Biological production of hydrogen has been investigated over the past 30 years with the ultimate goal of providing a clean, carbon-neutral fuel. However, based on an extensive literature search and the recommendations of several recent DOE- and DOD-sponsored expert review panels it is obvious that an important element of this research has been largely overlooked - the physiology and diversity of naturally occurring, H₂-producing bacteria. The main objective of this project is to develop a technique to extensively screen nitrogen fixing bacteria isolated from unique environments suspected of H₂ production. In the present work, a fairly new detection method is examined (Katsuda et al. 2006), in which a H₂ color indicator was prepared. This color indicator consists of Wilkinson’s catalyst and several redox sensitive dyes: methylene blue, crystal violet, neutral red, and methyl orange. Tris rhodium chloride, commonly known as Wilkinson’s catalyst was sulfonated to make it water soluble and was subsequently added to vials containing growth media and one redox dye. H₂ (100%, 20%, 5%, 2%, 1%, and 0.5%) was added to each vial, while negative controls were flushed with argon, and incubated 18 h at room temperature. A change of color was observed in all test solutions containing 5-100% H₂ compared to the negative controls. Methyl orange appears to be most sensitive to hydrogen gas, since its visual change of color was detected even with 1% H_2. In addition, spectrophotometric scans (visible spectrum) of each dye/H₂ combination were conducted. Currently, we are adapting the method as a high-throughput technique to screen bacterial H₂ production in 96-well plates.