protocol enables large-scale experiments in an easy and cost-effective method while maintaining cells vital, crucial factors for transition to industrial scales. For complete details on the use and execution of this protocol, please refer to Elman et al. (2022).